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No commercial re-use. See rights and permissions. Published by BMJ." /> <meta name="DC.AccessRights" content="restricted" /> <meta name="DC.Relation" content="10.1136/gutjnl-2023-331857" /> <meta name="DC.Relation" content="10.1136/gutjnl-2023-331857" /> <meta name="DC.Description" content="Objective Whether and how the PI3K-AKT pathway, a central node of metabolic homeostasis, is responsible for high-fat-induced non-alcoholic steatohepatitis (NASH) and hepatocellular carcinoma (HCC) remain a mystery. Characterisation of AKT regulation in this setting will provide new strategies to combat HCC. Design Metabolite library screening disclosed that palmitic acid (PA) could activate AKT. In vivo and in vitro palmitoylation assay were employed to detect AKT palmitoylation. Diverse cell and mouse models, including generation of AKT1C77S and AKT1C224S knock-in cells, Zdhhc17 and Zdhhc24 knockout mice and Akt1C224S knock-in mice were employed. Human liver tissues from patients with NASH and HCC, hydrodynamic transfection mouse model, high-fat/high-cholesterol diet (HFHCD)-induced NASH/HCC mouse model and high-fat and methionine/choline-deficient diet (HFMCD)-induced NASH mouse model were also further explored for our mechanism studies. Results By screening a metabolite library, PA has been defined to activate AKT by promoting its palmitoyl modification, an essential step for growth factor-induced AKT activation. Biologically, a high-fat diet could promote AKT kinase activity, thereby promoting NASH and liver cancer. Mechanistically, palmitoyl binding anchors AKT to the cell membrane in a PIP3-independent manner, in part by preventing AKT from assembling into an inactive polymer. The palmitoyltransferases ZDHHC17/24 were characterised to palmitoylate AKT to exert oncogenic effects. Interestingly, the anti-obesity drug orlistat or specific penetrating peptides can effectively attenuate AKT palmitoylation and activation by restricting PA synthesis or repressing AKT modification, respectively, thereby antagonising liver tumorigenesis. Conclusions Our findings elucidate a novel fine-tuned regulation of AKT by PA-ZDHHC17/24-mediated palmitoylation, and highlight tumour therapeutic strategies by taking PA-restricted diets, limiting PA synthesis, or directly targeting AKT palmitoylation. Data are available upon reasonable request." /> <meta name="DC.Contributor" content="Lang Bu" /> <meta name="DC.Contributor" content="Zhengkun Zhang" /> <meta name="DC.Contributor" content="Jianwen Chen" /> <meta name="DC.Contributor" content="Yizeng Fan" /> <meta name="DC.Contributor" content="Jinhe Guo" /> <meta name="DC.Contributor" content="Yaqing Su" /> <meta name="DC.Contributor" content="Huan Wang" /> <meta name="DC.Contributor" content="Xiaomei Zhang" /> <meta name="DC.Contributor" content="Xueji Wu" /> <meta name="DC.Contributor" content="Qiwei Jiang" /> <meta name="DC.Contributor" content="Bing Gao" /> <meta name="DC.Contributor" content="Lei Wang" /> <meta name="DC.Contributor" content="Kunpeng Hu" /> <meta name="DC.Contributor" content="Xiang Zhang" /> <meta name="DC.Contributor" content="Wei Xie" /> <meta name="DC.Contributor" content="Wenyi Wei" /> <meta name="DC.Contributor" content="Ming Kuang" /> <meta name="DC.Contributor" content="Jianping Guo" /> <meta name="article:published_time" content="2024-07-01" /> <meta name="article:section" content="Hepatology" /> <meta name="citation_title" content="High-fat diet promotes liver tumorigenesis via palmitoylation and activation of AKT" /> <meta name="citation_abstract" lang="en" content="<h3>Objective</h3> <p>Whether and how the PI3K-AKT pathway, a central node of metabolic homeostasis, is responsible for high-fat-induced non-alcoholic steatohepatitis (NASH) and hepatocellular carcinoma (HCC) remain a mystery. Characterisation of AKT regulation in this setting will provide new strategies to combat HCC.</p><h3>Design</h3> <p>Metabolite library screening disclosed that palmitic acid (PA) could activate AKT. In vivo and in vitro palmitoylation assay were employed to detect AKT palmitoylation. Diverse cell and mouse models, including generation of <i>AKT1<sup>C77S</sup></i> and <i>AKT1<sup>C224S</sup></i> knock-in cells, <i>Zdhhc17</i> and <i>Zdhhc24</i> knockout mice and <i>Akt1<sup>C224S</sup></i> knock-in mice were employed. Human liver tissues from patients with NASH and HCC, hydrodynamic transfection mouse model, high-fat/high-cholesterol diet (HFHCD)-induced NASH/HCC mouse model and high-fat and methionine/choline-deficient diet (HFMCD)-induced NASH mouse model were also further explored for our mechanism studies.</p><h3>Results</h3> <p>By screening a metabolite library, PA has been defined to activate AKT by promoting its palmitoyl modification, an essential step for growth factor-induced AKT activation. Biologically, a high-fat diet could promote AKT kinase activity, thereby promoting NASH and liver cancer. Mechanistically, palmitoyl binding anchors AKT to the cell membrane in a PIP3-independent manner, in part by preventing AKT from assembling into an inactive polymer. The palmitoyltransferases ZDHHC17/24 were characterised to palmitoylate AKT to exert oncogenic effects. Interestingly, the anti-obesity drug orlistat or specific penetrating peptides can effectively attenuate AKT palmitoylation and activation by restricting PA synthesis or repressing AKT modification, respectively, thereby antagonising liver tumorigenesis.</p><h3>Conclusions</h3> <p>Our findings elucidate a novel fine-tuned regulation of AKT by PA-ZDHHC17/24-mediated palmitoylation, and highlight tumour therapeutic strategies by taking PA-restricted diets, limiting PA synthesis, or directly targeting AKT palmitoylation.</p>" /> <meta name="citation_journal_title" content="Gut" /> <meta name="citation_publisher" content="BMJ Publishing Group" /> <meta name="citation_publication_date" content="2024/07/01" /> <meta name="citation_mjid" content="gutjnl;73/7/1156" /> <meta name="citation_id" content="73/7/1156" /> <meta name="citation_public_url" content="https://gut.bmj.com/content/73/7/1156" /> <meta name="citation_abstract_html_url" content="https://gut.bmj.com/content/73/7/1156.abstract" 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the First Affiliated Hospital, Sun Yat-sen University" /> <meta name="citation_author" content="Zhengkun Zhang" /> <meta name="citation_author_institution" content="Institute of Precision Medicine, the First Affiliated Hospital, Sun Yat-sen University" /> <meta name="citation_author" content="Jianwen Chen" /> <meta name="citation_author_institution" content="Institute of Precision Medicine, the First Affiliated Hospital, Sun Yat-sen University" /> <meta name="citation_author" content="Yizeng Fan" /> <meta name="citation_author_institution" content="Department of Pathology, Beth Israel Deaconess Medical Center, Harvard Medical School" /> <meta name="citation_author" content="Jinhe Guo" /> <meta name="citation_author_institution" content="Institute of Precision Medicine, the First Affiliated Hospital, Sun Yat-sen University" /> <meta name="citation_author" content="Yaqing Su" /> <meta name="citation_author_institution" content="Institute of Precision Medicine, the First Affiliated Hospital, Sun Yat-sen University" /> <meta name="citation_author" content="Huan Wang" /> <meta name="citation_author_institution" content="Institute of Precision Medicine, the First Affiliated Hospital, Sun Yat-sen University" /> <meta name="citation_author" content="Xiaomei Zhang" /> <meta name="citation_author_institution" content="Institute of Precision Medicine, the First Affiliated Hospital, Sun Yat-sen University" /> <meta name="citation_author" content="Xueji Wu" /> <meta name="citation_author_institution" content="Institute of Precision Medicine, the First Affiliated Hospital, Sun Yat-sen University" /> <meta name="citation_author" content="Qiwei Jiang" /> <meta name="citation_author_institution" content="Institute of Precision Medicine, the First Affiliated Hospital, Sun Yat-sen University" /> <meta name="citation_author" content="Bing Gao" /> <meta name="citation_author_institution" content="Institute of Precision Medicine, the First Affiliated Hospital, Sun Yat-sen University" /> <meta name="citation_author" content="Lei Wang" /> <meta name="citation_author_institution" content="Institute of Precision Medicine, the First Affiliated Hospital, Sun Yat-sen University" /> <meta name="citation_author" content="Kunpeng Hu" /> <meta name="citation_author_institution" content="Division of General Surgery, the Third Affiliated Hospital of Sun Yat-Sen University" /> <meta name="citation_author" content="Xiang Zhang" /> <meta name="citation_author_institution" content="State Key Laboratory of Digestive Disease, Institute of Digestive Disease and the Department of Medicine and Therapeutics, Li Ka Shing Institute of Health Sciences, the Chinese University of Hong Kong" /> <meta name="citation_author" content="Wei Xie" /> <meta name="citation_author_institution" content="Institute of Precision Medicine, the First Affiliated Hospital, Sun Yat-sen University" /> <meta name="citation_author" content="Wenyi Wei" /> <meta name="citation_author_institution" 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Shah;citation_author=NM. Iyengar;citation_title=Plant-based and ketogenic diets as diverging paths to address cancer: a review;citation_pages=1201-8;citation_volume=8;citation_year=2022" /> <meta name="citation_reference" content="citation_journal_title=Nature;citation_author=G. Pascual;citation_author=D. Domínguez;citation_author=M. Elosúa-Bayes;citation_title=Dietary palmitic acid promotes a prometastatic memory via schwann cells;citation_pages=485-90;citation_volume=599;citation_year=2021" /> <meta name="citation_reference" content="citation_journal_title=Gastroenterology;citation_author=J. Yang;citation_author=H. Wei;citation_author=Y. Zhou;citation_title=High-fat diet promotes colorectal tumorigenesis through Modulating gut Microbiota and metabolites;citation_pages=135-149;citation_volume=162;citation_year=2022;citation_pmid=http://www.n;citation_doi=10.1053/J.GASTRO.2021.08.041" /> <meta name="citation_reference" content="citation_journal_title=Lancet Diabetes Endocrinol;citation_title=The obesity-cancer link: of increasing concern;citation_pages=175;citation_volume=8;citation_year=2020" /> <meta name="citation_reference" content="citation_journal_title=Nature;citation_author=S. Chen;citation_author=B. Zhu;citation_author=C. Yin;citation_title=Palmitoylation-dependent activation of Mc1R prevents melanomagenesis;citation_pages=399-403;citation_volume=549;citation_year=2017;citation_pmid=28869973;citation_doi=10.1038/nature23887" /> <meta name="citation_reference" content="citation_journal_title=Nature;citation_author=M. Zhang;citation_author=L. Zhou;citation_author=Y. Xu;citation_title=A Stat3 palmitoylation cycle promotes Th17 differentiation and colitis;citation_pages=434-9;citation_volume=586;citation_year=2020;citation_doi=10.1038/s41586-020-2799-2" /> <meta name="citation_reference" content="citation_journal_title=Science;citation_author=O. Rocks;citation_author=A. Peyker;citation_author=M. Kahms;citation_title=An acylation cycle regulates localization and activity of palmitoylated Ras Isoforms;citation_volume=307;citation_year=2005;citation_pmid=15705808;citation_doi=10.1126/science.1105654" /> <meta name="citation_reference" content="citation_journal_title=Nat Rev Gastroenterol Hepatol;citation_author=QM. Anstee;citation_author=HL. Reeves;citation_author=E. Kotsiliti;citation_title=From NASH to HCC: Current concepts and future challenges;citation_pages=411-28;citation_volume=16;citation_year=2019;citation_pmid=31028350;citation_doi=10.1038/s41575-019-0145-7" /> <meta name="citation_reference" content="citation_journal_title=J Hepatol;citation_author=X. Wu;citation_author=KL. Poulsen;citation_author=C. Sanz-Garcia;citation_title=MLKL-dependent signaling regulates autophagic flux in a murine model of non-alcohol-associated fatty liver and steatohepatitis;citation_pages=616-27;citation_volume=73;citation_year=2020;citation_pmid=32220583;citation_doi=10.1016/j.jhep.2020.03.023" /> <meta name="citation_reference" content="citation_journal_title=Nat Rev Gastroenterol Hepatol;citation_author=F. Heymann;citation_author=F. Tacke;citation_title=Immunology in the liver--from homeostasis to disease;citation_pages=88-110;citation_volume=13;citation_year=2016;citation_pmid=26758786;citation_doi=10.1038/nrgastro.2015.200" /> <meta name="citation_reference" content="citation_journal_title=Gastroenterology;citation_author=DF. Calvisi;citation_author=C. Wang;citation_author=C. Ho;citation_title=Increased Lipogenesis, induced by AKT-Mtorc1-Rps6 signaling, promotes development of human hepatocellular carcinoma;citation_pages=1071-83;citation_volume=140;citation_year=2011;citation_pmid=21147110;citation_doi=10.1053/j.gastro.2010.12.006" /> <meta name="citation_reference" content="citation_journal_title=Biochem J;citation_author=B. Vanhaesebroeck;citation_author=DR. Alessi;citation_title=The Pi3K-Pdk1 connection: more than just a road to PKB;citation_pages=561-76;citation_volume=346 Pt 3;citation_year=2000;citation_pmid=http://www.n" /> <meta name="citation_reference" content="citation_journal_title=Cell;citation_author=BD. Manning;citation_author=A. Toker;citation_title=AKT/PKB signaling: navigating the network;citation_pages=381-405;citation_volume=169;citation_year=2017;citation_pmid=28431241;citation_doi=10.1016/j.cell.2017.04.001" /> <meta name="citation_reference" content="citation_journal_title=Nature;citation_author=P. Liu;citation_author=M. Begley;citation_author=W. Michowski;citation_title=Cell-cycle-regulated activation of AKT kinase by phosphorylation at its Carboxyl terminus;citation_pages=541-5;citation_volume=508;citation_year=2014;citation_pmid=24670654;citation_doi=10.1038/nature13079" /> <meta name="citation_reference" content="citation_journal_title=Science;citation_journal_abbrev=Science;citation_author=J. Guo;citation_author=AA. Chakraborty;citation_author=P. Liu;citation_title=pVHL suppresses kinase activity of Akt in a proline-hydroxylation-dependent manner;citation_pages=929-932;citation_volume=353;citation_year=2016;citation_issue=6302;citation_pmid=27563096;citation_doi=10.1126/science.aad5755" /> <meta name="citation_reference" content="citation_journal_title=Science;citation_journal_abbrev=Science;citation_author=L. Stephens;citation_author=K. Anderson;citation_author=D. Stokoe;citation_title=Protein Kinase B Kinases That Mediate Phosphatidylinositol 3,4,5-Trisphosphate-Dependent Activation of Protein Kinase B;citation_pages=710-714;citation_volume=279;citation_year=1998;citation_issue=5351;citation_pmid=9445477;citation_doi=10.1126/science.279.5351.710" /> <meta name="citation_reference" content="citation_journal_title=Experimental cell research;citation_journal_abbrev=Exp Cell Res;citation_author=B. Vanhaesebroeck;citation_author=MD. Waterfield;citation_title=Signaling by distinct classes of phosphoinositide 3-kinases.;citation_pages=239-254;citation_volume=253;citation_year=1999;citation_issue=1;citation_pmid=10579926;citation_doi=10.1006/excr.1999.4701" /> <meta name="citation_reference" content="citation_journal_title=Science;citation_journal_abbrev=Science;citation_author=W-L. Yang;citation_author=J. Wang;citation_author=C-H. Chan;citation_title=The E3 Ligase TRAF6 Regulates Akt Ubiquitination and Activation;citation_pages=1134-1138;citation_volume=325;citation_year=2009;citation_issue=5944;citation_pmid=19713527;citation_doi=10.1126/science.1175065" /> <meta name="citation_reference" content="citation_journal_title=Nat Cell Biol;citation_author=J. Guo;citation_author=X. Dai;citation_author=B. Laurent;citation_title=AKT methylation by Setdb1 promotes AKT kinase activity and Oncogenic functions;citation_pages=226-37;citation_volume=21;citation_year=2019;citation_pmid=30692625;citation_doi=10.1038/s41556-018-0261-6" /> <meta name="citation_reference" content="citation_journal_title=Sci Signal;citation_title=The Deacetylase Sirt1 promotes membrane localization and activation of AKT and Pdk1 during tumorigenesis and cardiac hypertrophy;citation_volume=4;citation_year=2011" /> <meta name="citation_reference" content="citation_journal_title=Nature;citation_author=A. Van Keymeulen;citation_author=MY. Lee;citation_author=M. Ousset;citation_title=Reactivation of multipotency by Oncogenic Pik3Ca induces breast tumour heterogeneity;citation_pages=119-23;citation_volume=525;citation_year=2015;citation_pmid=26266985;citation_doi=10.1038/nature14665" /> <meta name="citation_reference" content="citation_journal_title=Nat Cell Biol;citation_author=S. Mehta;citation_author=Y. Zhang;citation_author=RH. Roth;citation_title=Single-Fluorophore biosensors for sensitive and multiplexed detection of signalling activities;citation_pages=1215-25;citation_volume=20;citation_year=2018;citation_pmid=30250062;citation_doi=10.1038/s41556-018-0200-6" /> <meta name="citation_reference" content="citation_journal_title=Analytical Chemistry (Washington, DC);citation_journal_abbrev=Analytical Chemistry (Washington, DC);citation_author=R. Paulmurugan;citation_author=SS. Gambhir;citation_title=Monitoring protein-protein interactions using split synthetic renilla luciferase protein-fragment-assisted complementation.;citation_pages=1584-1589;citation_volume=75;citation_year=2003;citation_issue=7;citation_pmid=12705589;citation_doi=10.1021/ac020731c" /> <meta name="citation_reference" content="citation_title=AKT (V-AKT murine Thymoma viral Oncogene Homolog 1) and N-Ras (neuroblastoma Ras viral Oncogene Homolog) Coactivation in the Mouse liver promotes rapid carcinogenesis by way of mTOR;citation_pages=833-45;citation_volume=55;citation_year=2012" /> <meta name="citation_reference" content="citation_journal_title=Gut;citation_author=X. Zhang;citation_author=OO. Coker;citation_author=ES. 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Xu;citation_author=M. Xu;citation_author=P. Liu;citation_title=The Mtorc2-Akt1 Cascade is crucial for C-Myc to promote Hepatocarcinogenesis in mice and humans;citation_pages=1600-13;citation_volume=70;citation_year=2019" /> <meta name="citation_reference" content="citation_journal_title=Nat Rev Cancer;citation_author=AJ. Hoy;citation_author=SR. Nagarajan;citation_author=LM. Butler;citation_title=Tumour fatty acid metabolism in the context of therapy resistance and obesity;citation_pages=753-66;citation_volume=21;citation_year=2021;citation_pmid=34417571;citation_doi=10.1038/s41568-021-00388-4" /> <meta name="citation_reference" content="citation_journal_title=Gut;citation_author=L. Che;citation_author=W. Chi;citation_author=Y. Qiao;citation_title=Cholesterol biosynthesis supports the growth of Hepatocarcinoma lesions depleted of fatty acid synthase in mice and humans;citation_volume=69;citation_year=2020;citation_pmid=30954949;citation_doi=10.1136/gutjnl-2018-317581" /> <meta name="citation_fulltext_world_readable" content="" /> <meta name="twitter:title" content="High-fat diet promotes liver tumorigenesis via palmitoylation and activation of AKT" /> <meta name="twitter:card" content="summary_large_image" /> <meta name="twitter:image" content="https://gut.bmj.com/sites/default/files/highwire/gutjnl/73/7.cover-source.jpg" /> <meta name="twitter:description" content="Objective Whether and how the PI3K-AKT pathway, a central node of metabolic homeostasis, is responsible for high-fat-induced non-alcoholic steatohepatitis (NASH) and hepatocellular carcinoma (HCC) remain a mystery. Characterisation of AKT regulation in this setting will provide new strategies to combat HCC. Design Metabolite library screening disclosed that palmitic acid (PA) could activate AKT. In vivo and in vitro palmitoylation assay were employed to detect AKT palmitoylation. Diverse cell and mouse models, including generation of AKT1C77S and AKT1C224S knock-in cells, Zdhhc17 and Zdhhc24 knockout mice and Akt1C224S knock-in mice were employed. Human liver tissues from patients with NASH and HCC, hydrodynamic transfection mouse model, high-fat/high-cholesterol diet (HFHCD)-induced NASH/HCC mouse model and high-fat and methionine/choline-deficient diet (HFMCD)-induced NASH mouse model were also further explored for our mechanism studies. Results By screening a metabolite library, PA has been defined to activate AKT by promoting its palmitoyl modification, an essential step for growth factor-induced AKT activation. Biologically, a high-fat diet could promote AKT kinase activity, thereby promoting NASH and liver cancer. Mechanistically, palmitoyl binding anchors AKT to the cell membrane in a PIP3-independent manner, in part by preventing AKT from assembling into an inactive polymer. The palmitoyltransferases ZDHHC17/24 were characterised to palmitoylate AKT to exert oncogenic effects. Interestingly, the anti-obesity drug orlistat or specific penetrating peptides can effectively attenuate AKT palmitoylation and activation by restricting PA synthesis or repressing AKT modification, respectively, thereby antagonising liver tumorigenesis. Conclusions Our findings elucidate a novel fine-tuned regulation of AKT by PA-ZDHHC17/24-mediated palmitoylation, and highlight tumour therapeutic strategies by taking PA-restricted diets, limiting PA synthesis, or directly targeting AKT palmitoylation. Data are available upon reasonable request." /> <meta name="og-title" property="og:title" content="High-fat diet promotes liver tumorigenesis via palmitoylation and activation of AKT" /> <meta name="og-url" property="og:url" content="https://gut.bmj.com/content/73/7/1156" /> <meta name="og-site-name" property="og:site_name" content="Gut" /> <meta name="og-description" property="og:description" content="Objective Whether and how the PI3K-AKT pathway, a central node of metabolic homeostasis, is responsible for high-fat-induced non-alcoholic steatohepatitis (NASH) and hepatocellular carcinoma (HCC) remain a mystery. Characterisation of AKT regulation in this setting will provide new strategies to combat HCC. Design Metabolite library screening disclosed that palmitic acid (PA) could activate AKT. In vivo and in vitro palmitoylation assay were employed to detect AKT palmitoylation. Diverse cell and mouse models, including generation of AKT1C77S and AKT1C224S knock-in cells, Zdhhc17 and Zdhhc24 knockout mice and Akt1C224S knock-in mice were employed. Human liver tissues from patients with NASH and HCC, hydrodynamic transfection mouse model, high-fat/high-cholesterol diet (HFHCD)-induced NASH/HCC mouse model and high-fat and methionine/choline-deficient diet (HFMCD)-induced NASH mouse model were also further explored for our mechanism studies. Results By screening a metabolite library, PA has been defined to activate AKT by promoting its palmitoyl modification, an essential step for growth factor-induced AKT activation. Biologically, a high-fat diet could promote AKT kinase activity, thereby promoting NASH and liver cancer. Mechanistically, palmitoyl binding anchors AKT to the cell membrane in a PIP3-independent manner, in part by preventing AKT from assembling into an inactive polymer. The palmitoyltransferases ZDHHC17/24 were characterised to palmitoylate AKT to exert oncogenic effects. Interestingly, the anti-obesity drug orlistat or specific penetrating peptides can effectively attenuate AKT palmitoylation and activation by restricting PA synthesis or repressing AKT modification, respectively, thereby antagonising liver tumorigenesis. Conclusions Our findings elucidate a novel fine-tuned regulation of AKT by PA-ZDHHC17/24-mediated palmitoylation, and highlight tumour therapeutic strategies by taking PA-restricted diets, limiting PA synthesis, or directly targeting AKT palmitoylation. Data are available upon reasonable request." /> <meta name="og-type" property="og:type" content="article" /> <meta name="og-image" property="og:image" content="https://gut.bmj.com/sites/default/files/highwire/gutjnl/73/7.cover-source.jpg" /> <link rel="alternate" type="application/vnd.ms-powerpoint" title="Powerpoint" href="/content/73/7/1156.ppt" /> <title>High-fat diet promotes liver tumorigenesis via palmitoylation and activation of AKT | Gut</title> <link type="text/css" rel="stylesheet" href="/sites/default/files/advagg_css/css__SXdRkZmI6sImeXD-jEGdZfrJy7g85hSv3_c89En08-8__BkUQ0yKymMbiRZNLbV7c1Pg0jmMBE1pa-Dap8Tr0rIU__dGRtyN8GJBuhBvXCmXEhVK4mGQl3ZIewHeJUeSvfyQ0.css" media="all" />  <script src="https://cookie-cdn.cookiepro.com/scripttemplates/otSDKStub.js" type="text/javascript" charset="UTF-8" data-domain-script="565e50dd-f6e9-46d6-91cd-5c40a270264f"></script> <script type="text/javascript"> function OnetrustAdsConsent(){ if(googletag.apiReady){ //console.log("GoogleTag API ready") googletag.pubads().disableInitialLoad(); if(OptanonActiveGroups.match(/,4,/)){ googletag.pubads().setRequestNonPersonalizedAds(0); //googletag.pubads().refresh(); } else{ googletag.pubads().setRequestNonPersonalizedAds(1); googletag.pubads().refresh(); } } else{ OnetrustAdsConsent(); } } function OptanonWrapper() { OnetrustAdsConsent(); } </script>  <script type="text/javascript" src="/sites/default/files/advagg_js/js__DdxPgqGimsdUot2QWNz_pGsHzqNGpx6RMCWHpL8YWkg__c6zfK6oHHbLpTal5PdQxeQI3xGoCM8IXWtEHwIZr560__dGRtyN8GJBuhBvXCmXEhVK4mGQl3ZIewHeJUeSvfyQ0.js"></script> <script type="text/javascript" src="https://securepubads.g.doubleclick.net/tag/js/gpt.js"></script> <script type="text/javascript" src="/sites/default/files/advagg_js/js__YFRuiqUdvK1yvVNb1oSBcGhrzBqGejYMOH-ZrKlO98U__AFX4RXDIciSUvyQgiiXaw6-hxp_0ISKrjNzVx1LaIoA__dGRtyN8GJBuhBvXCmXEhVK4mGQl3ZIewHeJUeSvfyQ0.js"></script> <script type="text/javascript" src="https://d1bxh8uas1mnw7.cloudfront.net/assets/embed.js"></script> <script type="text/javascript" src="/sites/default/files/advagg_js/js__RxmsdK7-WlnwoO5NREEVy8MRi0OqO4ZhkOHaMokGNlw__IAyE4mioyYmepJfxC77gDsM0J1xDyesxXH7WjkUNmyc__dGRtyN8GJBuhBvXCmXEhVK4mGQl3ZIewHeJUeSvfyQ0.js"></script> <script type="text/javascript" defer="defer" async="async" src="//cdn.foxycart.com/bmjpg.ecommerce.highwire.org/loader.js"></script> <script type="text/javascript" async="async" src="https://scholar.google.com/scholar_js/casa.js"></script> <script type="text/javascript"> <![CDATA[//><!]]> </script> <script type="text/javascript"> <![CDATA[//><!]]> </script> <script type="text/javascript"> <![CDATA[//><!]]> </script> <script type="text/javascript"> <![CDATA[//><!]]> </script> <script type="text/javascript"> <![CDATA[//><!]]> </script> <script type="text/javascript"> <![CDATA[//><!]]> </script> <script type="text/javascript"> <![CDATA[//><!]]> </script> <script type="text/javascript"> <![CDATA[//><!]]> </script> <script type="text/javascript"> <![CDATA[//><!]]> </script> <script type="text/javascript"> <![CDATA[//><!]]> </script> <script type="text/javascript"> <![CDATA[//><!]]> </script>   <script> let cookies = parseCookies(); let bmj_uuid = cookies.bmj_uuid; let bmj_ppid = cookies.bmj_ppid; window.dataLayer = window.dataLayer || []; const anonMailId = 'e3b0c44298fc1c149afbf4c8996fb92427ae41e4649b934ca495991b7852b855'; function checkUserLoginStatus(dataLayer) { const userMail = dataLayer.find((item) => item.user)?.user?.mail; if (!userMail) { return false; } return userMail !== anonMailId; } let nonAnonMailId = ((dataLayer.find(item => item.user)?.user)?.mail) === anonMailId ? null : (dataLayer.find(item => item.user)?.user)?.mail; let userId = ((dataLayer.find(item => item.user)?.user)?.['data-ics']) ?? 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class="content-block-markup" xmlns:xhtml="http://www.w3.org/1999/xhtml"><div xmlns:xhtml="http://www.w3.org/1999/xhtml" class="contributors"><ol class="contributor-list" id="contrib-group-1"><li class="contributor" id="contrib-1"><span class="name">Lang Bu</span><a id="xref-aff-1-1" class="xref-aff" href="#aff-1">1</a><span class="xref-sep">,</span><a id="xref-aff-2-1" class="xref-aff" href="#aff-2">2</a>, </li><li class="contributor" id="contrib-2"><span class="name">Zhengkun Zhang</span><a id="xref-aff-2-2" class="xref-aff" href="#aff-2">2</a>, </li><li class="contributor" id="contrib-3"><span class="name">Jianwen Chen</span><a id="xref-aff-2-3" class="xref-aff" href="#aff-2">2</a>, </li><li class="contributor" id="contrib-4"><span class="name">Yizeng Fan</span><a id="xref-aff-3-1" class="xref-aff" href="#aff-3">3</a>, </li><li class="contributor" id="contrib-5"><span class="name">Jinhe Guo</span><a id="xref-aff-2-4" class="xref-aff" href="#aff-2">2</a>, </li><li class="contributor" id="contrib-6"><span class="name">Yaqing Su</span><a id="xref-aff-2-5" class="xref-aff" href="#aff-2">2</a>, </li><li class="contributor" id="contrib-7"><span class="name">Huan Wang</span><a id="xref-aff-2-6" class="xref-aff" href="#aff-2">2</a>, </li><li class="contributor" id="contrib-8"><span class="name">Xiaomei Zhang</span><a id="xref-aff-2-7" class="xref-aff" href="#aff-2">2</a>, </li><li class="contributor" id="contrib-9"><span class="name">Xueji Wu</span><a id="xref-aff-2-8" class="xref-aff" href="#aff-2">2</a>, </li><li class="contributor" id="contrib-10"><span class="name">Qiwei Jiang</span><a id="xref-aff-2-9" class="xref-aff" href="#aff-2">2</a>, </li><li class="contributor" id="contrib-11"><span class="name">Bing Gao</span><a id="xref-aff-2-10" class="xref-aff" href="#aff-2">2</a>, </li><li class="contributor" id="contrib-12"><span class="name">Lei Wang</span><a id="xref-aff-2-11" class="xref-aff" href="#aff-2">2</a>, </li><li class="contributor" id="contrib-13"><span class="name">Kunpeng Hu</span><a id="xref-aff-4-1" class="xref-aff" href="#aff-4">4</a>, </li><li class="contributor" id="contrib-14"><span class="name">Xiang Zhang</span><a id="xref-aff-5-1" class="xref-aff" href="#aff-5">5</a>, </li><li class="contributor" id="contrib-15"><span class="name">Wei Xie</span><a id="xref-aff-2-12" class="xref-aff" href="#aff-2">2</a>, </li><li class="contributor" id="contrib-16"><span class="name">Wenyi Wei</span><a id="xref-aff-3-2" class="xref-aff" href="#aff-3">3</a>, </li><li class="contributor" id="contrib-17"><a href="http://orcid.org/0000-0002-7397-5779" class="bmjj-markup-orcid-logo" target="_blank">http://orcid.org/0000-0002-7397-5779</a><span class="name">Ming Kuang</span><a id="xref-aff-1-2" class="xref-aff" href="#aff-1">1</a>, </li><li class="last" id="contrib-18"><a href="http://orcid.org/0000-0002-8158-0101" class="bmjj-markup-orcid-logo" target="_blank">http://orcid.org/0000-0002-8158-0101</a><span class="name">Jianping Guo</span><a id="xref-aff-2-13" class="xref-aff" href="#aff-2">2</a></li></ol><ol class="affiliation-list"><li class="aff"><a id="aff-1" name="aff-1"></a><address> <sup>1</sup> <span class="institution">Center of Hepato-Pancreate-Biliary Surgery, the First Affiliated Hospital, Sun Yat-sen University</span>, <span class="addr-line">Guangzhou, Guangdong</span>, China </address></li><li class="aff"><a id="aff-2" name="aff-2"></a><address> <sup>2</sup> <span class="institution">Institute of Precision Medicine, the First Affiliated Hospital, Sun Yat-sen University</span>, <span class="addr-line">Guangzhou, Guangdong</span>, China </address></li><li class="aff"><a id="aff-3" name="aff-3"></a><address> <sup>3</sup> <span class="institution">Department of Pathology, Beth Israel Deaconess Medical Center, Harvard Medical School</span>, <span class="addr-line">Boston</span>, <span class="addr-line">Massachusetts</span>, USA </address></li><li class="aff"><a id="aff-4" name="aff-4"></a><address> <sup>4</sup> <span class="institution">Division of General Surgery, the Third Affiliated Hospital of Sun Yat-Sen University</span>, <span class="addr-line">Guangzhou, Guangdong</span>, China </address></li><li class="aff"><a id="aff-5" name="aff-5"></a><address> <sup>5</sup> <span class="institution">State Key Laboratory of Digestive Disease, Institute of Digestive Disease and the Department of Medicine and Therapeutics, Li Ka Shing Institute of Health Sciences, the Chinese University of Hong Kong</span>, <span class="addr-line">Hong Kong</span>, China </address></li></ol><ol class="corresp-list"><li class="corresp" id="corresp-1"><span class="corresp-label">Correspondence to</span> Professor Jianping Guo; <span class="em-link"><span class="em-addr">guojp6{at}mail.sysu.edu.cn</span></span>; Professor Ming Kuang; <span class="em-link"><span class="em-addr">kuangm{at}mail.sysu.edu.cn</span></span> </li></ol></div></div></div> </div> </div> <div class="panel-separator"></div><div class="panel-pane pane-highwire-markup abstract-with-bc" > <div class="pane-content"> <div class="highwire-markup"><div xmlns="http://www.w3.org/1999/xhtml" id="content-block" xmlns:xhtml="http://www.w3.org/1999/xhtml"><div class="article abstract-view "><span class="highwire-journal-article-marker-start"></span><div class="section abstract" id="abstract-1"><h2>Abstract</h2><div id="sec-1" class="subsection"><p id="p-1"><strong>Objective</strong> Whether and how the PI3K-AKT pathway, a central node of metabolic homeostasis, is responsible for high-fat-induced non-alcoholic steatohepatitis (NASH) and hepatocellular carcinoma (HCC) remain a mystery. Characterisation of AKT regulation in this setting will provide new strategies to combat HCC.</p></div><div id="sec-2" class="subsection"><p id="p-2"><strong>Design</strong> Metabolite library screening disclosed that palmitic acid (PA) could activate AKT. In vivo and in vitro palmitoylation assay were employed to detect AKT palmitoylation. Diverse cell and mouse models, including generation of <em>AKT1<sup>C77S</sup> </em> and <em>AKT1<sup>C224S</sup> </em> knock-in cells, <em>Zdhhc17</em> and <em>Zdhhc24</em> knockout mice and <em>Akt1<sup>C224S</sup> </em> knock-in mice were employed. Human liver tissues from patients with NASH and HCC, hydrodynamic transfection mouse model, high-fat/high-cholesterol diet (HFHCD)-induced NASH/HCC mouse model and high-fat and methionine/choline-deficient diet (HFMCD)-induced NASH mouse model were also further explored for our mechanism studies.</p></div><div id="sec-3" class="subsection"><p id="p-3"><strong>Results</strong> By screening a metabolite library, PA has been defined to activate AKT by promoting its palmitoyl modification, an essential step for growth factor-induced AKT activation. Biologically, a high-fat diet could promote AKT kinase activity, thereby promoting NASH and liver cancer. Mechanistically, palmitoyl binding anchors AKT to the cell membrane in a PIP3-independent manner, in part by preventing AKT from assembling into an inactive polymer. The palmitoyltransferases ZDHHC17/24 were characterised to palmitoylate AKT to exert oncogenic effects. Interestingly, the anti-obesity drug orlistat or specific penetrating peptides can effectively attenuate AKT palmitoylation and activation by restricting PA synthesis or repressing AKT modification, respectively, thereby antagonising liver tumorigenesis.</p></div><div id="sec-4" class="subsection"><p id="p-4"><strong>Conclusions</strong> Our findings elucidate a novel fine-tuned regulation of AKT by PA-ZDHHC17/24-mediated palmitoylation, and highlight tumour therapeutic strategies by taking PA-restricted diets, limiting PA synthesis, or directly targeting AKT palmitoylation.</p></div></div><ul class="kwd-group"><li class="kwd">CELL SIGNALLING</li><li class="kwd">DRUG DEVELOPMENT</li><li class="kwd">FATTY ACID SUPPLEMENTATION</li><li class="kwd">HEPATOBILIARY CANCER</li><li class="kwd">MOLECULAR ONCOLOGY</li></ul><div class="section data-availability" id="sec-21"><h2 class="">Data availability statement</h2><p id="p-55">Data are available upon reasonable request.</p></div><span class="highwire-journal-article-marker-end"></span></div><span class="related-urls"></span></div></div> </div> </div> <div class="panel-separator"></div><div class="panel-pane pane-dfp-pane oas-ads oas-ads-mid pull-right" > <div class="pane-content"> <div id="dfp-ad-mpu-wrapper" class="dfp-tag-wrapper"> <div id="dfp-ad-mpu" class="dfp-tag-wrapper"> <script type="text/javascript"> googletag.cmd.push(function() { googletag.display("dfp-ad-mpu"); }); </script> </div> </div> </div> </div> <div class="panel-separator"></div><div class="panel-pane pane-custom pane-3" > <div class="pane-content"> <p><a href="https://doi.org/10.1136/gutjnl-2023-330826" target="_new">https://doi.org/10.1136/gutjnl-2023-330826</a></p> </div> </div> <div class="panel-separator"></div><div class="panel-pane pane-highwire-altmetrics" > <h2 class="pane-title">Statistics from Altmetric.com</h2> <div class="pane-content"> <div data-badge-details="right" data-badge-type="medium-donut" data-doi="10.1136/gutjnl-2023-330826" data-hide-no-mentions="true" class="altmetric-embed"></div> </div> </div> <div class="panel-separator"></div><div class="panel-pane pane-bmjj-jumplinks" > <div class="pane-content"> <div class="highwire-list-wrapper"><div class="highwire-list"><ul></ul></div></div> </div> </div> <div class="panel-separator"></div><div class="panel-pane pane-custom pane-4 permissions-box" > <h2 class="pane-title">Request Permissions</h2> <div class="pane-content"> <p>If you wish to reuse any or all of this article please use the link below which will take you to the Copyright Clearance Center’s RightsLink service. You will be able to get a quick price and instant permission to reuse the content in many different ways.</p> </div> </div> <div class="panel-separator"></div><div class="panel-pane pane-panels-ajax-pane-content permissions-box pane-highwire-permission-link" id="request-permissions"> <h2 class="pane-title"><span class='panels-ajax-pane-title' data-pid='new-70fd4c70-563e-4b94-ae3b-2a97f3980c13'></span></h2> <div class="pane-content"> <div class='panels-ajax-pane panels-ajax-pane-new-70fd4c70-563e-4b94-ae3b-2a97f3980c13' data-pid='new-70fd4c70-563e-4b94-ae3b-2a97f3980c13'></div> </div> </div> <div class="panel-separator"></div><div class="panel-pane pane-highwire-markup" > <div class="pane-content"> <div class="highwire-markup"><div xmlns="http://www.w3.org/1999/xhtml" class="content-block-markup" xmlns:xhtml="http://www.w3.org/1999/xhtml"><div class="article fulltext-view "><span class="highwire-journal-article-marker-start"></span><ul class="kwd-group"><li class="kwd"><a href="/keyword/cell-signalling" class="hw-term hw-article-keyword hw-article-keyword-cell-signalling" rel="nofollow">CELL SIGNALLING</a></li><li class="kwd"><a href="/keyword/drug-development" class="hw-term hw-article-keyword hw-article-keyword-drug-development" rel="nofollow">DRUG DEVELOPMENT</a></li><li class="kwd"><a href="/keyword/fatty-acid-supplementation" class="hw-term hw-article-keyword hw-article-keyword-fatty-acid-supplementation" rel="nofollow">FATTY ACID SUPPLEMENTATION</a></li><li class="kwd"><a href="/keyword/hepatobiliary-cancer" class="hw-term hw-article-keyword hw-article-keyword-hepatobiliary-cancer" rel="nofollow">HEPATOBILIARY CANCER</a></li><li class="kwd"><a href="/keyword/molecular-oncology" class="hw-term hw-article-keyword hw-article-keyword-molecular-oncology" rel="nofollow">MOLECULAR ONCOLOGY</a></li></ul><div class="boxed-text" id="boxed-text-1"><div id="sec-5" class="subsection"><h4>WHAT IS ALREADY KNOWN ON THIS TOPIC</h4><ul class="list-unord " id="list-1"><li id="list-item-1"><p id="p-5">AKT plays a leading role in physiological metabolic homeostasis and is closely related to liver metabolic disorders.</p></li><li id="list-item-2"><p id="p-6">High-fat diets play a key role in liver metabolic disorders, the detailed mechanisms of different fatty acids such as palmitic acid (PA) in these diseases remain unclear.</p></li><li id="list-item-3"><p id="p-7">AKT-mTOR activation induces lipogenesis and hepatocellular carcinoma (HCC) progression.</p></li></ul></div><div id="sec-6" class="subsection"><h4>WHAT THIS STUDY ADDS</h4><ul class="list-unord " id="list-2"><li id="list-item-4"><p id="p-8">AKT undergoes palmitoylation modification, which would contribute to high-fat diets (PA uptake)-induced AKT activation in cells, mice and human liver tissues (NASH and HCC).</p></li><li id="list-item-5"><p id="p-9">The palmitoyltransferases ZDHHC17/24 palmitoylate AKT to exert pathological effects in NASH formation and liver tumorigenesis.</p></li><li id="list-item-6"><p id="p-10">The anti-obesity drug orlistat or specific penetrating peptides can effectively attenuate AKT palmitoylation and activation, thereby antagonising liver tumorigenesis.</p></li></ul></div><div id="sec-7" class="subsection"><h4>HOW THIS STUDY MIGHT AFFECT RESEARCH, PRACTICE OR POLICY</h4><ul class="list-unord " id="list-3"><li id="list-item-7"><p id="p-11">This study encloses a fine-tuned regulation of AKT activity by PA-ZDHHC-mediated palmitoylation, providing the strategies for HCC therapy by limiting PA uptake, restricting PA synthesis, and targeting ZDHHC17/24.</p></li></ul></div></div><div class="section" id="sec-8"><h2 class="">One sentence statement</h2><p id="p-12">Adoption of a low-fat (palmitic acid (PA)-restricted) diet, restriction of PA synthesis, or direct targeting of ZDHHC17/24 would provide effective strategies for hepatocellular carcinoma (HCC) therapy by attenuating AKT palmitoylation and activation.</p></div><div class="section intro" id="sec-9"><h2 class="">Introduction</h2><p id="p-13">Diet not only provides nutrition and building blocks for the uncontrolled growth of tumour cells, but also promotes signal transduction to affect various aspects of tumours, so diet-changing therapies have recently been realised, including calorie restriction and low-calorie-fat diet.<a id="xref-ref-1-1" class="xref-bibr" href="#ref-1">1</a> It is worth noting that high-fat diets (HFDs) are primarily associated with metabolic diseases, including obesity and cancer, through diverse and unclear mechanisms.<a id="xref-ref-2-1" class="xref-bibr" href="#ref-2">2 3</a> Among them, palmitic oil is one of the most source of dietary oil and major cause of obesity, which has been considered as a metabolic disease and preferred to inducing tumours.<a id="xref-ref-4-1" class="xref-bibr" href="#ref-4">4</a> Of note, PA as a major component of palmitic and animal fat oil has been tightly linked to oral tumour metastasis with its metabolic function to shape cell plasma membrane.<a id="xref-ref-2-2" class="xref-bibr" href="#ref-2">2</a> Meanwhile, an active product of PA, palmitoyl-CoA, has been shown to induce palmitoyl modification in a variety of proteins, such as MC1R in response to UV-induced melanoma,<a id="xref-ref-5-1" class="xref-bibr" href="#ref-5">5</a> STAT3 in response to IL6-induced T<sub>h17</sub> differentiation and antiviral infection,<a id="xref-ref-6-1" class="xref-bibr" href="#ref-6">6</a> and N-RAS for colon cancers.<a id="xref-ref-7-1" class="xref-bibr" href="#ref-7">7</a> However, whether and how PA accumulation from synthesis or food uptake is involved in tumorigenesis, especially in organs involved in metabolic homeostasis, such as liver, is unclear.</p><p id="p-14">HFD has been considered as an unabated risk factor of non-alcoholic steatohepatitis (NASH), an advanced form of non-alcoholic fatty liver disease (NAFLD), now has also been termed as metabolic dysfunction-associated steatohepatitis. NASH involves lobular inﬂammation, ﬁbrosis and hepatocyte ballooning, which can progress to HCC.<a id="xref-ref-8-1" class="xref-bibr" href="#ref-8">8</a> Although multiple potential mechanisms such as autophagy and immune responses are associated with NASH,<a id="xref-ref-9-1" class="xref-bibr" href="#ref-9">9 10</a> whether and how the PI3K-AKT pathway, a central node of metabolic homeostasis and playing potent role in lipogenesis and HCC progression,<a id="xref-ref-11-1" class="xref-bibr" href="#ref-11">11</a> is responsible for high-fat-induced NASH and HCC remain a mystery.</p><p id="p-15">As a central signal in response to extracellular stimuli, the PI3K-AKT pathway plays pivotal roles in cell growth, survival and metabolic homeostasis physiologically, as well as in diabetes and tumorigenesis pathologically.<a id="xref-ref-12-1" class="xref-bibr" href="#ref-12">12 13</a> Therefore, the regulation of AKT, especially at the post-translational level, has been extensively established.<a id="xref-ref-14-1" class="xref-bibr" href="#ref-14">14 15</a> Notably, the binding of AKT PH domain to PI3K-generated PIP<sub>3</sub> is thought to be a critical step in the full activation of AKT by PDK1-mediated T308 and mTORC2-mediated S473 phosphorylation.<a id="xref-ref-16-1" class="xref-bibr" href="#ref-16">16 17</a> For example, ubiquitination, acetylation and methylation all contribute to the binding of AKT to PIP<sub>3</sub> for its further activation.<a id="xref-ref-18-1" class="xref-bibr" href="#ref-18">18–20</a> Thus, genetic alterations of PIP<sub>3</sub>-associated enzymes, including its kinase <em>PIK3CA</em>, phosphatase <em>PTEN</em>, and their upstream regulatory proteins, such as <em>RAS</em> and <em>EGFR</em>, have been observed in around 50% of diverse types of cancer and largely contribute to PIP<sub>3</sub> accumulation and subsequent AKT activation.<a id="xref-ref-13-1" class="xref-bibr" href="#ref-13">13</a> To this end, specific inhibitors against the <em>PIK3CA-H1047R</em> mutant have been approved for breast cancer intervention.<a id="xref-ref-21-1" class="xref-bibr" href="#ref-21">21</a> However, besides canonical PI3K-PIP<sub>3</sub> dependent AKT activation, whether and how AKT activation in a PIP<sub>3</sub>-independent manner remain unclear. In this study, we report that accumulating PA from HFDs or aberrant synthesis promotes ZDHHC17/24-catalysing palmitoylation and activation of AKT in a PIP<sub>3</sub>-independent manner, leading to NASH and liver tumorigenesis, and highlight the strategy to target PA-ZDHHC-AKT axis for combating HCC.</p></div><div class="section results" id="sec-10"><h2 class="">Results</h2><div id="sec-11" class="subsection"><h3>HFD activates AKT and promotes NASH and liver tumorigenesis</h3><p id="p-16">To systematically explore the regulatory role of metabolites on AKT kinase, we established an in vitro AKT activity screening approach based on and optimised from previous reports.<a id="xref-ref-22-1" class="xref-bibr" href="#ref-22">22 23</a> Briefly, using AKT recognised peptide (PRPRSCTWPDPRPEF) as a substrate, phosphorylation of that can be recognised by a specific FHA1 domain, leading to the assembly of a functional Renilla luciferase (<a id="xref-fig-1-1" class="xref-fig" href="#F1">figure 1A</a>). This reporter system was initially validated with an insulin-based positive control (<a id="xref-supplementary-material-1-1" class="xref-supplementary-material" href="#DC1">online supplemental figure S1A</a>), and then challenged with a metabolite library (<a id="xref-fig-1-2" class="xref-fig" href="#F1">figure 1B</a>). Among them, several metabolites were identified and validated to positively activate AKT, including equol, menadione, PA, propylparaben, xanthosine dihydrate, isohomovanillic acid, and 5'-adenylic acid (<a id="xref-fig-1-3" class="xref-fig" href="#F1">figure 1B–C</a> and <a id="xref-supplementary-material-1-2" class="xref-supplementary-material" href="#DC1">online supplemental figure S1B</a>). Notably, PA was shown on be the top-ranked metabolites for activating AKT (<a id="xref-fig-1-4" class="xref-fig" href="#F1">figure 1C</a> and <a id="xref-supplementary-material-1-3" class="xref-supplementary-material" href="#DC1">online supplemental figure S1B</a>), which was effectively antagonised by the palmitoyl-transferase inhibitor 2-bromopalmitate (2BP) (<a id="xref-supplementary-material-1-4" class="xref-supplementary-material" href="#DC1">online supplemental figure S1C</a>). Consistent with these findings, PA promoted, whereas 2BP attenuated AKT kinase activity (<a id="xref-supplementary-material-1-5" class="xref-supplementary-material" href="#DC1">online supplemental figure S1D-G</a>), its interaction with PDK1 and PIP<sub>3</sub> (<a id="xref-supplementary-material-1-6" class="xref-supplementary-material" href="#DC1">online supplemental figure S1H-I</a>), and its plasma membrane translocation (<a id="xref-supplementary-material-1-7" class="xref-supplementary-material" href="#DC1">online supplemental figure S1J</a>). RNA-sequencing results also demonstrated that PA could strongly enhance the activity of PI3K-AKT pathway (<a id="xref-supplementary-material-1-8" class="xref-supplementary-material" href="#DC1">online supplemental figure S1K-N</a>), thus, collectively suggesting PA as a potent activator of AKT.</p><div id="DC1" class="supplementary-material"><h3 class="">Supplemental material</h3><div> <span class="inline-linked-media-wrapper"><span id="DC1" class="inline-linked-media"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/DC1/embed/inline-supplementary-material-1.pdf?download=true" class="" data-icon-position="" data-hide-link-title="0"><span class="icon-download-alt"></span>[gutjnl-2023-330826supp001.pdf]</a></span></span> </div></div><div id="F1" class="fig pos-float odd"><div class="highwire-figure"><div class="fig-inline-img-wrapper"><div class="fig-inline-img"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F1.large.jpg?width=800&height=600&carousel=1" title="Palmitic acid (PA) activates AKT and promotes liver tumorigenesis. (A) Mode diagram of AKT activity detection system. (B) HEK293 cells were transfected with AKT-Sub-TK-FHA1 plasmid, serum-starved for 10 hours, then treated with more than 500 metabolites for 8 hours and subjected to luciferase assay. (C) HEK293 cells were transfected with AKT-Sub-TK-FHA1, serum-starved for 10 hours, then treated with screened metabolites for 8 hours and subjected to luciferase assay. Data are presented as the mean±SD of three independent experiments. **P" class="highwire-fragment fragment-images colorbox-load" rel="gallery-fragment-images-183578412" data-figure-caption="<div class="highwire-markup">Palmitic acid (PA) activates AKT and promotes liver tumorigenesis. (A) Mode diagram of AKT activity detection system. (B) HEK293 cells were transfected with AKT-Sub-TK-FHA1 plasmid, serum-starved for 10 hours, then treated with more than 500 metabolites for 8 hours and subjected to luciferase assay. (C) HEK293 cells were transfected with AKT-Sub-TK-FHA1, serum-starved for 10 hours, then treated with screened metabolites for 8 hours and subjected to luciferase assay. Data are presented as the mean±SD of three independent experiments. **P<0.01, ***p<0.001, ****p<0.0001 (Student’s t test). (D) Different hepatocarcinogenesis mouse models and the degree of tumorigenic capacity. (E) Representative image of livers harvested from mice which were hydrodynamic transfected with N‐RasV12, SB transposase, and AKT1 or myr-mAkt1 plasmids, then intraperitoneally injected or intragastric administration with or without PA (10 mg/kg) every day and fed on a normal diet (ND), or a high-fat diet (HFD) for 1 or 2 months. (F) The percentage of liver/body weight was shown. Data are presented as the mean±SD (n=6 mice per group). ***P<0.001, ****p<0.0001 (Student’s t test). (G) Liver sections were stained with H&E and analysed by immunohistochemistry (IHC) for pS6 (pS240/244), AFP, Ki67, and Cleaved-Caspase3 levels. (H) Liver tissues from above group were subjected to immunoblotting (IB) analysis. (I) Liver sections of C57BL/6 mice with HFHC diet for different months were stained with H&E, Masson, and analysed by IHC for pS6, Pan-CK, and CPS1 levels. (J) Liver tissues from above groups were subjected to IB analysis.</div>" data-icon-position="" data-hide-link-title="0"><span class="hw-responsive-img"><img class="highwire-fragment fragment-image lazyload" alt="Figure 1" src="data:image/gif;base64,R0lGODlhAQABAIAAAAAAAP///yH5BAEAAAAALAAAAAABAAEAAAIBRAA7" data-src="https://gut.bmj.com/content/gutjnl/73/7/1156/F1.medium.gif" width="405" height="440"/><noscript><img class="highwire-fragment fragment-image" alt="Figure 1" src="https://gut.bmj.com/content/gutjnl/73/7/1156/F1.medium.gif" width="405" height="440"/></noscript></span></a></div></div><ul class="highwire-figure-links inline"><li class="download-fig first"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F1.large.jpg?download=true" class="highwire-figure-link highwire-figure-link-download" title="Download Figure 1" data-icon-position="" data-hide-link-title="0">Download figure</a></li> <li class="new-tab"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F1.large.jpg" class="highwire-figure-link highwire-figure-link-newtab" target="_blank" data-icon-position="" data-hide-link-title="0">Open in new tab</a></li> <li class="download-ppt last"><a href="/highwire/powerpoint/261214" class="highwire-figure-link highwire-figure-link-ppt" data-icon-position="" data-hide-link-title="0">Download powerpoint</a></li> </ul></div><div class="fig-caption" xmlns:xhtml="http://www.w3.org/1999/xhtml"><span class="fig-label">Figure 1</span> <p id="p-18" class="first-child">Palmitic acid (PA) activates AKT and promotes liver tumorigenesis. (A) Mode diagram of AKT activity detection system. (B) HEK293 cells were transfected with AKT-Sub-TK-FHA1 plasmid, serum-starved for 10 hours, then treated with more than 500 metabolites for 8 hours and subjected to luciferase assay. (C) HEK293 cells were transfected with AKT-Sub-TK-FHA1, serum-starved for 10 hours, then treated with screened metabolites for 8 hours and subjected to luciferase assay. Data are presented as the mean±SD of three independent experiments. **P<0.01, ***p<0.001, ****p<0.0001 (Student’s t test). (D) Different hepatocarcinogenesis mouse models and the degree of tumorigenic capacity. (E) Representative image of livers harvested from mice which were hydrodynamic transfected with N‐RasV12, SB transposase, and AKT1 or myr-mAkt1 plasmids, then intraperitoneally injected or intragastric administration with or without PA (10 mg/kg) every day and fed on a normal diet (ND), or a high-fat diet (HFD) for 1 or 2 months. (F) The percentage of liver/body weight was shown. Data are presented as the mean±SD (n=6 mice per group). ***P<0.001, ****p<0.0001 (Student’s t test). (G) Liver sections were stained with H&E and analysed by immunohistochemistry (IHC) for pS6 (pS240/244), AFP, Ki67, and Cleaved-Caspase3 levels. (H) Liver tissues from above group were subjected to immunoblotting (IB) analysis. (I) Liver sections of C57BL/6 mice with HFHC diet for different months were stained with H&E, Masson, and analysed by IHC for pS6, Pan-CK, and CPS1 levels. (J) Liver tissues from above groups were subjected to IB analysis.</p><div class="sb-div caption-clear"></div></div></div><p id="p-19">Despite recent reports that PA promotes metastasis of oral cancer in mice,<a id="xref-ref-2-3" class="xref-bibr" href="#ref-2">2</a> we sought to determine whether PA promotes tumorigenesis through activation of AKT. To this end, a well-established tail-vein hydrodynamic transfection liver tumour mouse model (transfection with hyperactive mouse Akt1 (myr-mAkt1), mutated N-Ras (N-Ras<sup>V12</sup>) and Sleeping Beauty transposon) was adopted and further optimised. Consistent with previous findings, cocktail constructs containing myr-mAkt1, but not wild-type (WT) AKT1 could promote liver tumorigenesis (<a id="xref-fig-1-5" class="xref-fig" href="#F1">figure 1D</a>).<a id="xref-ref-11-2" class="xref-bibr" href="#ref-11">11 24</a> Interestingly, in our optimised model, HFD or administration of PA (I.G. or I.P.), both significantly promoted WT-AKT1-induced NASH (<a id="xref-fig-1-6" class="xref-fig" href="#F1">figure 1E–F</a>), coupled with increased AKT activity and hepatocyte proliferation (<a id="xref-fig-1-7" class="xref-fig" href="#F1">figure 1G–H</a>). Consistent with these findings, we obtained liver tissues from a reported HFLC-feeding induced NAFLD/NASH/HCC murine model,<a id="xref-ref-25-1" class="xref-bibr" href="#ref-25">25</a> and observed that AKT activity was increasingly elevated on progression of NASH to HCC (<a id="xref-fig-1-8" class="xref-fig" href="#F1">figure 1I–J</a>). These results suggest that HFD or PA administration could activate AKT, thereby potentially promoting NASH formation and liver tumorigenesis.</p></div><div id="sec-12" class="subsection"><h3>AKT undergoes palmitoyl-modification</h3><p id="p-20">Although PA has been shown to factilitate PI3K-AKT signalling,<a id="xref-ref-26-1" class="xref-bibr" href="#ref-26">26</a> we observed that 2BP, which has the function of antagonising palmitoyltransferase, readily inhibited PA-induced AKT activation (<a id="xref-supplementary-material-1-9" class="xref-supplementary-material" href="#DC1">online supplemental figure S1C-J</a>). Therefore, we hypothesised that AKT undergoes palmitoyl-modification. To this end, two palmitoyl-conjugation assays were employed, the click chemistry reaction to biotin-azide assay and acyl-biotin exchange (ABE) palmitoylation assay. Both ectopically expressed and endogenous AKT1 were proved to undergo palmitoyl-modification (<a id="xref-fig-2-1" class="xref-fig" href="#F2">figure 2A–B</a> and <a id="xref-supplementary-material-1-10" class="xref-supplementary-material" href="#DC1">online supplemental figure S2A-D</a>). Similar results were observed in AKT2 and AKT3 subtypes (<a id="xref-supplementary-material-1-11" class="xref-supplementary-material" href="#DC1">online supplemental figure S2E</a>). Importantly, AKT1 palmitoylation was significantly induced by PA stimulation, which was antagonised by 2BP (<a id="xref-fig-2-2" class="xref-fig" href="#F2">figure 2C</a> and <a id="xref-supplementary-material-1-12" class="xref-supplementary-material" href="#DC1">online supplemental figure S2F</a>). In contrast, challenges with insulin, PI3K inhibitors, or enforced AKT membrane translocation (myr-AKT1) had mild effects on AKT palmitoyl-conjugation (<a id="xref-supplementary-material-1-13" class="xref-supplementary-material" href="#DC1">online supplemental figure S2G-J</a>).</p><div id="F2" class="fig pos-float odd"><div class="highwire-figure"><div class="fig-inline-img-wrapper"><div class="fig-inline-img"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F2.large.jpg?width=800&height=600&carousel=1" title="AKT1 is palmitoylated at C77/C224 to enhance its kinase activity and oncogenic function. (A) Huh7 cells were labelled with dimethyl sulfoxide (DMSO) or alk-C16 (50 µM) for 4 hours, followed by click chemistry reaction to biotin-azide assay. (B) Whole cell lysates of Huh7 cells were incubated with α-AKT1 antibody and processed for acyl-biotin exchange (ABE) palmitoylation assay. (C) HepG2 cells were treated with palmitic acid (PA) (100 µM) with or without 2BP (50 µM) for 4 hours, incubated with α-AKT1 antibody and processed for ABE palmitoylation assay. (D) Mouse liver tissues obtained from figure 1E were subjected for ABE palmitoylation assay. (E) Four non-alcoholic steatohepatitis (NASH)-induced hepatocellular carcinoma (HCC) with their adjacent normal tissues were subjected for ABE palmitoylation assay. N, indicates adjacent normal tissue; T, indicates liver tumour; Mock, indicates no hydroxylamine (HAM) treatment. (F) Mouse liver tissues obtained from figure 1I were subjected for ABE palmitoylation assay. (G) Human liver sections with no steatosis, NASH and HCC were stained with H&E, Masson, and analysed by IHC for pS6 levels. (H) Liver tissues from above groups were subjected to ABE palmitoylation assay, and the relative palmitoylation of AKT was quantified with total AKT1 and normalised with no steatosis tissues. Data are presented as the mean±SD (n=7 for no steatosis tissues, n=8 for NASH, n=8 for HCC). **P" class="highwire-fragment fragment-images colorbox-load" rel="gallery-fragment-images-183578412" data-figure-caption="<div class="highwire-markup"><div xmlns="http://www.w3.org/1999/xhtml">AKT1 is palmitoylated at C77/C224 to enhance its kinase activity and oncogenic function. (A) Huh7 cells were labelled with dimethyl sulfoxide (DMSO) or alk-C16 (50 µM) for 4 hours, followed by click chemistry reaction to biotin-azide assay. (B) Whole cell lysates of Huh7 cells were incubated with α-AKT1 antibody and processed for acyl-biotin exchange (ABE) palmitoylation assay. (C) HepG2 cells were treated with palmitic acid (PA) (100 µM) with or without 2BP (50 µM) for 4 hours, incubated with α-AKT1 antibody and processed for ABE palmitoylation assay. (D) Mouse liver tissues obtained from figure 1E were subjected for ABE palmitoylation assay. (E) Four non-alcoholic steatohepatitis (NASH)-induced hepatocellular carcinoma (HCC) with their adjacent normal tissues were subjected for ABE palmitoylation assay. N, indicates adjacent normal tissue; T, indicates liver tumour; Mock, indicates no hydroxylamine (HAM) treatment. (F) Mouse liver tissues obtained from figure 1I were subjected for ABE palmitoylation assay. (G) Human liver sections with no steatosis, NASH and HCC were stained with H&E, Masson, and analysed by IHC for pS6 levels. (H) Liver tissues from above groups were subjected to ABE palmitoylation assay, and the relative palmitoylation of AKT was quantified with total AKT1 and normalised with no steatosis tissues. Data are presented as the mean±SD (n=7 for no steatosis tissues, n=8 for NASH, n=8 for HCC). **P<0.01, ***p<0.001, ****p<0.0001 (Student’s t test). (I) <em>AKT1<sup>WT</sup> </em>, <em>AKT1<sup>C77S</sup> </em> and <em>AKT1<sup>C224S</sup> </em> HEK293 cells were treated with or without PA for 4 hours, followed by ABE palmitoylation assay. (J) <em>AKT1<sup>WT</sup> </em>, <em>AKT1<sup>C77S</sup> </em> and <em>AKT1<sup>C224S</sup> </em> HEK293 cells were treated with or without PA and alk-C16 for 4 hours, subjected to click chemistry reaction to biotin-azide assay. (K) DLD1-<em>AKT1/2<sup>-/-</sup>-EV,</em> DLD1-<em>AKT1/2<sup>-/-</sup>-Flag-AKT1-WT</em>, <em>C77S</em>, <em>C224S</em>, and <em>C77/224S</em> cells were subjected to colony formation (I) and soft agar assays (J). Data are presented as the mean±SD of three independent experiments. **P<0.01, ***p<0.001, ****p<0.0001 (Student’s t test). (L) Representative image of livers harvested from hydrodynamic transfection of EV, myr-mAkt1, or myr-mAkt1-C77/224S and N‐RasV12, SB plasmids groups, then fed on a normal diet (ND) for 1 month. The percentage of liver/body weight was shown. Data are presented as the mean±SD (n=6 mice per group). **P<0.01, ***p<0.001 (Student’s t test). (M) Liver tissues from above group were subjected to IB analysis. (N) Representative image of livers harvested from mice which were hydrodynamic transfected with N‐RasV12, SB transposase, and Flag-AKT1-WT, C77S, C224S, or C77/224S plasmids, then fed on ND or a high fat diet (HFD) for 2 months. The percentage of liver/body weight was shown. Data are presented as the mean±SD (n=8 mice per group). ***P<0.001, ****p<0.0001 (Student’s t test). (O) Liver tissues from above group were subjected to IB analysis. (P) Representative images of 4-week-old <em>Akt1<sup>WT</sup> </em> and <em>Akt1<sup>C224S</sup> </em> mice. Representative liver images and the percentage of liver/body weight were shown. Data are presented as the mean±SD (n=5 mice per group). ***P<0.001 (Student’s t test). (Q) Representative images and body weight of male mice fed with HFD for 11 weeks. Data are presented as the mean±SD (n=6 mice per group). ****p<0.0001 (two-way analysis of variance analysis). (R) Weights of liver, visceral adipose tissue (VAT), and brown adipose tissue (BAT) were shown as percentage of body weight. Data are presented as the mean±SD (n=6 mice per group). **P<0.01, ***p<0.001 (Student’s t test). (S) Liver tissues, VAT and BAT in (R) were stained with H&E, Oil Red, or analysed by IHC for pS6 (pS240/S244) levels. (T) Liver tissues in (R) were subjected to IB analysis. (U) <em>Akt1<sup>WT</sup> </em> and <em>Akt1<sup>C224S</sup> </em> mice fed with ND or HFD, then subjected to ABE palmitoylation assay.</div></div>" data-icon-position="" data-hide-link-title="0"><span class="hw-responsive-img"><img class="highwire-fragment fragment-image lazyload" alt="Figure 2" src="data:image/gif;base64,R0lGODlhAQABAIAAAAAAAP///yH5BAEAAAAALAAAAAABAAEAAAIBRAA7" data-src="https://gut.bmj.com/content/gutjnl/73/7/1156/F2.medium.gif" width="383" height="440"/><noscript><img class="highwire-fragment fragment-image" alt="Figure 2" src="https://gut.bmj.com/content/gutjnl/73/7/1156/F2.medium.gif" width="383" height="440"/></noscript></span></a></div></div><ul class="highwire-figure-links inline"><li class="download-fig first"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F2.large.jpg?download=true" class="highwire-figure-link highwire-figure-link-download" title="Download Figure 2" data-icon-position="" data-hide-link-title="0">Download figure</a></li> <li class="new-tab"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F2.large.jpg" class="highwire-figure-link highwire-figure-link-newtab" target="_blank" data-icon-position="" data-hide-link-title="0">Open in new tab</a></li> <li class="download-ppt last"><a href="/highwire/powerpoint/261208" class="highwire-figure-link highwire-figure-link-ppt" data-icon-position="" data-hide-link-title="0">Download powerpoint</a></li> </ul></div><div class="fig-caption"><span class="fig-label">Figure 2</span> <p id="p-21" class="first-child">AKT1 is palmitoylated at C77/C224 to enhance its kinase activity and oncogenic function. (A) Huh7 cells were labelled with dimethyl sulfoxide (DMSO) or alk-C16 (50 µM) for 4 hours, followed by click chemistry reaction to biotin-azide assay. (B) Whole cell lysates of Huh7 cells were incubated with α-AKT1 antibody and processed for acyl-biotin exchange (ABE) palmitoylation assay. (C) HepG2 cells were treated with palmitic acid (PA) (100 µM) with or without 2BP (50 µM) for 4 hours, incubated with α-AKT1 antibody and processed for ABE palmitoylation assay. (D) Mouse liver tissues obtained from <a id="xref-fig-1-9" class="xref-fig" href="#F1">figure 1E</a> were subjected for ABE palmitoylation assay. (E) Four non-alcoholic steatohepatitis (NASH)-induced hepatocellular carcinoma (HCC) with their adjacent normal tissues were subjected for ABE palmitoylation assay. N, indicates adjacent normal tissue; T, indicates liver tumour; Mock, indicates no hydroxylamine (HAM) treatment. (F) Mouse liver tissues obtained from <a id="xref-fig-1-10" class="xref-fig" href="#F1">figure 1I</a> were subjected for ABE palmitoylation assay. (G) Human liver sections with no steatosis, NASH and HCC were stained with H&E, Masson, and analysed by IHC for pS6 levels. (H) Liver tissues from above groups were subjected to ABE palmitoylation assay, and the relative palmitoylation of AKT was quantified with total AKT1 and normalised with no steatosis tissues. Data are presented as the mean±SD (n=7 for no steatosis tissues, n=8 for NASH, n=8 for HCC). **P<0.01, ***p<0.001, ****p<0.0001 (Student’s t test). (I) <em>AKT1<sup>WT</sup> </em>, <em>AKT1<sup>C77S</sup> </em> and <em>AKT1<sup>C224S</sup> </em> HEK293 cells were treated with or without PA for 4 hours, followed by ABE palmitoylation assay. (J) <em>AKT1<sup>WT</sup> </em>, <em>AKT1<sup>C77S</sup> </em> and <em>AKT1<sup>C224S</sup> </em> HEK293 cells were treated with or without PA and alk-C16 for 4 hours, subjected to click chemistry reaction to biotin-azide assay. (K) DLD1-<em>AKT1/2<sup>-/-</sup>-EV,</em> DLD1-<em>AKT1/2<sup>-/-</sup>-Flag-AKT1-WT</em>, <em>C77S</em>, <em>C224S</em>, and <em>C77/224S</em> cells were subjected to colony formation (I) and soft agar assays (J). Data are presented as the mean±SD of three independent experiments. **P<0.01, ***p<0.001, ****p<0.0001 (Student’s t test). (L) Representative image of livers harvested from hydrodynamic transfection of EV, myr-mAkt1, or myr-mAkt1-C77/224S and N‐RasV12, SB plasmids groups, then fed on a normal diet (ND) for 1 month. The percentage of liver/body weight was shown. Data are presented as the mean±SD (n=6 mice per group). **P<0.01, ***p<0.001 (Student’s t test). (M) Liver tissues from above group were subjected to IB analysis. (N) Representative image of livers harvested from mice which were hydrodynamic transfected with N‐RasV12, SB transposase, and Flag-AKT1-WT, C77S, C224S, or C77/224S plasmids, then fed on ND or a high fat diet (HFD) for 2 months. The percentage of liver/body weight was shown. Data are presented as the mean±SD (n=8 mice per group). ***P<0.001, ****p<0.0001 (Student’s t test). (O) Liver tissues from above group were subjected to IB analysis. (P) Representative images of 4-week-old <em>Akt1<sup>WT</sup> </em> and <em>Akt1<sup>C224S</sup> </em> mice. Representative liver images and the percentage of liver/body weight were shown. Data are presented as the mean±SD (n=5 mice per group). ***P<0.001 (Student’s t test). (Q) Representative images and body weight of male mice fed with HFD for 11 weeks. Data are presented as the mean±SD (n=6 mice per group). ****p<0.0001 (two-way analysis of variance analysis). (R) Weights of liver, visceral adipose tissue (VAT), and brown adipose tissue (BAT) were shown as percentage of body weight. Data are presented as the mean±SD (n=6 mice per group). **P<0.01, ***p<0.001 (Student’s t test). (S) Liver tissues, VAT and BAT in (R) were stained with H&E, Oil Red, or analysed by IHC for pS6 (pS240/S244) levels. (T) Liver tissues in (R) were subjected to IB analysis. (U) <em>Akt1<sup>WT</sup> </em> and <em>Akt1<sup>C224S</sup> </em> mice fed with ND or HFD, then subjected to ABE palmitoylation assay.</p><div class="sb-div caption-clear"></div></div></div><p id="p-22">To investigate AKT palmitoylation in vivo, we harvested liver tissues from PA or HFD-feeding mice as we mentioned above (<a id="xref-fig-1-11" class="xref-fig" href="#F1">figure 1D</a>), and observed that PA and HFD markedly enhanced AKT palmitoylation (<a id="xref-fig-2-3" class="xref-fig" href="#F2">figure 2D</a>). More importantly, compared with adjacent normal liver tissues, NASH-induced HCC displayed higher AKT palmitoylation levels (<a id="xref-fig-2-4" class="xref-fig" href="#F2">figure 2E</a>). Consisnt with the association of AKT activity with NASH/HCC progression (<a id="xref-fig-1-12" class="xref-fig" href="#F1">figure 1I</a>), we observed a progressive increase of AKT palmitoylation following HFLC-feeding time (<a id="xref-fig-2-5" class="xref-fig" href="#F2">figure 2F</a>). Consistently, NASH tissue specimens also displayed increased AKT palmitoylation, accompanied with elevated AKT activity, by contrast, HCC tissues exhibited more stronger AKT palmitoylation and activation (<a id="xref-fig-2-6" class="xref-fig" href="#F2">figure 2G–H</a> and <a id="xref-supplementary-material-1-14" class="xref-supplementary-material" href="#DC1">online supplemental figure S2K-L</a>). These observations together suggest that AKT undergoes palmitoylation modification both in NASH and HCC.</p><p id="p-23">To point out potential palmitoyl-residues, the amino acid sequence of AKT was analysed (<a id="xref-supplementary-material-1-15" class="xref-supplementary-material" href="#DC1">online supplemental figure S3A</a>). Of note, mutation of C77 or C224, but not other cystines, including previously reported C344,<a id="xref-ref-27-1" class="xref-bibr" href="#ref-27">27</a> significantly abolished AKT phosphorylation (<a id="xref-supplementary-material-1-16" class="xref-supplementary-material" href="#DC1">online supplemental figure S3B</a>), and reduced their binding to PDK1 (<a id="xref-supplementary-material-1-17" class="xref-supplementary-material" href="#DC1">online supplemental figure S3C</a>). Interestingly, C77S or/and C224S mutations clearly abolished baseline or PA-induced AKT palmitoylation (<a id="xref-supplementary-material-1-18" class="xref-supplementary-material" href="#DC1">online supplemental figure S3D-H</a>). To further investigate the physiological role of palmitoylation on AKT, we generated C77S and C224S-AKT1 knock-in HEK293 cells (termed as <em>AKT1<sup>C77S</sup> </em> and <em>AKT1<sup>C224S</sup> </em>) (<a id="xref-supplementary-material-1-19" class="xref-supplementary-material" href="#DC1">online supplemental figure S3I</a>), which showed readily reduced PA-induced AKT palmitoylation (<a id="xref-fig-2-7" class="xref-fig" href="#F2">figure 2I–J</a>), indicating that AKT undergoes palmitoyl-modification at both C77 and C224 residues.</p></div><div id="sec-13" class="subsection"><h3>Palmitoylation deficiency represses AKT kinase activity and oncogenic functions</h3><p id="p-24">Next, we examined whether PA activates AKT by enhancing its palmitoylation at C77 and C224. The results showed that C77S or/and C224S mutant significantly inhibited PA-induced or insulin-induced AKT activity (<a id="xref-supplementary-material-1-20" class="xref-supplementary-material" href="#DC1">online supplemental figure S4A</a>), phosphorylation and palmitoylation in HEK293 (<a id="xref-supplementary-material-1-21" class="xref-supplementary-material" href="#DC1">online supplemental figure S4B-D</a>) and <em>AKT1/2</em> double knockout DLD1 (DLD1-<em>AKT1/2<sup>-/-</sup> </em>) cells (<a id="xref-supplementary-material-1-22" class="xref-supplementary-material" href="#DC1">online supplemental figure S4E-G</a>). In support of this observation, ectopic expression of C77S or/and C224S-AKT1 mutants dramatically reduced colony formation and anchorage growth (<a id="xref-fig-2-8" class="xref-fig" href="#F2">figure 2K</a> and <a id="xref-supplementary-material-1-23" class="xref-supplementary-material" href="#DC1">online supplemental figure S4H-I</a>), as well as tumour growth in vivo (<a id="xref-supplementary-material-1-24" class="xref-supplementary-material" href="#DC1">online supplemental figure S4J-L</a>), coupled with decreased AKT phosphorylation (<a id="xref-supplementary-material-1-25" class="xref-supplementary-material" href="#DC1">online supplemental figure S4M</a>) in DLD1-<em>AKT1/2<sup>-/-</sup> </em> cells. Similarly, HEK293-<em>AKT1<sup>C77S</sup> </em> and <em>AKT1<sup>C224S</sup> </em> cells also exhibited decreased AKT phosphorylation on insulin or PA stimulation (<a id="xref-supplementary-material-1-26" class="xref-supplementary-material" href="#DC1">online supplemental figure S4N-R</a>), accompanied with attenuated its interaction with PDK1 (<a id="xref-supplementary-material-1-27" class="xref-supplementary-material" href="#DC1">online supplemental figure S4S-T</a>), decreased cell colony formation (<a id="xref-supplementary-material-1-28" class="xref-supplementary-material" href="#DC1">online supplemental figure S4U</a>). Interestingly, RNA-sequencing results showed that PA could strongly enhance insulin signalling, NAFLD, and activate PI3K-AKT signalling in HEK293-<em>AKT1<sup>WT</sup> </em> cells compared with <em>AKT1<sup>C224S</sup> </em> cells (<a id="xref-supplementary-material-1-29" class="xref-supplementary-material" href="#DC1">online supplemental figure S4V-W</a>), indicating the potential link of PA to AKT activation and fatty liver disease. Notably, the C77S and/or C224S mutants reduced AKT kinase activity measured by in vitro kinase assays (<a id="xref-supplementary-material-1-30" class="xref-supplementary-material" href="#DC1">online supplemental figure S4X</a>), collectively suggesting that PA partially activates AKT by promoting AKT palmitoylation at C77 and C224.</p><p id="p-25">Due to the critical role of fat deposition in NASH and its-induced liver tumorigenesis, we sought to investigate whether palmitoylation deficiency defects AKT-induced tumorigenesis in vivo. To this end, we employed the canonical hydrodynamic transfection mouse model, in comparation with intact myr-mAkt1, palmitoylation deficient mutations (myr-mAkt1-C77/224S) could significantly attenuate HCC formation (<a id="xref-fig-2-9" class="xref-fig" href="#F2">figure 2L</a>), coupled with decreased Akt phosphorylation and activation (<a id="xref-fig-2-10" class="xref-fig" href="#F2">figure 2M</a> and <a id="xref-supplementary-material-1-31" class="xref-supplementary-material" href="#DC1">online supplemental figure S5A</a>). Furthermore, to determine whether HFD-induced NASH and HCC through palmitoylation of AKT, we used the optimal murine model in combination with HFD-feeding (<a id="xref-fig-1-13" class="xref-fig" href="#F1">figure 1D</a>), and observed that injected AKT1 construct bearing different mutants (C77S and/or C224S) could obsviously alleviate HFD-induced NASH (<a id="xref-fig-2-11" class="xref-fig" href="#F2">figure 2N</a>), accompanied with decreased AKT phosphorylation, palmitoylation and downstream activation (<a id="xref-fig-2-12" class="xref-fig" href="#F2">figure 2O</a> and <a id="xref-supplementary-material-1-32" class="xref-supplementary-material" href="#DC1">online supplemental figure S5B-C</a>), indicating that palmitoylation of AKT at C77 and C224 is critical for NASH and HCC.</p><p id="p-26">To further explore the physiological role of AKT palmitoylation in vivo, we generated Akt1-C224S knock-in mice (<em>Akt1<sup>C224S</sup> </em>) (<a id="xref-supplementary-material-1-33" class="xref-supplementary-material" href="#DC1">online supplemental figure S6A-B</a>). Similar to the phenotype of <em>Akt1</em>-KO mice (<em>Akt1<sup>-/-</sup> </em>),<a id="xref-ref-28-1" class="xref-bibr" href="#ref-28">28 29</a> <em>Akt1<sup>C224S</sup> </em> mice were born with expected Mendelian ratio (<a id="xref-supplementary-material-1-34" class="xref-supplementary-material" href="#DC1">online supplemental figure S6C</a>), and displayed lower growth rate and decreased body size/weight, organ size/weight and Akt1 activation compared with intact mice (<a id="xref-fig-2-13" class="xref-fig" href="#F2">figure 2P</a> and <a id="xref-supplementary-material-1-35" class="xref-supplementary-material" href="#DC1">online supplemental figure S6D-K</a>). A similar phenomenon was also observed in mice fed a HFD (<a id="xref-fig-2-14" class="xref-fig" href="#F2">figure 2Q–T</a>), accompanied with promoting Akt1 palmitoylation (<a id="xref-fig-2-15" class="xref-fig" href="#F2">figure 2U</a>). Notably, insulin or PA stimulation had a slight effect on pT308-AKT in liver tissues or MEFs from <em>Akt1<sup>C224S</sup> </em> mice compared with the counterpart mice (<a id="xref-supplementary-material-1-36" class="xref-supplementary-material" href="#DC1">online supplemental figure S6L-O</a>). These findings suggest that defective palmitoylation impairs AKT kinase activity and physiological function.</p></div><div id="sec-14" class="subsection"><h3>Palmitoylation promotes AKT membrane localisation in a PI3K-PIP<sub>3</sub> independent manner</h3><p id="p-27">It is previously reported that palmitoyl-conjugation prefers to anchoring target proteins, such as MC1R, STAT3, N-RAS and NOD1/2, to membrane.<a id="xref-ref-5-2" class="xref-bibr" href="#ref-5">5 30–32</a> Therefore, we tended to define whether palmitoyl-conjugation could promote AKT membrane anchoring to further enhance its interaction with PDK1 (<a id="xref-supplementary-material-1-37" class="xref-supplementary-material" href="#DC1">online supplemental figure S1H-J</a>), which mainly occurs in the plasma membrane in a PIP<sub>3</sub>-dependent manner.<a id="xref-ref-16-2" class="xref-bibr" href="#ref-16">16</a> Expectedly, similar to insulin, PA also apparently enhanced AKT membrane localisation (<a id="xref-fig-3-1" class="xref-fig" href="#F3">figure 3A</a> and <a id="xref-supplementary-material-1-38" class="xref-supplementary-material" href="#DC1">online supplemental figure S7A</a>), whereas 2BP administration (<a id="xref-supplementary-material-1-39" class="xref-supplementary-material" href="#DC1">online supplemental figure S1J</a>) or C77S and/or C224S mutant (<a id="xref-fig-3-2" class="xref-fig" href="#F3">figure 3A</a> and <a id="xref-supplementary-material-1-40" class="xref-supplementary-material" href="#DC1">online supplemental figure S7A</a>) markedly impaired insulin or PA-induced AKT membrane localisation. In addition, cell fractionations also demonstrated that HEK293-<em>AKT1<sup>C77S</sup> </em> and <em>AKT1<sup>C224S</sup> </em>, as well as MEFs-<em>Akt1<sup>C224S</sup> </em> exhibited a marked decrease in AKT membrane localisation induced by insulin or PA (<a id="xref-fig-3-3" class="xref-fig" href="#F3">figure 3B</a> and <a id="xref-supplementary-material-1-41" class="xref-supplementary-material" href="#DC1">online supplemental figure S7B-D</a>), concurrently decreased AKT binding with PIP<sub>3</sub> (<a id="xref-fig-3-4" class="xref-fig" href="#F3">figure 3C–D</a> and <a id="xref-supplementary-material-1-42" class="xref-supplementary-material" href="#DC1">online supplemental figure S7E-F</a>). These findings suggest that the canonical function of insulin-activated AKT depends at least in part on AKT palmitoyl-conjugation.</p><div id="F3" class="fig pos-float odd"><div class="highwire-figure"><div class="fig-inline-img-wrapper"><div class="fig-inline-img"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F3.large.jpg?width=800&height=600&carousel=1" title="Palmitoylation conjugation enhances AKT membrane translocation independent of PIP3. (A) AKT1WT , AKT1C77S , and AKT1C224S HEK293 cells were serum-starved for 18 hours before treated with or without insulin (1 µg/mL) for 10 min, palmitic acid (PA) (100 µM) for 4 hours, or wortmannin (1 µM) for 2 hours, then subjected to endogenous IF analysis. Scale bars, 10 µm. (B) AKT1WT , AKT1C77S and AKT1C224S HEK293 cells were serum-starved for 18 hours before treated with or without PA for 4 hours, then subjected to cell membrane separation assay. (C and D) AKT1WT , AKT1C77S and AKT1C224S HEK293 cells were serum-starved for 18 hours before treated with or without insulin for 10 min (C) or PA for 4 hours (D), followed by PIP3 pull-down assay. (E) HEK293 cells were transfected with Flag-AKT1-PH or Flag-AKT1-PH-C77S, serum-starved for 18 hours before treated with or without insulin, then subjected to PIP3 pull-down assay. (F) HEK293 cells were transfected with Flag-AKT1-PH or Flag-AKT1-PH-C77S, serum-starved for 18 hours before treated with or without insulin or PA, then subjected to IF analysis. Scale bars, 10 µm. (G) HEK293 cells were serum-starved for 18 hours before treated with or without insulin for 10 min, PA for 4 hours, or wortmannin for 2 hours, followed by endogenous IF analysis. Scale bars, 10 µm." class="highwire-fragment fragment-images colorbox-load" rel="gallery-fragment-images-183578412" data-figure-caption="<div class="highwire-markup"><div xmlns="http://www.w3.org/1999/xhtml">Palmitoylation conjugation enhances AKT membrane translocation independent of PIP<sub>3</sub>. (A) <em>AKT1<sup>WT</sup> </em>, <em>AKT1<sup>C77S</sup> </em>, and <em>AKT1<sup>C224S</sup> </em> HEK293 cells were serum-starved for 18 hours before treated with or without insulin (1 µg/mL) for 10 min, palmitic acid (PA) (100 µM) for 4 hours, or wortmannin (1 µM) for 2 hours, then subjected to endogenous IF analysis. Scale bars, 10 µm. (B) <em>AKT1<sup>WT</sup> </em>, <em>AKT1<sup>C77S</sup> </em> and <em>AKT1<sup>C224S</sup> </em> HEK293 cells were serum-starved for 18 hours before treated with or without PA for 4 hours, then subjected to cell membrane separation assay. (C and D) <em>AKT1<sup>WT</sup> </em>, <em>AKT1<sup>C77S</sup> </em> and <em>AKT1<sup>C224S</sup> </em> HEK293 cells were serum-starved for 18 hours before treated with or without insulin for 10 min (C) or PA for 4 hours (D), followed by PIP3 pull-down assay. (E) HEK293 cells were transfected with Flag-AKT1-PH or Flag-AKT1-PH-C77S, serum-starved for 18 hours before treated with or without insulin, then subjected to PIP<sub>3</sub> pull-down assay. (F) HEK293 cells were transfected with Flag-AKT1-PH or Flag-AKT1-PH-C77S, serum-starved for 18 hours before treated with or without insulin or PA, then subjected to IF analysis. Scale bars, 10 µm. (G) HEK293 cells were serum-starved for 18 hours before treated with or without insulin for 10 min, PA for 4 hours, or wortmannin for 2 hours, followed by endogenous IF analysis. Scale bars, 10 µm.</div></div>" data-icon-position="" data-hide-link-title="0"><span class="hw-responsive-img"><img class="highwire-fragment fragment-image lazyload" alt="Figure 3" src="data:image/gif;base64,R0lGODlhAQABAIAAAAAAAP///yH5BAEAAAAALAAAAAABAAEAAAIBRAA7" data-src="https://gut.bmj.com/content/gutjnl/73/7/1156/F3.medium.gif" width="440" height="267"/><noscript><img class="highwire-fragment fragment-image" alt="Figure 3" src="https://gut.bmj.com/content/gutjnl/73/7/1156/F3.medium.gif" width="440" height="267"/></noscript></span></a></div></div><ul class="highwire-figure-links inline"><li class="download-fig first"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F3.large.jpg?download=true" class="highwire-figure-link highwire-figure-link-download" title="Download Figure 3" data-icon-position="" data-hide-link-title="0">Download figure</a></li> <li class="new-tab"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F3.large.jpg" class="highwire-figure-link highwire-figure-link-newtab" target="_blank" data-icon-position="" data-hide-link-title="0">Open in new tab</a></li> <li class="download-ppt last"><a href="/highwire/powerpoint/261320" class="highwire-figure-link highwire-figure-link-ppt" data-icon-position="" data-hide-link-title="0">Download powerpoint</a></li> </ul></div><div class="fig-caption"><span class="fig-label">Figure 3</span> <p id="p-28" class="first-child">Palmitoylation conjugation enhances AKT membrane translocation independent of PIP<sub>3</sub>. (A) <em>AKT1<sup>WT</sup> </em>, <em>AKT1<sup>C77S</sup> </em>, and <em>AKT1<sup>C224S</sup> </em> HEK293 cells were serum-starved for 18 hours before treated with or without insulin (1 µg/mL) for 10 min, palmitic acid (PA) (100 µM) for 4 hours, or wortmannin (1 µM) for 2 hours, then subjected to endogenous IF analysis. Scale bars, 10 µm. (B) <em>AKT1<sup>WT</sup> </em>, <em>AKT1<sup>C77S</sup> </em> and <em>AKT1<sup>C224S</sup> </em> HEK293 cells were serum-starved for 18 hours before treated with or without PA for 4 hours, then subjected to cell membrane separation assay. (C and D) <em>AKT1<sup>WT</sup> </em>, <em>AKT1<sup>C77S</sup> </em> and <em>AKT1<sup>C224S</sup> </em> HEK293 cells were serum-starved for 18 hours before treated with or without insulin for 10 min (C) or PA for 4 hours (D), followed by PIP3 pull-down assay. (E) HEK293 cells were transfected with Flag-AKT1-PH or Flag-AKT1-PH-C77S, serum-starved for 18 hours before treated with or without insulin, then subjected to PIP<sub>3</sub> pull-down assay. (F) HEK293 cells were transfected with Flag-AKT1-PH or Flag-AKT1-PH-C77S, serum-starved for 18 hours before treated with or without insulin or PA, then subjected to IF analysis. Scale bars, 10 µm. (G) HEK293 cells were serum-starved for 18 hours before treated with or without insulin for 10 min, PA for 4 hours, or wortmannin for 2 hours, followed by endogenous IF analysis. Scale bars, 10 µm.</p><div class="sb-div caption-clear"></div></div></div><p id="p-29">To elucidate how palmitoyl-conjugation affects AKT membrane translocation, we observed that depletion of PIP<sub>3</sub> by PI3K inhibitors attenuated insulin but not PA-induced AKT membrane localisation (<a id="xref-fig-3-5" class="xref-fig" href="#F3">figure 3A</a>, <a id="xref-supplementary-material-1-43" class="xref-supplementary-material" href="#DC1">online supplemental figure S7D,G</a>). On the other hand, PI3K inhibitors readily disrupted PA-induced AKT interaction with PIP<sub>3</sub> and PDK1 (<a id="xref-supplementary-material-1-44" class="xref-supplementary-material" href="#DC1">online supplemental figure S7H-I</a>) but not their membrane location (<a id="xref-supplementary-material-1-45" class="xref-supplementary-material" href="#DC1">online supplemental figure S7J-K</a>). Due to the important role of AKT-PH domain in mediating its interaction with PIP<sub>3</sub> and further membrane location,<a id="xref-ref-13-2" class="xref-bibr" href="#ref-13">13</a> both insulin-induced and PA-induced AKT-PH domain interaction with PIP<sub>3</sub> and membrane translocation could be markedly abolished by C77S mutation (<a id="xref-fig-3-6" class="xref-fig" href="#F3">figure 3E–F</a>, and <a id="xref-supplementary-material-1-46" class="xref-supplementary-material" href="#DC1">online supplemental figure S7L</a>). To directly detect whether PA-induced AKT membrane location through binding with PIP<sub>3</sub>, specific staining for PIP<sub>3</sub> was performed, and only partial colocalisation of PIP3 with PA-induced AKT membrane localisation was observed (<a id="xref-fig-3-7" class="xref-fig" href="#F3">figure 3G</a> and <a id="xref-supplementary-material-1-47" class="xref-supplementary-material" href="#DC1">online supplemental figure S7M</a>). Insulin-induced colocalisation of AKT and PIP<sub>3</sub> could be attenuated by PI3K inhibitors, while PI3K inhibitor did not impair PA-induced AKT membrane localisation (<a id="xref-fig-3-8" class="xref-fig" href="#F3">figure 3G</a> and <a id="xref-supplementary-material-1-48" class="xref-supplementary-material" href="#DC1">online supplemental figure S7M</a>). Consistent with the property of palmitoylation to anchor proteins directly at the membrane, our results indicate that palmitoylation enhances AKT membrane localisation in a PI3K-PIP<sub>3</sub> independent manner.</p></div><div id="sec-15" class="subsection"><h3>Palmitoylation deficiency facilitates AKT inactive polymer formation</h3><p id="p-30">Although palmitoylation at C77 of AKT-PH domain has been shown to promote AKT membrane anchoring, the function of palmitoylation at C224 remain unclear. As protein dimerisation/polymerisation plays a key role in its biological function, this has been shown to be regulated by palmitoyl-modification, including STAT3.<a id="xref-ref-30-1" class="xref-bibr" href="#ref-30">30</a> Therefore, we sought to detect whether palmitoylation at C77 or C224 affects AKT dimer/polymerisation. To this end, split synthetic renilla luciferase, GST pull-down, fluorescent resonance energy transfer (FRET), and disuccinimidyl suberate (DSS) crosslinking assays shown that AKT formed dimer/polymer both in vitro and in cells and C224S mutant enhanced AKT1 dimerisation (<a id="xref-fig-4-1" class="xref-fig" href="#F4">figure 4A–F</a> and <a id="xref-supplementary-material-1-49" class="xref-supplementary-material" href="#DC1">online supplemental figure S8A-B</a>). Interestingly, PA stimulation reduced AKT dimer/polymer, which was antagonised by 2BP (<a id="xref-fig-4-2" class="xref-fig" href="#F4">figure 4G</a> and <a id="xref-supplementary-material-1-50" class="xref-supplementary-material" href="#DC1">online supplemental figure S8C-E</a>). Meanwhile, the C224S but not the C77S mutant enhanced AKT dimerisation compared with WT-AKT (<a id="xref-fig-4-3" class="xref-fig" href="#F4">figure 4H</a> and <a id="xref-supplementary-material-1-51" class="xref-supplementary-material" href="#DC1">online supplemental figure S8F-G</a>). In support of this finding, C224S (but not C77S) strongly enhanced AKT aggregate formation detected by gel filtration (<a id="xref-fig-4-4" class="xref-fig" href="#F4">figure 4I–J</a> and <a id="xref-supplementary-material-1-52" class="xref-supplementary-material" href="#DC1">online supplemental figure S8H-I</a>), with a large complex containing AKT inactive species (<a id="xref-supplementary-material-1-53" class="xref-supplementary-material" href="#DC1">online supplemental figure S8H-I</a>). To further validate the effect of palmitoyl-modification for AKT dimer/polymer, we employed both insect cell and bacteria purified systems, in which palmitoyl-transferases was very low or evolutionarily lost, respectively. We observed that, similar to C224S, purified WT-AKT protein displayed large-sized probable aggregates in vitro (<a id="xref-fig-4-5" class="xref-fig" href="#F4">figure 4K–L</a> and <a id="xref-supplementary-material-1-54" class="xref-supplementary-material" href="#DC1">S8J</a>). These observations imply that palmitoylation at C224 antagonises the formation of AKT aggregates.</p><div id="F4" class="fig pos-float odd"><div class="highwire-figure"><div class="fig-inline-img-wrapper"><div class="fig-inline-img"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F4.large.jpg?width=800&height=600&carousel=1" title="Palmitoylation defects AKT polymerisation. (A) Split synthetic Renilla luciferase system. (B) HEK293 cells were transfected with EV, Renilla-N, Renilla-C, AKT1-Renilla-N, AKT1-Renilla-C, or AKT1-C224S-Renilla-C, then subjected to luciferase assay. Data are presented as the mean±SD of three independent experiments. ***P" class="highwire-fragment fragment-images colorbox-load" rel="gallery-fragment-images-183578412" data-figure-caption="<div class="highwire-markup">Palmitoylation defects AKT polymerisation. (A) Split synthetic Renilla luciferase system. (B) HEK293 cells were transfected with EV, Renilla-N, Renilla-C, AKT1-Renilla-N, AKT1-Renilla-C, or AKT1-C224S-Renilla-C, then subjected to luciferase assay. Data are presented as the mean±SD of three independent experiments. ***P<0.001, ****p<0.0001 (Student’s t test and two-way analysis of variance analysis). (C) Bacterially purified His-AKT1 protein and glutathione S-transferase (GST) or GST-AKT1 proteins were subjected to GST pull-down assay. (D) HEK293 cells were transfected with cyan fluorescent protein (CFP)-AKT1 or CFP-AKT1-C224S and yellow fluorescent protein (YFP)-AKT1 plasmids, then subjected to fluorescent resonance energy transfer (FRET) assay (the red boxes were bleached, the green boxes were not bleached, and the blue boxes were negative control). Scale bars, 10 µm. (E) The quantifications of CFP changes after YFP photobleaching. Data are presented as the mean±SD of three independent experiments. **P<0.01, ***p<0.001, ****p<0.0001 (Student’s t test and two-way analysis of variance analysis). (F) Huh7 and Jhh7 were subjected to disuccinimidyl suberate (DSS) crosslinking assay. (G) HEK293 cells were transfected with EV or Flag-AKT1, treated with PA or 2BP for 4 hours, the whole cell extract was immunoprecipitated with anti-Flag agarose and eluted with 3×Flag peptide, then subjected to Native PAGE assay. (H) HEK293 cells were transfected with EV, Flag-AKT1-WT, C77S, C224S, or C77/224S, the whole cell extract was immunoprecipitated with anti-Flag agarose and eluted with 3×Flag peptide, then subjected to Native PAGE assay. (I and J) HEK293 cells were transfected with Flag-AKT1-WT, C77S, C224S, or C77/224S, then the whole cell extract was separated by gel filtration and subjected to IB analysis (I). The quantitative trendline results of protein expression levels were quantified by Image J software (J). (K) Bacterially purified His-AKT1 protein was subjected to gel filtration and IB analysis. (L) Bacterially purified His-AKT1 was subject to in vitro palmitoylation assays with purified GST-ZDHHC24. The reaction was analysed with Native PAGE. SE, shorter exposure; LE, longer exposure.</div>" data-icon-position="" data-hide-link-title="0"><span class="hw-responsive-img"><img class="highwire-fragment fragment-image lazyload" alt="Figure 4" src="data:image/gif;base64,R0lGODlhAQABAIAAAAAAAP///yH5BAEAAAAALAAAAAABAAEAAAIBRAA7" data-src="https://gut.bmj.com/content/gutjnl/73/7/1156/F4.medium.gif" width="440" height="403"/><noscript><img class="highwire-fragment fragment-image" alt="Figure 4" src="https://gut.bmj.com/content/gutjnl/73/7/1156/F4.medium.gif" width="440" height="403"/></noscript></span></a></div></div><ul class="highwire-figure-links inline"><li class="download-fig first"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F4.large.jpg?download=true" class="highwire-figure-link highwire-figure-link-download" title="Download Figure 4" data-icon-position="" data-hide-link-title="0">Download figure</a></li> <li class="new-tab"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F4.large.jpg" class="highwire-figure-link highwire-figure-link-newtab" target="_blank" data-icon-position="" data-hide-link-title="0">Open in new tab</a></li> <li class="download-ppt last"><a href="/highwire/powerpoint/261211" class="highwire-figure-link highwire-figure-link-ppt" data-icon-position="" data-hide-link-title="0">Download powerpoint</a></li> </ul></div><div class="fig-caption"><span class="fig-label">Figure 4</span> <p id="p-31" class="first-child">Palmitoylation defects AKT polymerisation. (A) Split synthetic Renilla luciferase system. (B) HEK293 cells were transfected with EV, Renilla-N, Renilla-C, AKT1-Renilla-N, AKT1-Renilla-C, or AKT1-C224S-Renilla-C, then subjected to luciferase assay. Data are presented as the mean±SD of three independent experiments. ***P<0.001, ****p<0.0001 (Student’s t test and two-way analysis of variance analysis). (C) Bacterially purified His-AKT1 protein and glutathione S-transferase (GST) or GST-AKT1 proteins were subjected to GST pull-down assay. (D) HEK293 cells were transfected with cyan fluorescent protein (CFP)-AKT1 or CFP-AKT1-C224S and yellow fluorescent protein (YFP)-AKT1 plasmids, then subjected to fluorescent resonance energy transfer (FRET) assay (the red boxes were bleached, the green boxes were not bleached, and the blue boxes were negative control). Scale bars, 10 µm. (E) The quantifications of CFP changes after YFP photobleaching. Data are presented as the mean±SD of three independent experiments. **P<0.01, ***p<0.001, ****p<0.0001 (Student’s t test and two-way analysis of variance analysis). (F) Huh7 and Jhh7 were subjected to disuccinimidyl suberate (DSS) crosslinking assay. (G) HEK293 cells were transfected with EV or Flag-AKT1, treated with PA or 2BP for 4 hours, the whole cell extract was immunoprecipitated with anti-Flag agarose and eluted with 3×Flag peptide, then subjected to Native PAGE assay. (H) HEK293 cells were transfected with EV, Flag-AKT1-WT, C77S, C224S, or C77/224S, the whole cell extract was immunoprecipitated with anti-Flag agarose and eluted with 3×Flag peptide, then subjected to Native PAGE assay. (I and J) HEK293 cells were transfected with Flag-AKT1-WT, C77S, C224S, or C77/224S, then the whole cell extract was separated by gel filtration and subjected to IB analysis (I). The quantitative trendline results of protein expression levels were quantified by Image J software (J). (K) Bacterially purified His-AKT1 protein was subjected to gel filtration and IB analysis. (L) Bacterially purified His-AKT1 was subject to in vitro palmitoylation assays with purified GST-ZDHHC24. The reaction was analysed with Native PAGE. SE, shorter exposure; LE, longer exposure.</p><div class="sb-div caption-clear"></div></div></div></div><div id="sec-16" class="subsection"><h3>ZDHHC17/24 interact with and palmitoylate AKT</h3><p id="p-32">Next, we screened a panel of palmitoyl-transferases (ZDHHCs) to point out potential enzymes for AKT palmitoylation. Notably, certain ZDHHCs readily associated with AKT, among that ZDHHC17 and ZDHHC24 were further shown to colocalise with AKT and promote AKT palmitoylation (<a id="xref-supplementary-material-1-55" class="xref-supplementary-material" href="#DC1">online supplemental figure S9A-C</a>). Thereafter, we focused on investigating the role of ZDHHC17/24 in AKT activation. First, the PH domain of AKT was narrowed to interact with ZDHHC17, while both the PH and kinase domain bound to ZDHHC24 (<a id="xref-supplementary-material-1-56" class="xref-supplementary-material" href="#DC1">online supplemental figure S9D-E</a>). Furthermore, direct interaction of AKT with ZDHHC17/24 was demonstrated both in vitro and in cells (<a id="xref-fig-5-1" class="xref-fig" href="#F5">figure 5A</a> and <a id="xref-supplementary-material-1-57" class="xref-supplementary-material" href="#DC1">online supplemental figure S9F-G</a>). Interestingly, the binding of AKT to ZDHHC17/24 was significantly increased after PA stimulation and attenuated after 2BP administration (<a id="xref-fig-5-2" class="xref-fig" href="#F5">figure 5B</a> and <a id="xref-supplementary-material-1-58" class="xref-supplementary-material" href="#DC1">online supplemental figure S9H-I</a>), whereas insulin and PI3K inhibitors did not affect their interaction (<a id="xref-supplementary-material-1-59" class="xref-supplementary-material" href="#DC1">online supplemental figure S9J-M</a>). Moreover, WT, but not the catalytically inactive ZDHHC17/24 mutant (C467S for ZDHHC17 and C124S for ZDHHC24), could promote AKT palmitoylation in cells (<a id="xref-supplementary-material-1-60" class="xref-supplementary-material" href="#DC1">online supplemental figure S10A-H</a>). Similar results were observed in the in vitro palmitoylation assay (online supplemental figure S10I-M). Notably, ZDHHC17 and ZDHHC24 promoted AKT palmitoylation at C77 and C224, respectively, in vitro (<a id="xref-fig-5-3" class="xref-fig" href="#F5">figure 5C–D</a> and <a id="xref-supplementary-material-1-61" class="xref-supplementary-material" href="#DC1">online supplemental figure S10N-O</a>). However, the C224S mutant also decreased ZDHHC17-mediated AKT palmitoylation in cells (<a id="xref-supplementary-material-1-62" class="xref-supplementary-material" href="#DC1">online supplemental figure S10P-Q</a>), which may be due to its role in promoting the assembly of AKT polymers to evade recognition by ZDHHC17. To this end, coexpression of ZDHHC17 and ZDHHC24 further enhanced AKT palmitoylation, PIP<sub>3</sub> binding and membrane localisation (<a id="xref-supplementary-material-1-63" class="xref-supplementary-material" href="#DC1">online supplemental figure S10R-T</a>). Interestingly, consistent with the finding above that lack of palmitoyl-transferases leading to bacterially purified AKT forming polymer, ZDHHC24-mediated in vitro palmitoylation significantly reduced purified AKT aggregates (<a id="xref-fig-4-6" class="xref-fig" href="#F4">figure 4L</a>). These results suggest that ZDHHC17 and ZDHHC24-mediated palmitoylation of AKT may perform different functions (<a id="xref-supplementary-material-1-64" class="xref-supplementary-material" href="#DC1">online supplemental figure S10U</a>).</p><div id="F5" class="fig pos-float odd"><div class="highwire-figure"><div class="fig-inline-img-wrapper"><div class="fig-inline-img"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F5.large.jpg?width=800&height=600&carousel=1" title="ZDHHC24 promotes AKT palmitoylation and performs oncogenic functions. (A) Whole cell lysates of HepG2 cells were immunoprecipitated with α-ZDHHC24 antibody and subjected to IB analysis. (B) HepG2 cells were treated with or without PA or 2BP for 4 hours, then subjected to endogenous IP analysis. (C and D) Bacterially purified His-AKT1-WT, C77S, C224S, or C77/224S and GST, GST-ZDHHC24 (C), or GST-ZDHHC17 (D) proteins were subjected to in vitro palmitoylation modification assay. (E) Liver tissues of Zdhhc17+/+ , Zdhhc17-/- , Zdhhc24+/+ , Zdhhc24+/- and Zdhhc24-/- mice were subjected to IB analysis. (F) Liver tissues from above group were stained with H&E and analysed by IHC for pS6 levels. (G) Gene Set Enrichment Analysis (GSEA) analysis of non-alcoholic fatty liver disease and PI3K-AKT signalling pathway in liver tissues from Zdhhc24+/+ or Zdhhc24-/- mice that were profiled by RNA-Seq. (H) Representative image of livers harvested from Zdhhc24+/+ , Zdhhc24-/- , Zdhhc17+/+ , or Zdhhc17-/- mice which were hydrodynamic transfected with N‐RasV12, SB transposase, and Flag-AKT1 plasmids, then fed on a normal diet (ND) or a high-fat diet for 2 months. The percentage of liver/body weight was shown. Data are presented as the mean±SD (n=8 mice per group). ***P" class="highwire-fragment fragment-images colorbox-load" rel="gallery-fragment-images-183578412" data-figure-caption="<div class="highwire-markup"><div xmlns="http://www.w3.org/1999/xhtml">ZDHHC24 promotes AKT palmitoylation and performs oncogenic functions. (A) Whole cell lysates of HepG2 cells were immunoprecipitated with α-ZDHHC24 antibody and subjected to IB analysis. (B) HepG2 cells were treated with or without PA or 2BP for 4 hours, then subjected to endogenous IP analysis. (C and D) Bacterially purified His-AKT1-WT, C77S, C224S, or C77/224S and GST, GST-ZDHHC24 (C), or GST-ZDHHC17 (D) proteins were subjected to in vitro palmitoylation modification assay. (E) Liver tissues of <em>Zdhhc17<sup>+/+</sup> </em>, <em>Zdhhc17<sup>-/-</sup> </em>, <em>Zdhhc24<sup>+/+</sup> </em>, <em>Zdhhc24<sup>+/-</sup> </em> and <em>Zdhhc24<sup>-/-</sup> </em> mice were subjected to IB analysis. (F) Liver tissues from above group were stained with H&E and analysed by IHC for pS6 levels. (G) Gene Set Enrichment Analysis (GSEA) analysis of non-alcoholic fatty liver disease and PI3K-AKT signalling pathway in liver tissues from <em>Zdhhc24<sup>+/+</sup> </em> or <em>Zdhhc24<sup>-/-</sup> </em> mice that were profiled by RNA-Seq. (H) Representative image of livers harvested from <em>Zdhhc24<sup>+/+</sup> </em>, <em>Zdhhc24<sup>-/-</sup> </em>, <em>Zdhhc17<sup>+/+</sup> </em>, or <em>Zdhhc17<sup>-/-</sup> </em> mice which were hydrodynamic transfected with N‐RasV12, SB transposase, and Flag-AKT1 plasmids, then fed on a normal diet (ND) or a high-fat diet for 2 months. The percentage of liver/body weight was shown. Data are presented as the mean±SD (n=8 mice per group). ***P<0.001, ****p<0.0001 (Student’s t test). (I) Liver sections were stained with H&E and analysed by IHC for pS6, AFP, Ki67, and Cleaved-Caspase3 levels. (J) Liver tissues from above mice were subjected to ABE palmitoylation assay. (K) Representative image of livers harvested from <em>Zdhhc24<sup>+/+</sup> </em> or <em>Zdhhc24<sup>-/-</sup> </em> mice which were fed with a ND or a HFMCD diet. The percentage of liver/body weight was shown. Data are presented as the mean±SD (n=6 mice per group). **P<0.01, ***p<0.001, ****p<0.0001 (Student’s t test and two-way analysis of variance analysis). (L) Liver sections from above mice were stained with H&E, Masson and analysed by IHC for pS6, AFP, F4/80 and Ki67 levels. (M) Liver tissues from above mice were subjected to ABE palmitoylation assay. (N) Liver tissues from above mice were subjected to IB analysis.</div></div>" data-icon-position="" data-hide-link-title="0"><span class="hw-responsive-img"><img class="highwire-fragment fragment-image lazyload" alt="Figure 5" src="data:image/gif;base64,R0lGODlhAQABAIAAAAAAAP///yH5BAEAAAAALAAAAAABAAEAAAIBRAA7" data-src="https://gut.bmj.com/content/gutjnl/73/7/1156/F5.medium.gif" width="338" height="440"/><noscript><img class="highwire-fragment fragment-image" alt="Figure 5" src="https://gut.bmj.com/content/gutjnl/73/7/1156/F5.medium.gif" width="338" height="440"/></noscript></span></a></div></div><ul class="highwire-figure-links inline"><li class="download-fig first"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F5.large.jpg?download=true" class="highwire-figure-link highwire-figure-link-download" title="Download Figure 5" data-icon-position="" data-hide-link-title="0">Download figure</a></li> <li class="new-tab"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F5.large.jpg" class="highwire-figure-link highwire-figure-link-newtab" target="_blank" data-icon-position="" data-hide-link-title="0">Open in new tab</a></li> <li class="download-ppt last"><a href="/highwire/powerpoint/261253" class="highwire-figure-link highwire-figure-link-ppt" data-icon-position="" data-hide-link-title="0">Download powerpoint</a></li> </ul></div><div class="fig-caption"><span class="fig-label">Figure 5</span> <p id="p-33" class="first-child">ZDHHC24 promotes AKT palmitoylation and performs oncogenic functions. (A) Whole cell lysates of HepG2 cells were immunoprecipitated with α-ZDHHC24 antibody and subjected to IB analysis. (B) HepG2 cells were treated with or without PA or 2BP for 4 hours, then subjected to endogenous IP analysis. (C and D) Bacterially purified His-AKT1-WT, C77S, C224S, or C77/224S and GST, GST-ZDHHC24 (C), or GST-ZDHHC17 (D) proteins were subjected to in vitro palmitoylation modification assay. (E) Liver tissues of <em>Zdhhc17<sup>+/+</sup> </em>, <em>Zdhhc17<sup>-/-</sup> </em>, <em>Zdhhc24<sup>+/+</sup> </em>, <em>Zdhhc24<sup>+/-</sup> </em> and <em>Zdhhc24<sup>-/-</sup> </em> mice were subjected to IB analysis. (F) Liver tissues from above group were stained with H&E and analysed by IHC for pS6 levels. (G) Gene Set Enrichment Analysis (GSEA) analysis of non-alcoholic fatty liver disease and PI3K-AKT signalling pathway in liver tissues from <em>Zdhhc24<sup>+/+</sup> </em> or <em>Zdhhc24<sup>-/-</sup> </em> mice that were profiled by RNA-Seq. (H) Representative image of livers harvested from <em>Zdhhc24<sup>+/+</sup> </em>, <em>Zdhhc24<sup>-/-</sup> </em>, <em>Zdhhc17<sup>+/+</sup> </em>, or <em>Zdhhc17<sup>-/-</sup> </em> mice which were hydrodynamic transfected with N‐RasV12, SB transposase, and Flag-AKT1 plasmids, then fed on a normal diet (ND) or a high-fat diet for 2 months. The percentage of liver/body weight was shown. Data are presented as the mean±SD (n=8 mice per group). ***P<0.001, ****p<0.0001 (Student’s t test). (I) Liver sections were stained with H&E and analysed by IHC for pS6, AFP, Ki67, and Cleaved-Caspase3 levels. (J) Liver tissues from above mice were subjected to ABE palmitoylation assay. (K) Representative image of livers harvested from <em>Zdhhc24<sup>+/+</sup> </em> or <em>Zdhhc24<sup>-/-</sup> </em> mice which were fed with a ND or a HFMCD diet. The percentage of liver/body weight was shown. Data are presented as the mean±SD (n=6 mice per group). **P<0.01, ***p<0.001, ****p<0.0001 (Student’s t test and two-way analysis of variance analysis). (L) Liver sections from above mice were stained with H&E, Masson and analysed by IHC for pS6, AFP, F4/80 and Ki67 levels. (M) Liver tissues from above mice were subjected to ABE palmitoylation assay. (N) Liver tissues from above mice were subjected to IB analysis.</p><div class="sb-div caption-clear"></div></div></div></div><div id="sec-17" class="subsection"><h3>ZDHHC17/24 play oncogenic roles by activating AKT</h3><p id="p-34">Next, we tended to explore the biological functions of ZDHHC17/24 in activating AKT. To this end, ectopic expression of both ZDHHC17 and ZDHHC24 enhanced, whereas depletion of <em>ZDHHC17</em> or <em>ZDHHC24</em> reduced AKT phosphorylation on insulin or PA stimulation (<a id="xref-supplementary-material-1-65" class="xref-supplementary-material" href="#DC1">online supplemental figure S11A-G</a>). Consistently, ZDHHC17/24 significantly enhanced the interaction of AKT and PDK1 (<a id="xref-supplementary-material-1-66" class="xref-supplementary-material" href="#DC1">online supplemental figure S11H-I</a>), whereas expression of WT but not catalytically inactive ZDHHC24 reduced AKT dimerisation (<a id="xref-supplementary-material-1-67" class="xref-supplementary-material" href="#DC1">online supplemental figure S11J</a>). Furthermore, depletion of <em>ZDHHC24</em> largely enhanced AKT polymerisation compared with counterpart cells (<a id="xref-supplementary-material-1-68" class="xref-supplementary-material" href="#DC1">online supplemental figure S11K</a>). To investigate the biological functions of ZDHHC17/24, ectopic expression of WT but not catalytically inactive ZDHHC17/24 enhanced the growth and colony formation of normal hepatic cell THLE3 (<a id="xref-supplementary-material-1-69" class="xref-supplementary-material" href="#DC1">online supplemental figure S12A-E</a>). Furthermore, depletion of <em>ZDHHC17</em> or <em>ZDHHC24</em> significantly reduced malignancy and tumour growth in HCC cells (<a id="xref-supplementary-material-1-70" class="xref-supplementary-material" href="#DC1">online supplemental figure S12F-R</a>), indicating the oncogenic roles of ZDHHC17 and ZDHHC24. To examine whether ZDHHC17/24 exert oncogenic effects through palmitoylation and activation of AKT, we found that the AKT inhibitor MK2206 significantly inhibited WT but only slightly affected ZDHHC17/24-induced colony formation (<a id="xref-supplementary-material-1-71" class="xref-supplementary-material" href="#DC1">online supplemental figure S13A-B</a>). Furthermore, ectopic expression of ZDHHC17/24 markedly promoted cell growth in WT, but not in C77S or C224S mutant AKT1, rescued DLD1-<em>AKT1/2<sup>-/-</sup> </em> cells (<a id="xref-supplementary-material-1-72" class="xref-supplementary-material" href="#DC1">online supplemental figure S13C-D</a>).</p><p id="p-35">To explore the physiological role of ZDHHC17/24 in vivo, <em>Zdhhc17</em> and <em>Zdhhc24</em> knockout mice (termed <em>Zdhhc17<sup>-/</sup> </em> <sup>-</sup> and <em>Zdhhc24<sup>-/-</sup> </em>) were generated. Of note, both <em>Zdhhc17<sup>-/</sup> </em> <sup>-</sup> and <em>Zdhhc24<sup>-/</sup> </em> <sup>-</sup> mice exhibited reduced Akt phosphorylation (<a id="xref-fig-5-4" class="xref-fig" href="#F5">figure 5E–F</a> and <a id="xref-supplementary-material-1-73" class="xref-supplementary-material" href="#DC1">online supplemental figure S14A-M</a>), along with reduced Akt palmitoylation (<a id="xref-supplementary-material-1-74" class="xref-supplementary-material" href="#DC1">online supplemental figure S14N</a>). Furthermore, RNA-sequencing-based transcriptomic analysis of liver tissues from <em>Zdhhc24<sup>-/-</sup> </em> and counterpart mice showed that depletion of <em>Zdhhc24</em> severely impaired the metabolic functions in liver tissues, such as reducing NAFLD, and repressing PI3K-AKT signalling pathway (<a id="xref-fig-5-5" class="xref-fig" href="#F5">figure 5G</a> and <a id="xref-supplementary-material-1-75" class="xref-supplementary-material" href="#DC1">online supplemental figure S15A-E</a>), suggesting a potent role of ZDHHC24 in AKT activation and liver diseases. Furthermore, in the optimised HFD-fed hydrodynamics transfection NASH/HCC mouse model, ablation of <em>Zdhhc17</em> and <em>Zdhhc24</em> resisted NASH induced by Akt-transfection (<a id="xref-fig-5-6" class="xref-fig" href="#F5">figure 5H</a>), coupled with reduced Akt phosphorylation, palmitoylation and its downstream activation (<a id="xref-fig-5-7" class="xref-fig" href="#F5">figure 5I–J</a> and <a id="xref-supplementary-material-1-76" class="xref-supplementary-material" href="#DC1">online supplemental figure S15F-G</a>). To confirm the potential role of ZDHHC17/24 in NASH, we employed HFMCD-based NASH mouse model.<a id="xref-ref-33-1" class="xref-bibr" href="#ref-33">33 34</a> The results showed that deletion of <em>Zdhhc17</em> or <em>Zdhhc24</em> could protect HFMCD-induced NASH (<a id="xref-fig-5-8" class="xref-fig" href="#F5">figure 5K–L</a> and <a id="xref-supplementary-material-1-77" class="xref-supplementary-material" href="#DC1">online supplemental figure S15H-I</a>), coupled with decreased AKT palmitoylation and activation (<a id="xref-fig-5-9" class="xref-fig" href="#F5">figure 5M–N</a> and <a id="xref-supplementary-material-1-78" class="xref-supplementary-material" href="#DC1">online supplemental figure S15J-K</a>). Together, these results suggest that ZDHHC17/24-mediated palmitoylation and activation of AKT is important for their function in promoting NASH and HCC.</p><p id="p-36">Bioinformatically, <em>ZDHHC17/24</em> were observed to be amplified in various cancers, including about 2% of HCC (<a id="xref-supplementary-material-1-79" class="xref-supplementary-material" href="#DC1">online supplemental figure S16A</a>), which mutually excluded with AKT activation pathways, such as alterations in <em>PIK3CA</em>, <em>PTEN</em>, <em>AKT1</em> and <em>EGFR</em> (<a id="xref-supplementary-material-1-80" class="xref-supplementary-material" href="#DC1">online supplemental figure S16B</a>). To test this finding, expression of ZDHHC17/24 and activation of AKT (indicated by pS6) were detected in samples of patient with HCC. As a result, ZDHHC17/24 were relatively highly expressed in HCC compared with adjacent normal tissues (<a id="xref-fig-6-1" class="xref-fig" href="#F6">figure 6A</a> and <a id="xref-supplementary-material-1-81" class="xref-supplementary-material" href="#DC1">online supplemental figure S16C</a>), and positively correlated with AKT activation (<a id="xref-fig-6-2" class="xref-fig" href="#F6">figure 6B</a> and <a id="xref-supplementary-material-1-82" class="xref-supplementary-material" href="#DC1">online supplemental figure S16D</a>). More importantly, both the expression of ZDHHC17 and ZDHHC24 were significantly associated with the poor survival rate of patients with HCC (<a id="xref-supplementary-material-1-83" class="xref-supplementary-material" href="#DC1">online supplemental figure S16E</a>), suggesting that amplification of <em>ZDHHC17/24</em> may promote HCC through activation of AKT.</p><div id="F6" class="fig pos-float odd"><div class="highwire-figure"><div class="fig-inline-img-wrapper"><div class="fig-inline-img"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F6.large.jpg?width=800&height=600&carousel=1" title="Targeting AKT palmitoylation to repress AKT activity and oncogenic roles in hepatocellular carcinoma (HCC). (A) IHC staining of ZDHHC17, ZDHHC24 and pS6 (pS240/S244) were performed in liver tumour and adjacent tissues. (B) The correlation of pS6 (pS240/S244) and ZDHHC17 or ZDHHC24 protein levels was calculated in liver carcinoma cirrhosis microarray. ****P" class="highwire-fragment fragment-images colorbox-load" rel="gallery-fragment-images-183578412" data-figure-caption="<div class="highwire-markup"><div xmlns="http://www.w3.org/1999/xhtml">Targeting AKT palmitoylation to repress AKT activity and oncogenic roles in hepatocellular carcinoma (HCC). (A) IHC staining of ZDHHC17, ZDHHC24 and pS6 (pS240/S244) were performed in liver tumour and adjacent tissues. (B) The correlation of pS6 (pS240/S244) and ZDHHC17 or ZDHHC24 protein levels was calculated in liver carcinoma cirrhosis microarray. ****P<0.0001 (χ<sup>2</sup> test). (C) Schematic of AKT1-C224 and AKT1-S224 peptides. The different residues are shown in red. CPPtat, CPP from HIV-1 Tat protein. (D) HEK293 cells were transfected with Flag-AKT1 or Flag-AKT1-C224S, treated with or without AKT1-C224 and AKT1-S224 peptides (10 µM) for 8 hours, then subjected to acyl-biotin exchange (ABE) palmitoylation assay. (E) Huh7 cells were serum-starved for 10 hours, treated with AKT1-C224 and AKT1-S224 peptides (10 µM) for 8 hours, before stimulated with or without palmitic acid (PA), then subjected to IB analysis. (F–H) Mice bearing Huh7 xenografts were treated with AKT1-C224 and AKT1-S224 peptides (10 mg/kg/day, I.P.), and tumour sizes were monitored (F). Data are presented as the mean±SD (n=8 mice per group). N.s, no significant; ***P<0.001 (two-way analysis of variance analysis). Tumours were dissected (G) and weighed (H). Data are presented as the mean±SD (n=8 mice per group). ***P<0.001 (Student’s t test). (I) Tumours tissues in (G) were subjected to ABE palmitoylation assay. (J) Illustration of the flow for treating Cebpb-Tta-TetO-Myc mice with AKT competing peptides. Briefly, female mice were terminated doxycycline water (100 μg/mL) at age 4 weeks, and then injected intraperitoneally with PBS, AKT1-C224, or AKT1-S224 peptides (10 mg/kg/day) for 10 days. The livers were harvest for further analysis. (K and L) Macroscopic liver images of Tet-on C-Myc mice with different treatments (K). And the ration of liver and bodyweight was calculated (L). (n=7 mice per group). ***P<0.001 (Student’s t test). (M) Cebpb-Tta-TetO-Myc female mice were terminated doxycycline water (100 µg/mL) at age 4 weeks, and then injected intraperitoneally with phosphate buffered saline (PBS), AKT1-C224, or AKT1-S224 peptides (10 mg/kg/day) until death, and then survival analysis of these mice was performed. (n=8 mice per group). ***P<0.001 (Log-rank Mantel-Cox test). (N–P) Liver sections were stained with H&E, Masson, and analysed by IHC for pS6 (pS240/244), pGSK3β, AFP, Ki67, Cleaved-Caspase3, Pan-CK, and CPS1 levels (N). The dissected tissues were harvested for IB analysis (O) and palmitoylation assay (P).</div></div>" data-icon-position="" data-hide-link-title="0"><span class="hw-responsive-img"><img class="highwire-fragment fragment-image lazyload" alt="Figure 6" src="data:image/gif;base64,R0lGODlhAQABAIAAAAAAAP///yH5BAEAAAAALAAAAAABAAEAAAIBRAA7" data-src="https://gut.bmj.com/content/gutjnl/73/7/1156/F6.medium.gif" width="367" height="440"/><noscript><img class="highwire-fragment fragment-image" alt="Figure 6" src="https://gut.bmj.com/content/gutjnl/73/7/1156/F6.medium.gif" width="367" height="440"/></noscript></span></a></div></div><ul class="highwire-figure-links inline"><li class="download-fig first"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F6.large.jpg?download=true" class="highwire-figure-link highwire-figure-link-download" title="Download Figure 6" data-icon-position="" data-hide-link-title="0">Download figure</a></li> <li class="new-tab"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F6.large.jpg" class="highwire-figure-link highwire-figure-link-newtab" target="_blank" data-icon-position="" data-hide-link-title="0">Open in new tab</a></li> <li class="download-ppt last"><a href="/highwire/powerpoint/261201" class="highwire-figure-link highwire-figure-link-ppt" data-icon-position="" data-hide-link-title="0">Download powerpoint</a></li> </ul></div><div class="fig-caption"><span class="fig-label">Figure 6</span> <p id="p-37" class="first-child">Targeting AKT palmitoylation to repress AKT activity and oncogenic roles in hepatocellular carcinoma (HCC). (A) IHC staining of ZDHHC17, ZDHHC24 and pS6 (pS240/S244) were performed in liver tumour and adjacent tissues. (B) The correlation of pS6 (pS240/S244) and ZDHHC17 or ZDHHC24 protein levels was calculated in liver carcinoma cirrhosis microarray. ****P<0.0001 (χ<sup>2</sup> test). (C) Schematic of AKT1-C224 and AKT1-S224 peptides. The different residues are shown in red. CPPtat, CPP from HIV-1 Tat protein. (D) HEK293 cells were transfected with Flag-AKT1 or Flag-AKT1-C224S, treated with or without AKT1-C224 and AKT1-S224 peptides (10 µM) for 8 hours, then subjected to acyl-biotin exchange (ABE) palmitoylation assay. (E) Huh7 cells were serum-starved for 10 hours, treated with AKT1-C224 and AKT1-S224 peptides (10 µM) for 8 hours, before stimulated with or without palmitic acid (PA), then subjected to IB analysis. (F–H) Mice bearing Huh7 xenografts were treated with AKT1-C224 and AKT1-S224 peptides (10 mg/kg/day, I.P.), and tumour sizes were monitored (F). Data are presented as the mean±SD (n=8 mice per group). N.s, no significant; ***P<0.001 (two-way analysis of variance analysis). Tumours were dissected (G) and weighed (H). Data are presented as the mean±SD (n=8 mice per group). ***P<0.001 (Student’s t test). (I) Tumours tissues in (G) were subjected to ABE palmitoylation assay. (J) Illustration of the flow for treating Cebpb-Tta-TetO-Myc mice with AKT competing peptides. Briefly, female mice were terminated doxycycline water (100 μg/mL) at age 4 weeks, and then injected intraperitoneally with PBS, AKT1-C224, or AKT1-S224 peptides (10 mg/kg/day) for 10 days. The livers were harvest for further analysis. (K and L) Macroscopic liver images of Tet-on C-Myc mice with different treatments (K). And the ration of liver and bodyweight was calculated (L). (n=7 mice per group). ***P<0.001 (Student’s t test). (M) Cebpb-Tta-TetO-Myc female mice were terminated doxycycline water (100 µg/mL) at age 4 weeks, and then injected intraperitoneally with phosphate buffered saline (PBS), AKT1-C224, or AKT1-S224 peptides (10 mg/kg/day) until death, and then survival analysis of these mice was performed. (n=8 mice per group). ***P<0.001 (Log-rank Mantel-Cox test). (N–P) Liver sections were stained with H&E, Masson, and analysed by IHC for pS6 (pS240/244), pGSK3β, AFP, Ki67, Cleaved-Caspase3, Pan-CK, and CPS1 levels (N). The dissected tissues were harvested for IB analysis (O) and palmitoylation assay (P).</p><div class="sb-div caption-clear"></div></div></div></div><div id="sec-18" class="subsection"><h3>APT2 reduces AKT palmitoylation and oncogenic functions</h3><p id="p-38">Palmitoylation is the only reversible fatty modification that can be converted by acyl-protein thioesterases such as APT1 and APT2.<a id="xref-ref-35-1" class="xref-bibr" href="#ref-35">35 36</a> To this end, APT2, but not APT1, was identified to interact with AKT mainly in its PH domain (<a id="xref-supplementary-material-1-84" class="xref-supplementary-material" href="#DC1">online supplemental figure S17A-D</a>). As a result, PA-induced AKT palmitoylation could be attenuated by APT2, which was further observed as a main eraser of AKT palmitoylation (<a id="xref-supplementary-material-1-85" class="xref-supplementary-material" href="#DC1">online supplemental figure S17E-F</a>). Consistent with this finding, ectopic expression of APT2 reduced AKT phosphorylation, membrane translocation, and PDK1 interaction on insulin or PA stimulation (<a id="xref-supplementary-material-1-86" class="xref-supplementary-material" href="#DC1">online supplemental figure S17G-K</a>). However, depletion of <em>APT2</em> markedly elevated cell proliferation while enhancing AKT palmitoylation and phosphorylation (<a id="xref-supplementary-material-1-87" class="xref-supplementary-material" href="#DC1">online supplemental figure S17L-N</a>). Moreover, ectopic expression of APT2 could enhance AKT polymerisation (<a id="xref-supplementary-material-1-88" class="xref-supplementary-material" href="#DC1">online supplemental figure S17O</a>). These findings suggest that APT2 plays an opposite role to ZDHHC17/24 in regulating AKT palmitoylation, activity and oncogenic function.</p></div><div id="sec-19" class="subsection"><h3>Targeting AKT palmitoylation to repress HCC tumour growth</h3><p id="p-39">Since PA can be synthesised by the tricarboxylic acid (TCA) cycle with citrate as the source and fatty acid synthase (FASN) as the restrict enzyme,<a id="xref-ref-37-1" class="xref-bibr" href="#ref-37">37</a> abnormal homeostasis of this process also leads to PA accumulation, especially in tumours. Previous reports have shown that, in fact, liposynthetic enzymes including FASN were elevated in NASH and HCC.<a id="xref-ref-38-1" class="xref-bibr" href="#ref-38">38</a> Furthermore, amplification of <em>FASN</em> tended to be mutually exclusive with AKT activation and <em>ZDHHC17/24</em> amplification in HCC (<a id="xref-supplementary-material-1-89" class="xref-supplementary-material" href="#DC1">online supplemental figure S18A</a>), indicating that abnormal FASN would lead to synthesis of PA, in turn promoting NASH and HCC. Consistent with previous findings, the approved anti-obesity drug FASN inhibitor orlistat can fight NASH, and synergise with chemotherapeutics to treat cancer, possibly by activating AKT.<a id="xref-ref-39-1" class="xref-bibr" href="#ref-39">39</a> Therefore, we hypnotised that the elevation of FASN may palmitoylate and activate AKT through the synthesis of PA. To this end, we observed that orlistat attenuated AKT palmitoylation, activation and membrane location (<a id="xref-supplementary-material-1-90" class="xref-supplementary-material" href="#DC1">online supplemental figure S18B-D</a>). Meanwhile, orlistat could significantly attenuate AKT palmitoylation, activation and the proliferation and colony formation of cells bearing WT but not C77/224S-AKT (<a id="xref-supplementary-material-1-91" class="xref-supplementary-material" href="#DC1">online supplemental figure S18E-F</a>). Consistent with these findings, deletion of <em>ZDHHC24</em> was resistant to orlistat treatment both in cells (<a id="xref-supplementary-material-1-92" class="xref-supplementary-material" href="#DC1">online supplemental figure S18G</a>) and in vivo (<a id="xref-supplementary-material-1-93" class="xref-supplementary-material" href="#DC1">online supplemental figure S18H-K</a>), indicating that aberrant synthesis of PA can be eliminated by FASN inhibitors, resulting in reduced AKT palmitoylation and activation against according cancers.</p><p id="p-40">As intake of PA from food bypasses FASN, we observed that orlistat could not efficiently block PA accumulation on PA stimulation (<a id="xref-supplementary-material-1-94" class="xref-supplementary-material" href="#DC1">online supplemental figure S19A-B</a>). As a result, orlistat displayed inefficacy to block free PA-induced AKT phosphorylation and PIP<sub>3</sub> interaction (<a id="xref-supplementary-material-1-95" class="xref-supplementary-material" href="#DC1">online supplemental figure S19C-D</a>). Alternatively, aim to directly block AKT palmitoylation, we synthesised candidate peptides containing the intact (C224) or mutant (S224) region of AKT1 (<a id="xref-fig-6-3" class="xref-fig" href="#F6">figure 6C</a>), wherein the CPPtat sequence was included to enable the peptides penetrating cells.<a id="xref-ref-40-1" class="xref-bibr" href="#ref-40">40</a> Notably, the C224 but not the S224 peptide significantly attenuated AKT palmitoylation (<a id="xref-fig-6-4" class="xref-fig" href="#F6">figure 6D</a>), and insulin/PA-induced AKT activation (<a id="xref-fig-6-5" class="xref-fig" href="#F6">figure 6E</a> and <a id="xref-supplementary-material-1-96" class="xref-supplementary-material" href="#DC1">online supplemental figure S19E</a>). Furthermore, administration of C224 but not S224 peptides markedly reduced hepatoma cell growth and colony formation (<a id="xref-supplementary-material-1-97" class="xref-supplementary-material" href="#DC1">online supplemental figure S19F-G</a>) and in vivo tumour growth (<a id="xref-fig-6-6" class="xref-fig" href="#F6">figure 6F–H</a>), coupled with decreased AKT palmitoylation, activation (<a id="xref-fig-6-7" class="xref-fig" href="#F6">figure 6I</a> and <a id="xref-supplementary-material-1-98" class="xref-supplementary-material" href="#DC1">online supplemental figure S19H</a>) and promotion of proliferation index (Ki67), as well as enhanced cell apoptosis (Cleaved Caspase 3 and PARP) (<a id="xref-supplementary-material-1-99" class="xref-supplementary-material" href="#DC1">online supplemental figure S19I</a>). Since blocking AKT could diminish <em>C-Myc</em> transgenic mice HCC tumorigenesis,<a id="xref-ref-41-1" class="xref-bibr" href="#ref-41">41</a> we employed this murine model to validate AKT target peptides (<a id="xref-fig-6-8" class="xref-fig" href="#F6">figure 6J</a>). The results showed that C224 but not the S224 peptides attenuated <em>C-Myc</em>-induced HCC formation (<a id="xref-fig-6-9" class="xref-fig" href="#F6">figure 6K–L</a> and <a id="xref-supplementary-material-1-100" class="xref-supplementary-material" href="#DC1">online supplemental figure S19J</a>) and mouse survival (<a id="xref-fig-6-10" class="xref-fig" href="#F6">figure 6M</a>), coupled with decreased AKT activity and palmitoylation (<a id="xref-fig-6-11" class="xref-fig" href="#F6">figure 6N–P</a>). These findings suggest that specific peptides against AKT palmitoylation may benefit PA-induced tumour growth.</p><p id="p-41">Thereby, amplified ZDHHC17/24 or accumulated PA by HFD or amplification of <em>FASN</em>-mediated synthesis promotes AKT activation through S-palmitoylation, promoting NASH and further liver tumorigenesis. Therefore, adopting a low-fat (PA-restricted) diet, restricting PA synthesis (with orlistat), directly targeting AKT palmitoylation (AKT peptides) or ZDHHC17/24 (2BP or specific inhibitors) will provide effective strategies for HCC therapies (<a id="xref-fig-7-1" class="xref-fig" href="#F7">figure 7</a>).</p><div id="F7" class="fig pos-float odd"><div class="highwire-figure"><div class="fig-inline-img-wrapper"><div class="fig-inline-img"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F7.large.jpg?width=800&height=600&carousel=1" title="Model of palmitoylation enhancing AKT activity and oncogenic function. Schematic showing that palmitic acid (PA) uptake, and amplified ZDHHC or FASN could promote AKT activation through S-palmitoylation to promote non-alcoholic steatohepatitis (NASH) and subsequent liver cancer, which could be ameliorated by PA restriction and orlistat-mediated PA blocking, or targeting AKT palmitoylation by ZDHHC inhibitors or specific AKT peptides." class="highwire-fragment fragment-images colorbox-load" rel="gallery-fragment-images-183578412" data-figure-caption="<div class="highwire-markup"><div xmlns="http://www.w3.org/1999/xhtml">Model of palmitoylation enhancing AKT activity and oncogenic function. Schematic showing that palmitic acid (PA) uptake, and amplified <em>ZDHHC</em> or <em>FASN</em> could promote AKT activation through S-palmitoylation to promote non-alcoholic steatohepatitis (NASH) and subsequent liver cancer, which could be ameliorated by PA restriction and orlistat-mediated PA blocking, or targeting AKT palmitoylation by ZDHHC inhibitors or specific AKT peptides.</div></div>" data-icon-position="" data-hide-link-title="0"><span class="hw-responsive-img"><img class="highwire-fragment fragment-image lazyload" alt="Figure 7" src="data:image/gif;base64,R0lGODlhAQABAIAAAAAAAP///yH5BAEAAAAALAAAAAABAAEAAAIBRAA7" data-src="https://gut.bmj.com/content/gutjnl/73/7/1156/F7.medium.gif" width="440" height="225"/><noscript><img class="highwire-fragment fragment-image" alt="Figure 7" src="https://gut.bmj.com/content/gutjnl/73/7/1156/F7.medium.gif" width="440" height="225"/></noscript></span></a></div></div><ul class="highwire-figure-links inline"><li class="download-fig first"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F7.large.jpg?download=true" class="highwire-figure-link highwire-figure-link-download" title="Download Figure 7" data-icon-position="" data-hide-link-title="0">Download figure</a></li> <li class="new-tab"><a href="https://gut.bmj.com/content/gutjnl/73/7/1156/F7.large.jpg" class="highwire-figure-link highwire-figure-link-newtab" target="_blank" data-icon-position="" data-hide-link-title="0">Open in new tab</a></li> <li class="download-ppt last"><a href="/highwire/powerpoint/261247" class="highwire-figure-link highwire-figure-link-ppt" data-icon-position="" data-hide-link-title="0">Download powerpoint</a></li> </ul></div><div class="fig-caption"><span class="fig-label">Figure 7</span> <p id="p-42" class="first-child">Model of palmitoylation enhancing AKT activity and oncogenic function. Schematic showing that palmitic acid (PA) uptake, and amplified <em>ZDHHC</em> or <em>FASN</em> could promote AKT activation through S-palmitoylation to promote non-alcoholic steatohepatitis (NASH) and subsequent liver cancer, which could be ameliorated by PA restriction and orlistat-mediated PA blocking, or targeting AKT palmitoylation by ZDHHC inhibitors or specific AKT peptides.</p><div class="sb-div caption-clear"></div></div></div></div></div><div class="section discussion" id="sec-20"><h2 class="">Discussion</h2><p id="p-43">Here, based on the HFLC mouse model of NAFLD and human liver specimens, we observed that AKT palmitoylation and activation are positively associated with the progression from NASH to HCC. Importantly, HFD (or PA stimulation) can promote the formation of NASH and HCC in a WT Akt hydrodynamically transfected mouse model. These findings were confirmed in mouse models under palmitoylation-deficient Akt<em>-C77/224S</em> hydrodynamic transfection or <em>Zdhhc17/24</em> knockout conditions. We further established a HFMCD-based NASH mouse model, in which depletion of <em>Zdhhc17/24</em> effectively alleviates HFMCD-feeding-induced NASH. These observations suggest that AKT palmitoylation and activation play an important role in promoting NASH and HCC. Although hydrodynamic transfection HCC mouse models have shown the important role of high-fat induced AKT activation in NASH/HCC, generatinon of the liver-specific <em>Akt</em> knockout or <em>Akt-C224S</em> knockin mice, using for HFLC and HFMCD models, are important to further confirm AKT roles in the progression of NAFLD (from NASH to HCC).</p><p id="p-44">Although HFD-induced obesity play a key role in tumorigenesis, progression, metastasis and drug resistance mainly through manipulation of metabolic homeostasis and inflammation,<a id="xref-ref-42-1" class="xref-bibr" href="#ref-42">42</a> the detailed mechanisms of different fatty acids in these malignancies remain unclear. Although cholesterol synthesis compensates for fatty acid inhibition in HCC,<a id="xref-ref-43-1" class="xref-bibr" href="#ref-43">43</a> we have not observed obviously compensatory activation of the cholesterol pathway following inhibition of palmitoylation (data not shown). Alternatively, in this study, we focused on one long-chain saturated fatty acid, PA, which is a major component of palmitic oil and has been reported to play a potential role in oral tumour metastasis.<a id="xref-ref-2-4" class="xref-bibr" href="#ref-2">2</a> In addition to its metabolic role in supporting lipogenesis, here we observed that PA can directly activate AKT through palmitoyl-modification. More importantly, the baseline of AKT palmitoylation is important for AKT response to insulin or other growth factor stimulation.</p><p id="p-45">Since PA is mainly derived from two pathways, FASN-mediated synthesis following the TCA cycle and intake from food. We used the FASN inhibitor orlistat, which has been approved for clinical obesity treatment. We found that orlistat could largely attenuate AKT palmitoyl-modification and activation by reducing basal PA levels, especially under insulin-stimulation conditions. However, orlistat was unable to block PA-induced AKT activity, suggesting that orlistat was not effective in overcoming PA intake-induced obesity or cancer, a possibility of tumour resistance to orlistat, while potent amplification of FASN-induced PA accumulation and AKT activation may benefit from this inhibitor. Due to the important physiological roles of PA in providing lipids for neuroprotection, orlistat would cause potential side effects. Alternatively, we develop small peptides that can compete ZDHHC24-mediated AKT palmitoylation and activation, resulting in decreased its oncogenic functions, with the potent efficacy for HCC therapy. As we demonstrate above, <em>ZDHHC17/24</em> are also observed amplification in HCC, indicating poor outcome for patients with HCC and acting as oncogenes through activation of AKT, which provides a promising target for HCC therapy. Thus, developing the specific inhibitors to target ZDHHC17/24 would provide a promising strategy for cancer therapy.</p><p id="p-46">In sum, our findings not only enclose a fine-tuned regulation of AKT activity by PA-ZDHHC-mediated palmitoylation, but also highlight strategies for HCC therapy by limiting PA uptake, restricting PA synthesis and targeting ZDHHC17/24.</p></div><div class="section data-availability" id="sec-21"><h2 class="">Data availability statement</h2><p id="p-55">Data are available upon reasonable request.</p></div><div class="section ethics-statement" id="sec-22"><h2 class="">Ethics statements</h2><div class="section" id="sec-23"><h3>Patient consent for publication</h3><p id="p-56" class="ethics-consent-to-publish">Not applicable.</p></div><div class="section" id="sec-24"><h3>Ethics approval</h3><p id="p-57" class="ethics-approval">Not applicable.</p></div></div><div class="section ack" id="ack-1"><h2>Acknowledgments</h2><p id="p-47">We thank members of the Guo laboratory for critical reading and kind suggestion of the manuscript.</p></div><div class="section ref-list" id="ref-list-1"><h2 class="">References</h2><ol class="cit-list"><li><a class="rev-xref-ref" href="#xref-ref-1-1" title="View reference 1 in text" id="ref-1">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.1"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Shah</span> <span class="cit-name-given-names">UA</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Iyengar</span> <span class="cit-name-given-names">NM</span> </span> </li></ol><cite>. <span class="cit-article-title">Plant-based and ketogenic diets as diverging paths to address cancer: a review</span>. <abbr class="cit-jnl-abbrev">JAMA Oncol</abbr> <span class="cit-pub-date">2022</span>;<span class="cit-vol">8</span>:<span class="cit-fpage">1201</span>–<span class="cit-lpage">8</span>. <a href="http://dx.doi.org/10.1001/jamaoncol.2022.1769">doi:10.1001/jamaoncol.2022.1769</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DJAMA%2BOncol%26rft.volume%253D8%26rft.spage%253D1201%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-2-1" title="View reference 2 in text" id="ref-2">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.2"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Pascual</span> <span class="cit-name-given-names">G</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Domínguez</span> <span class="cit-name-given-names">D</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Elosúa-Bayes</span> <span class="cit-name-given-names">M</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Dietary palmitic acid promotes a prometastatic memory via schwann cells</span>. <abbr class="cit-jnl-abbrev">Nature</abbr> <span class="cit-pub-date">2021</span>;<span class="cit-vol">599</span>:<span class="cit-fpage">485</span>–<span class="cit-lpage">90</span>. <a href="http://dx.doi.org/10.1038/s41586-021-04075-0">doi:10.1038/s41586-021-04075-0</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DNature%26rft.volume%253D599%26rft.spage%253D485%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-2-1" title="View reference 3 in text" id="ref-3">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.3" data-doi="10.1053/J.GASTRO.2021.08.041"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Yang</span> <span class="cit-name-given-names">J</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Wei</span> <span class="cit-name-given-names">H</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Zhou</span> <span class="cit-name-given-names">Y</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">High-fat diet promotes colorectal tumorigenesis through Modulating gut Microbiota and metabolites</span>. <abbr class="cit-jnl-abbrev">Gastroenterology</abbr> <span class="cit-pub-date">2022</span>;<span class="cit-vol">162</span>:<span class="cit-fpage">135</span>–<span class="cit-lpage">149</span>.<span class="cit-elocation-id">S0016-5085(21)03439-9</span>. <a href="http://dx.doi.org/10.1053/j.gastro.2021.08.041">doi:10.1053/j.gastro.2021.08.041</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DGastroenterology%26rft.volume%253D162%26rft.spage%253D135%26rft_id%253Dinfo%253Adoi%252F10.1053%252FJ.GASTRO.2021.08.041%26rft_id%253Dinfo%253Apmid%252Fhttp%253A%252F%252Fwww.n%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1053/J.GASTRO.2021.08.041&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a><a href="/lookup/external-ref?access_num=http://www.n&link_type=MED&atom=%2Fgutjnl%2F73%2F7%2F1156.atom" class="cit-ref-sprinkles cit-ref-sprinkles-medline"><span>PubMed</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-4-1" title="View reference 4 in text" id="ref-4">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.4"><div class="cit-metadata"><ol class="cit-auth-list"><li><span class="cit-auth cit-collab">The Lancet Diabetes</span></li></ol><cite>. <span class="cit-article-title">The obesity-cancer link: of increasing concern</span>. <abbr class="cit-jnl-abbrev">Lancet Diabetes Endocrinol</abbr> <span class="cit-pub-date">2020</span>;<span class="cit-vol">8</span>:<span class="cit-fpage">175</span>. <a href="http://dx.doi.org/10.1016/S2213-8587(20)30031-0">doi:10.1016/S2213-8587(20)30031-0</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DLancet%2BDiabetes%2BEndocrinol%26rft.volume%253D8%26rft.spage%253D175%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-5-1" title="View reference 5 in text" id="ref-5">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.5" data-doi="10.1038/nature23887"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Chen</span> <span class="cit-name-given-names">S</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Zhu</span> <span class="cit-name-given-names">B</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Yin</span> <span class="cit-name-given-names">C</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Palmitoylation-dependent activation of Mc1R prevents melanomagenesis</span>. <abbr class="cit-jnl-abbrev">Nature</abbr> <span class="cit-pub-date">2017</span>;<span class="cit-vol">549</span>:<span class="cit-fpage">399</span>–<span class="cit-lpage">403</span>. <a href="http://dx.doi.org/10.1038/nature23887">doi:10.1038/nature23887</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DNature%26rft.volume%253D549%26rft.spage%253D399%26rft_id%253Dinfo%253Adoi%252F10.1038%252Fnature23887%26rft_id%253Dinfo%253Apmid%252F28869973%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1038/nature23887&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a><a href="/lookup/external-ref?access_num=28869973&link_type=MED&atom=%2Fgutjnl%2F73%2F7%2F1156.atom" class="cit-ref-sprinkles cit-ref-sprinkles-medline"><span>PubMed</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-6-1" title="View reference 6 in text" id="ref-6">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.6" data-doi="10.1038/s41586-020-2799-2"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Zhang</span> <span class="cit-name-given-names">M</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Zhou</span> <span class="cit-name-given-names">L</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Xu</span> <span class="cit-name-given-names">Y</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">A Stat3 palmitoylation cycle promotes Th17 differentiation and colitis</span>. <abbr class="cit-jnl-abbrev">Nature</abbr> <span class="cit-pub-date">2020</span>;<span class="cit-vol">586</span>:<span class="cit-fpage">434</span>–<span class="cit-lpage">9</span>. <a href="http://dx.doi.org/10.1038/s41586-020-2799-2">doi:10.1038/s41586-020-2799-2</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DNature%26rft.volume%253D586%26rft.spage%253D434%26rft_id%253Dinfo%253Adoi%252F10.1038%252Fs41586-020-2799-2%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1038/s41586-020-2799-2&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-7-1" title="View reference 7 in text" id="ref-7">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.7" data-doi="10.1126/science.1105654"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Rocks</span> <span class="cit-name-given-names">O</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Peyker</span> <span class="cit-name-given-names">A</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Kahms</span> <span class="cit-name-given-names">M</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">An acylation cycle regulates localization and activity of palmitoylated Ras Isoforms</span>. <abbr class="cit-jnl-abbrev">Science</abbr> <span class="cit-pub-date">2005</span>;<span class="cit-vol">307</span>:<span class="cit-fpage">1746</span>–<span class="cit-lpage">52</span>. <a href="http://dx.doi.org/10.1126/science.1105654">doi:10.1126/science.1105654</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DScience%26rft_id%253Dinfo%253Adoi%252F10.1126%252Fscience.1105654%26rft_id%253Dinfo%253Apmid%252F15705808%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/ijlink/YTozOntzOjQ6InBhdGgiO3M6MTQ6Ii9sb29rdXAvaWpsaW5rIjtzOjU6InF1ZXJ5IjthOjQ6e3M6ODoibGlua1R5cGUiO3M6NDoiQUJTVCI7czoxMToiam91cm5hbENvZGUiO3M6Mzoic2NpIjtzOjU6InJlc2lkIjtzOjEzOiIzMDcvNTcxNi8xNzQ2IjtzOjQ6ImF0b20iO3M6MjI6Ii9ndXRqbmwvNzMvNy8xMTU2LmF0b20iO31zOjg6ImZyYWdtZW50IjtzOjA6IiI7fQ==" class="cit-ref-sprinkles cit-ref-sprinkles-ijlink"><span><span class="cit-reflinks-abstract">Abstract</span><span class="cit-sep cit-reflinks-variant-name-sep">/</span><span class="cit-reflinks-full-text"><span class="free-full-text">FREE </span>Full Text</span></span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-8-1" title="View reference 8 in text" id="ref-8">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.8" data-doi="10.1038/s41575-019-0145-7"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Anstee</span> <span class="cit-name-given-names">QM</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Reeves</span> <span class="cit-name-given-names">HL</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Kotsiliti</span> <span class="cit-name-given-names">E</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">From NASH to HCC: Current concepts and future challenges</span>. <abbr class="cit-jnl-abbrev">Nat Rev Gastroenterol Hepatol</abbr> <span class="cit-pub-date">2019</span>;<span class="cit-vol">16</span>:<span class="cit-fpage">411</span>–<span class="cit-lpage">28</span>. <a href="http://dx.doi.org/10.1038/s41575-019-0145-7">doi:10.1038/s41575-019-0145-7</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DNat%2BRev%2BGastroenterol%2BHepatol%26rft.volume%253D16%26rft.spage%253D411%26rft_id%253Dinfo%253Adoi%252F10.1038%252Fs41575-019-0145-7%26rft_id%253Dinfo%253Apmid%252F31028350%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1038/s41575-019-0145-7&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a><a href="/lookup/external-ref?access_num=31028350&link_type=MED&atom=%2Fgutjnl%2F73%2F7%2F1156.atom" class="cit-ref-sprinkles cit-ref-sprinkles-medline"><span>PubMed</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-9-1" title="View reference 9 in text" id="ref-9">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.9" data-doi="10.1016/j.jhep.2020.03.023"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Wu</span> <span class="cit-name-given-names">X</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Poulsen</span> <span class="cit-name-given-names">KL</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Sanz-Garcia</span> <span class="cit-name-given-names">C</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">MLKL-dependent signaling regulates autophagic flux in a murine model of non-alcohol-associated fatty liver and steatohepatitis</span>. <abbr class="cit-jnl-abbrev">J Hepatol</abbr> <span class="cit-pub-date">2020</span>;<span class="cit-vol">73</span>:<span class="cit-fpage">616</span>–<span class="cit-lpage">27</span>. <a href="http://dx.doi.org/10.1016/j.jhep.2020.03.023">doi:10.1016/j.jhep.2020.03.023</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DJ%2BHepatol%26rft.volume%253D73%26rft.spage%253D616%26rft_id%253Dinfo%253Adoi%252F10.1016%252Fj.jhep.2020.03.023%26rft_id%253Dinfo%253Apmid%252F32220583%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1016/j.jhep.2020.03.023&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a><a href="/lookup/external-ref?access_num=32220583&link_type=MED&atom=%2Fgutjnl%2F73%2F7%2F1156.atom" class="cit-ref-sprinkles cit-ref-sprinkles-medline"><span>PubMed</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-9-1" title="View reference 10 in text" id="ref-10">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.10" data-doi="10.1038/nrgastro.2015.200"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Heymann</span> <span class="cit-name-given-names">F</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Tacke</span> <span class="cit-name-given-names">F</span> </span> </li></ol><cite>. <span class="cit-article-title">Immunology in the liver--from homeostasis to disease</span>. <abbr class="cit-jnl-abbrev">Nat Rev Gastroenterol Hepatol</abbr> <span class="cit-pub-date">2016</span>;<span class="cit-vol">13</span>:<span class="cit-fpage">88</span>–<span class="cit-lpage">110</span>. <a href="http://dx.doi.org/10.1038/nrgastro.2015.200">doi:10.1038/nrgastro.2015.200</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DNat%2BRev%2BGastroenterol%2BHepatol%26rft.volume%253D13%26rft.spage%253D88%26rft_id%253Dinfo%253Adoi%252F10.1038%252Fnrgastro.2015.200%26rft_id%253Dinfo%253Apmid%252F26758786%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1038/nrgastro.2015.200&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a><a href="/lookup/external-ref?access_num=26758786&link_type=MED&atom=%2Fgutjnl%2F73%2F7%2F1156.atom" class="cit-ref-sprinkles cit-ref-sprinkles-medline"><span>PubMed</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-11-1" title="View reference 11 in text" id="ref-11">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.11" data-doi="10.1053/j.gastro.2010.12.006"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Calvisi</span> <span class="cit-name-given-names">DF</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Wang</span> <span class="cit-name-given-names">C</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Ho</span> <span class="cit-name-given-names">C</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Increased Lipogenesis, induced by AKT-Mtorc1-Rps6 signaling, promotes development of human hepatocellular carcinoma</span>. <abbr class="cit-jnl-abbrev">Gastroenterology</abbr> <span class="cit-pub-date">2011</span>;<span class="cit-vol">140</span>:<span class="cit-fpage">1071</span>–<span class="cit-lpage">83</span>. <a href="http://dx.doi.org/10.1053/j.gastro.2010.12.006">doi:10.1053/j.gastro.2010.12.006</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DGastroenterology%26rft.volume%253D140%26rft.spage%253D1071%26rft_id%253Dinfo%253Adoi%252F10.1053%252Fj.gastro.2010.12.006%26rft_id%253Dinfo%253Apmid%252F21147110%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1053/j.gastro.2010.12.006&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a><a href="/lookup/external-ref?access_num=21147110&link_type=MED&atom=%2Fgutjnl%2F73%2F7%2F1156.atom" class="cit-ref-sprinkles cit-ref-sprinkles-medline"><span>PubMed</span></a><a href="/lookup/external-ref?access_num=000288014700047&link_type=ISI" class="cit-ref-sprinkles cit-ref-sprinkles-newisilink cit-ref-sprinkles-webofscience"><span>Web of Science</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-12-1" title="View reference 12 in text" id="ref-12">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.12"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Vanhaesebroeck</span> <span class="cit-name-given-names">B</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Alessi</span> <span class="cit-name-given-names">DR</span> </span> </li></ol><cite>. <span class="cit-article-title">The Pi3K-Pdk1 connection: more than just a road to PKB</span>. <abbr class="cit-jnl-abbrev">Biochem J</abbr> <span class="cit-pub-date">2000</span>;<span class="cit-vol">346 Pt 3</span>:<span class="cit-fpage">561</span>–<span class="cit-lpage">76</span>.</cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DBiochem%2BJ%26rft.volume%253D3463%26rft.spage%253D561%26rft_id%253Dinfo%253Apmid%252Fhttp%253A%252F%252Fwww.n%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=http://www.n&link_type=MED&atom=%2Fgutjnl%2F73%2F7%2F1156.atom" class="cit-ref-sprinkles cit-ref-sprinkles-medline"><span>PubMed</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-12-1" title="View reference 13 in text" id="ref-13">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.13" data-doi="10.1016/j.cell.2017.04.001"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Manning</span> <span class="cit-name-given-names">BD</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Toker</span> <span class="cit-name-given-names">A</span> </span> </li></ol><cite>. <span class="cit-article-title">AKT/PKB signaling: navigating the network</span>. <abbr class="cit-jnl-abbrev">Cell</abbr> <span class="cit-pub-date">2017</span>;<span class="cit-vol">169</span>:<span class="cit-fpage">381</span>–<span class="cit-lpage">405</span>. <a href="http://dx.doi.org/10.1016/j.cell.2017.04.001">doi:10.1016/j.cell.2017.04.001</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DCell%26rft.volume%253D169%26rft.spage%253D381%26rft_id%253Dinfo%253Adoi%252F10.1016%252Fj.cell.2017.04.001%26rft_id%253Dinfo%253Apmid%252F28431241%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1016/j.cell.2017.04.001&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a><a href="/lookup/external-ref?access_num=28431241&link_type=MED&atom=%2Fgutjnl%2F73%2F7%2F1156.atom" class="cit-ref-sprinkles cit-ref-sprinkles-medline"><span>PubMed</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-14-1" title="View reference 14 in text" id="ref-14">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.14" data-doi="10.1038/nature13079"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Liu</span> <span class="cit-name-given-names">P</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Begley</span> <span class="cit-name-given-names">M</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Michowski</span> <span class="cit-name-given-names">W</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Cell-cycle-regulated activation of AKT kinase by phosphorylation at its Carboxyl terminus</span>. <abbr class="cit-jnl-abbrev">Nature</abbr> <span class="cit-pub-date">2014</span>;<span class="cit-vol">508</span>:<span class="cit-fpage">541</span>–<span class="cit-lpage">5</span>. <a href="http://dx.doi.org/10.1038/nature13079">doi:10.1038/nature13079</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DNature%26rft.volume%253D508%26rft.spage%253D541%26rft_id%253Dinfo%253Adoi%252F10.1038%252Fnature13079%26rft_id%253Dinfo%253Apmid%252F24670654%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1038/nature13079&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a><a href="/lookup/external-ref?access_num=24670654&link_type=MED&atom=%2Fgutjnl%2F73%2F7%2F1156.atom" class="cit-ref-sprinkles cit-ref-sprinkles-medline"><span>PubMed</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-14-1" title="View reference 15 in text" id="ref-15">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.15" data-doi="10.1126/science.aad5755"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Guo</span> <span class="cit-name-given-names">J</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Chakraborty</span> <span class="cit-name-given-names">AA</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Liu</span> <span class="cit-name-given-names">P</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Wei, pVHL suppresses kinase activity of AKT in a proline-hydroxylation-dependent manner</span>. <abbr class="cit-jnl-abbrev">Science</abbr> <span class="cit-pub-date">2016</span>;<span class="cit-vol">353</span>:<span class="cit-fpage">929</span>–<span class="cit-lpage">32</span>. <a href="http://dx.doi.org/10.1126/science.aad5755">doi:10.1126/science.aad5755</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DScience%26rft.stitle%253DScience%26rft.aulast%253DGuo%26rft.auinit1%253DJ.%26rft.volume%253D353%26rft.issue%253D6302%26rft.spage%253D929%26rft.epage%253D932%26rft.atitle%253DpVHL%2Bsuppresses%2Bkinase%2Bactivity%2Bof%2BAkt%2Bin%2Ba%2Bproline-hydroxylation-dependent%2Bmanner%26rft_id%253Dinfo%253Adoi%252F10.1126%252Fscience.aad5755%26rft_id%253Dinfo%253Apmid%252F27563096%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/ijlink/YTozOntzOjQ6InBhdGgiO3M6MTQ6Ii9sb29rdXAvaWpsaW5rIjtzOjU6InF1ZXJ5IjthOjQ6e3M6ODoibGlua1R5cGUiO3M6NDoiQUJTVCI7czoxMToiam91cm5hbENvZGUiO3M6Mzoic2NpIjtzOjU6InJlc2lkIjtzOjEyOiIzNTMvNjMwMi85MjkiO3M6NDoiYXRvbSI7czoyMjoiL2d1dGpubC83My83LzExNTYuYXRvbSI7fXM6ODoiZnJhZ21lbnQiO3M6MDoiIjt9" class="cit-ref-sprinkles cit-ref-sprinkles-ijlink"><span><span class="cit-reflinks-abstract">Abstract</span><span class="cit-sep cit-reflinks-variant-name-sep">/</span><span class="cit-reflinks-full-text"><span class="free-full-text">FREE </span>Full Text</span></span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-16-1" title="View reference 16 in text" id="ref-16">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.16" data-doi="10.1126/science.279.5351.710"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Stephens</span> <span class="cit-name-given-names">L</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Anderson</span> <span class="cit-name-given-names">K</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Stokoe</span> <span class="cit-name-given-names">D</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Protein kinase B Kinases that mediate Phosphatidylinositol 3,4,5-Trisphosphate-dependent activation of protein kinase B</span>. <abbr class="cit-jnl-abbrev">Science</abbr> <span class="cit-pub-date">1998</span>;<span class="cit-vol">279</span>:<span class="cit-fpage">710</span>–<span class="cit-lpage">4</span>. <a href="http://dx.doi.org/10.1126/science.279.5351.710">doi:10.1126/science.279.5351.710</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DScience%26rft.stitle%253DScience%26rft.aulast%253DStephens%26rft.auinit1%253DL.%26rft.volume%253D279%26rft.issue%253D5351%26rft.spage%253D710%26rft.epage%253D714%26rft.atitle%253DProtein%2BKinase%2BB%2BKinases%2BThat%2BMediate%2BPhosphatidylinositol%2B3%252C4%252C5-Trisphosphate-Dependent%2BActivation%2Bof%2BProtein%2BKinase%2BB%26rft_id%253Dinfo%253Adoi%252F10.1126%252Fscience.279.5351.710%26rft_id%253Dinfo%253Apmid%252F9445477%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/ijlink/YTozOntzOjQ6InBhdGgiO3M6MTQ6Ii9sb29rdXAvaWpsaW5rIjtzOjU6InF1ZXJ5IjthOjQ6e3M6ODoibGlua1R5cGUiO3M6NDoiQUJTVCI7czoxMToiam91cm5hbENvZGUiO3M6Mzoic2NpIjtzOjU6InJlc2lkIjtzOjEyOiIyNzkvNTM1MS83MTAiO3M6NDoiYXRvbSI7czoyMjoiL2d1dGpubC83My83LzExNTYuYXRvbSI7fXM6ODoiZnJhZ21lbnQiO3M6MDoiIjt9" class="cit-ref-sprinkles cit-ref-sprinkles-ijlink"><span><span class="cit-reflinks-abstract">Abstract</span><span class="cit-sep cit-reflinks-variant-name-sep">/</span><span class="cit-reflinks-full-text"><span class="free-full-text">FREE </span>Full Text</span></span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-16-1" title="View reference 17 in text" id="ref-17">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.17" data-doi="10.1006/excr.1999.4701"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Vanhaesebroeck</span> <span class="cit-name-given-names">B</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Waterfield</span> <span class="cit-name-given-names">MD</span> </span> </li></ol><cite>. <span class="cit-article-title">Signaling by distinct classes of Phosphoinositide 3-Kinases</span>. <abbr class="cit-jnl-abbrev">Exp Cell Res</abbr> <span class="cit-pub-date">1999</span>;<span class="cit-vol">253</span>:<span class="cit-fpage">239</span>–<span class="cit-lpage">54</span>. <a href="http://dx.doi.org/10.1006/excr.1999.4701">doi:10.1006/excr.1999.4701</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DExperimental%2Bcell%2Bresearch%26rft.stitle%253DExp%2BCell%2BRes%26rft.aulast%253DVanhaesebroeck%26rft.auinit1%253DB.%26rft.volume%253D253%26rft.issue%253D1%26rft.spage%253D239%26rft.epage%253D254%26rft.atitle%253DSignaling%2Bby%2Bdistinct%2Bclasses%2Bof%2Bphosphoinositide%2B3-kinases.%26rft_id%253Dinfo%253Adoi%252F10.1006%252Fexcr.1999.4701%26rft_id%253Dinfo%253Apmid%252F10579926%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1006/excr.1999.4701&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a><a href="/lookup/external-ref?access_num=10579926&link_type=MED&atom=%2Fgutjnl%2F73%2F7%2F1156.atom" class="cit-ref-sprinkles cit-ref-sprinkles-medline"><span>PubMed</span></a><a href="/lookup/external-ref?access_num=000083999600022&link_type=ISI" class="cit-ref-sprinkles cit-ref-sprinkles-newisilink cit-ref-sprinkles-webofscience"><span>Web of Science</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-18-1" title="View reference 18 in text" id="ref-18">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.18" data-doi="10.1126/science.1175065"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Yang</span> <span class="cit-name-given-names">W-L</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Wang</span> <span class="cit-name-given-names">J</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Chan</span> <span class="cit-name-given-names">C-H</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">The E3 Ligase Traf6 regulates AKT Ubiquitination and activation</span>. <abbr class="cit-jnl-abbrev">Science</abbr> <span class="cit-pub-date">2009</span>;<span class="cit-vol">325</span>:<span class="cit-fpage">1134</span>–<span class="cit-lpage">8</span>. <a href="http://dx.doi.org/10.1126/science.1175065">doi:10.1126/science.1175065</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DScience%26rft.stitle%253DScience%26rft.aulast%253DYang%26rft.auinit1%253DW.-L.%26rft.volume%253D325%26rft.issue%253D5944%26rft.spage%253D1134%26rft.epage%253D1138%26rft.atitle%253DThe%2BE3%2BLigase%2BTRAF6%2BRegulates%2BAkt%2BUbiquitination%2Band%2BActivation%26rft_id%253Dinfo%253Adoi%252F10.1126%252Fscience.1175065%26rft_id%253Dinfo%253Apmid%252F19713527%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/ijlink/YTozOntzOjQ6InBhdGgiO3M6MTQ6Ii9sb29rdXAvaWpsaW5rIjtzOjU6InF1ZXJ5IjthOjQ6e3M6ODoibGlua1R5cGUiO3M6NDoiQUJTVCI7czoxMToiam91cm5hbENvZGUiO3M6Mzoic2NpIjtzOjU6InJlc2lkIjtzOjEzOiIzMjUvNTk0NC8xMTM0IjtzOjQ6ImF0b20iO3M6MjI6Ii9ndXRqbmwvNzMvNy8xMTU2LmF0b20iO31zOjg6ImZyYWdtZW50IjtzOjA6IiI7fQ==" class="cit-ref-sprinkles cit-ref-sprinkles-ijlink"><span><span class="cit-reflinks-abstract">Abstract</span><span class="cit-sep cit-reflinks-variant-name-sep">/</span><span class="cit-reflinks-full-text"><span class="free-full-text">FREE </span>Full Text</span></span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-18-1" title="View reference 19 in text" id="ref-19">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.19" data-doi="10.1038/s41556-018-0261-6"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Guo</span> <span class="cit-name-given-names">J</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Dai</span> <span class="cit-name-given-names">X</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Laurent</span> <span class="cit-name-given-names">B</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">AKT methylation by Setdb1 promotes AKT kinase activity and Oncogenic functions</span>. <abbr class="cit-jnl-abbrev">Nat Cell Biol</abbr> <span class="cit-pub-date">2019</span>;<span class="cit-vol">21</span>:<span class="cit-fpage">226</span>–<span class="cit-lpage">37</span>. <a href="http://dx.doi.org/10.1038/s41556-018-0261-6">doi:10.1038/s41556-018-0261-6</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DNat%2BCell%2BBiol%26rft.volume%253D21%26rft.spage%253D226%26rft_id%253Dinfo%253Adoi%252F10.1038%252Fs41556-018-0261-6%26rft_id%253Dinfo%253Apmid%252F30692625%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1038/s41556-018-0261-6&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a><a href="/lookup/external-ref?access_num=30692625&link_type=MED&atom=%2Fgutjnl%2F73%2F7%2F1156.atom" class="cit-ref-sprinkles cit-ref-sprinkles-medline"><span>PubMed</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-18-1" title="View reference 20 in text" id="ref-20">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.20"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Sundaresan</span> <span class="cit-name-given-names">NR</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Pillai</span> <span class="cit-name-given-names">VB</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Wolfgeher</span> <span class="cit-name-given-names">D</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">The Deacetylase Sirt1 promotes membrane localization and activation of AKT and Pdk1 during tumorigenesis and cardiac hypertrophy</span>. <abbr class="cit-jnl-abbrev">Sci Signal</abbr> <span class="cit-pub-date">2011</span>;<span class="cit-vol">4</span>:<span class="cit-elocation-id">ra46</span>. <a href="http://dx.doi.org/10.1126/scisignal.2001465">doi:10.1126/scisignal.2001465</a> </cite></div><div class="cit-extra"></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-21-1" title="View reference 21 in text" id="ref-21">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.21" data-doi="10.1038/nature14665"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Van Keymeulen</span> <span class="cit-name-given-names">A</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Lee</span> <span class="cit-name-given-names">MY</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Ousset</span> <span class="cit-name-given-names">M</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Reactivation of multipotency by Oncogenic Pik3Ca induces breast tumour heterogeneity</span>. <abbr class="cit-jnl-abbrev">Nature</abbr> <span class="cit-pub-date">2015</span>;<span class="cit-vol">525</span>:<span class="cit-fpage">119</span>–<span class="cit-lpage">23</span>. <a href="http://dx.doi.org/10.1038/nature14665">doi:10.1038/nature14665</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DNature%26rft.volume%253D525%26rft.spage%253D119%26rft_id%253Dinfo%253Adoi%252F10.1038%252Fnature14665%26rft_id%253Dinfo%253Apmid%252F26266985%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1038/nature14665&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a><a href="/lookup/external-ref?access_num=26266985&link_type=MED&atom=%2Fgutjnl%2F73%2F7%2F1156.atom" class="cit-ref-sprinkles cit-ref-sprinkles-medline"><span>PubMed</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-22-1" title="View reference 22 in text" id="ref-22">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.22" data-doi="10.1038/s41556-018-0200-6"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Mehta</span> <span class="cit-name-given-names">S</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Zhang</span> <span class="cit-name-given-names">Y</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Roth</span> <span class="cit-name-given-names">RH</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Single-Fluorophore biosensors for sensitive and multiplexed detection of signalling activities</span>. <abbr class="cit-jnl-abbrev">Nat Cell Biol</abbr> <span class="cit-pub-date">2018</span>;<span class="cit-vol">20</span>:<span class="cit-fpage">1215</span>–<span class="cit-lpage">25</span>. <a href="http://dx.doi.org/10.1038/s41556-018-0200-6">doi:10.1038/s41556-018-0200-6</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DNat%2BCell%2BBiol%26rft.volume%253D20%26rft.spage%253D1215%26rft_id%253Dinfo%253Adoi%252F10.1038%252Fs41556-018-0200-6%26rft_id%253Dinfo%253Apmid%252F30250062%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1038/s41556-018-0200-6&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a><a href="/lookup/external-ref?access_num=30250062&link_type=MED&atom=%2Fgutjnl%2F73%2F7%2F1156.atom" class="cit-ref-sprinkles cit-ref-sprinkles-medline"><span>PubMed</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-22-1" title="View reference 23 in text" id="ref-23">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.23" data-doi="10.1021/ac020731c"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Paulmurugan</span> <span class="cit-name-given-names">R</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Gambhir</span> <span class="cit-name-given-names">SS</span> </span> </li></ol><cite>. <span class="cit-article-title">Monitoring protein-protein interactions using split synthetic Renilla Luciferase protein-fragment-assisted complementation</span>. <abbr class="cit-jnl-abbrev">Anal Chem</abbr> <span class="cit-pub-date">2003</span>;<span class="cit-vol">75</span>:<span class="cit-fpage">1584</span>–<span class="cit-lpage">9</span>. <a href="http://dx.doi.org/10.1021/ac020731c">doi:10.1021/ac020731c</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DAnalytical%2BChemistry%2B%2528Washington%252C%2BDC%2529%26rft.stitle%253DAnalytical%2BChemistry%2B%2528Washington%252C%2BDC%2529%26rft.aulast%253DPaulmurugan%26rft.auinit1%253DR.%26rft.volume%253D75%26rft.issue%253D7%26rft.spage%253D1584%26rft.epage%253D1589%26rft.atitle%253DMonitoring%2Bprotein-protein%2Binteractions%2Busing%2Bsplit%2Bsynthetic%2Brenilla%2Bluciferase%2Bprotein-fragment-assisted%2Bcomplementation.%26rft_id%253Dinfo%253Adoi%252F10.1021%252Fac020731c%26rft_id%253Dinfo%253Apmid%252F12705589%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1021/ac020731c&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a><a href="/lookup/external-ref?access_num=12705589&link_type=MED&atom=%2Fgutjnl%2F73%2F7%2F1156.atom" class="cit-ref-sprinkles cit-ref-sprinkles-medline"><span>PubMed</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-11-2" title="View reference 24 in text" id="ref-24">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.24"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Ho</span> <span class="cit-name-given-names">C</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Wang</span> <span class="cit-name-given-names">C</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Mattu</span> <span class="cit-name-given-names">S</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">AKT (V-AKT murine Thymoma viral Oncogene Homolog 1) and N-Ras (neuroblastoma Ras viral Oncogene Homolog) Coactivation in the Mouse liver promotes rapid carcinogenesis by way of mTOR</span>. <span class="cit-pub-date">2012</span>;<span class="cit-vol">55</span>:<span class="cit-fpage">833</span>–<span class="cit-lpage">45</span>. <a href="http://dx.doi.org/10.1002/hep.24736">doi:10.1002/hep.24736</a> </cite></div><div class="cit-extra"></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-25-1" title="View reference 25 in text" id="ref-25">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.25" data-doi="10.1136/gutjnl-2019-319664"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Zhang</span> <span class="cit-name-given-names">X</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Coker</span> <span class="cit-name-given-names">OO</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Chu</span> <span class="cit-name-given-names">ES</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Dietary cholesterol drives fatty liver-associated liver cancer by Modulating gut Microbiota and metabolites</span>. <abbr class="cit-jnl-abbrev">Gut</abbr> <span class="cit-pub-date">2021</span>;<span class="cit-vol">70</span>:<span class="cit-fpage">761</span>–<span class="cit-lpage">74</span>. <a href="http://dx.doi.org/10.1136/gutjnl-2019-319664">doi:10.1136/gutjnl-2019-319664</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DGut%26rft_id%253Dinfo%253Adoi%252F10.1136%252Fgutjnl-2019-319664%26rft_id%253Dinfo%253Apmid%252F32694178%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/ijlink/YTozOntzOjQ6InBhdGgiO3M6MTQ6Ii9sb29rdXAvaWpsaW5rIjtzOjU6InF1ZXJ5IjthOjQ6e3M6ODoibGlua1R5cGUiO3M6NDoiQUJTVCI7czoxMToiam91cm5hbENvZGUiO3M6NjoiZ3V0am5sIjtzOjU6InJlc2lkIjtzOjg6IjcwLzQvNzYxIjtzOjQ6ImF0b20iO3M6MjI6Ii9ndXRqbmwvNzMvNy8xMTU2LmF0b20iO31zOjg6ImZyYWdtZW50IjtzOjA6IiI7fQ==" class="cit-ref-sprinkles cit-ref-sprinkles-ijlink"><span><span class="cit-reflinks-abstract">Abstract</span><span class="cit-sep cit-reflinks-variant-name-sep">/</span><span class="cit-reflinks-full-text"><span class="free-full-text">FREE </span>Full Text</span></span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-26-1" title="View reference 26 in text" id="ref-26">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.26"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Pan</span> <span class="cit-name-given-names">J</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Fan</span> <span class="cit-name-given-names">Z</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Wang</span> <span class="cit-name-given-names">Z</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Cd36 mediates Palmitate acid-induced metastasis of gastric cancer via AKT/GSK-3Beta/beta-Catenin pathway</span>. <abbr class="cit-jnl-abbrev">J Exp Clin Cancer Res</abbr> <span class="cit-pub-date">2019</span>;<span class="cit-vol">38</span>:<span class="cit-elocation-id">52</span>. <a href="http://dx.doi.org/10.1186/s13046-019-1049-7">doi:10.1186/s13046-019-1049-7</a> </cite></div><div class="cit-extra"></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-27-1" title="View reference 27 in text" id="ref-27">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.27"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Blaustein</span> <span class="cit-name-given-names">M</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Piegari</span> <span class="cit-name-given-names">E</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Martínez Calejman</span> <span class="cit-name-given-names">C</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Akt is S-Palmitoylated: A new layer of regulation for AKT</span>. <abbr class="cit-jnl-abbrev">Front Cell Dev Biol</abbr> <span class="cit-pub-date">2021</span>;<span class="cit-vol">9</span>:<span class="cit-elocation-id">626404</span>. <a href="http://dx.doi.org/10.3389/fcell.2021.626404">doi:10.3389/fcell.2021.626404</a> </cite></div><div class="cit-extra"></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-28-1" title="View reference 28 in text" id="ref-28">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.28" data-doi="10.1101/gad.913901"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Chen</span> <span class="cit-name-given-names">WS</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Xu</span> <span class="cit-name-given-names">PZ</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Gottlob</span> <span class="cit-name-given-names">K</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Growth retardation and increased apoptosis in mice with homozygous disruption of the Akt1 gene</span>. <abbr class="cit-jnl-abbrev">Genes Dev</abbr> <span class="cit-pub-date">2001</span>;<span class="cit-vol">15</span>:<span class="cit-fpage">2203</span>–<span class="cit-lpage">8</span>. <a href="http://dx.doi.org/10.1101/gad.913901">doi:10.1101/gad.913901</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DGenes%2B%2526%2BDevelopment%26rft.stitle%253DGenes%2BDev.%26rft.aulast%253DChen%26rft.auinit1%253DW.%2BS.%26rft.volume%253D15%26rft.issue%253D17%26rft.spage%253D2203%26rft.epage%253D2208%26rft.atitle%253DGrowth%2Bretardation%2Band%2Bincreased%2Bapoptosis%2Bin%2Bmice%2Bwith%2Bhomozygous%2Bdisruption%2Bof%2Bthe%2Bakt1%2Bgene%26rft_id%253Dinfo%253Adoi%252F10.1101%252Fgad.913901%26rft_id%253Dinfo%253Apmid%252F11544177%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/ijlink/YTozOntzOjQ6InBhdGgiO3M6MTQ6Ii9sb29rdXAvaWpsaW5rIjtzOjU6InF1ZXJ5IjthOjQ6e3M6ODoibGlua1R5cGUiO3M6NDoiQUJTVCI7czoxMToiam91cm5hbENvZGUiO3M6ODoiZ2VuZXNkZXYiO3M6NToicmVzaWQiO3M6MTA6IjE1LzE3LzIyMDMiO3M6NDoiYXRvbSI7czoyMjoiL2d1dGpubC83My83LzExNTYuYXRvbSI7fXM6ODoiZnJhZ21lbnQiO3M6MDoiIjt9" class="cit-ref-sprinkles cit-ref-sprinkles-ijlink"><span><span class="cit-reflinks-abstract">Abstract</span><span class="cit-sep cit-reflinks-variant-name-sep">/</span><span class="cit-reflinks-full-text"><span class="free-full-text">FREE </span>Full Text</span></span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-28-1" title="View reference 29 in text" id="ref-29">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.29" data-doi="10.1074/jbc.C100462200"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Cho</span> <span class="cit-name-given-names">H</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Thorvaldsen</span> <span class="cit-name-given-names">JL</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Chu</span> <span class="cit-name-given-names">Q</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Akt1/Pkbalpha is required for normal growth but Dispensable for maintenance of glucose homeostasis in mice</span>. <abbr class="cit-jnl-abbrev">J Biol Chem</abbr> <span class="cit-pub-date">2001</span>;<span class="cit-vol">276</span>:<span class="cit-fpage">38349</span>–<span class="cit-lpage">52</span>. <a href="http://dx.doi.org/10.1074/jbc.C100462200">doi:10.1074/jbc.C100462200</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DJ%2BBiol%2BChem%26rft_id%253Dinfo%253Adoi%252F10.1074%252Fjbc.C100462200%26rft_id%253Dinfo%253Apmid%252F11533044%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/ijlink/YTozOntzOjQ6InBhdGgiO3M6MTQ6Ii9sb29rdXAvaWpsaW5rIjtzOjU6InF1ZXJ5IjthOjQ6e3M6ODoibGlua1R5cGUiO3M6NDoiQUJTVCI7czoxMToiam91cm5hbENvZGUiO3M6MzoiamJjIjtzOjU6InJlc2lkIjtzOjEyOiIyNzYvNDIvMzgzNDkiO3M6NDoiYXRvbSI7czoyMjoiL2d1dGpubC83My83LzExNTYuYXRvbSI7fXM6ODoiZnJhZ21lbnQiO3M6MDoiIjt9" class="cit-ref-sprinkles cit-ref-sprinkles-ijlink"><span><span class="cit-reflinks-abstract">Abstract</span><span class="cit-sep cit-reflinks-variant-name-sep">/</span><span class="cit-reflinks-full-text"><span class="free-full-text">FREE </span>Full Text</span></span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-5-2" title="View reference 30 in text" id="ref-30">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.30"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Niu</span> <span class="cit-name-given-names">J</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Sun</span> <span class="cit-name-given-names">Y</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Chen</span> <span class="cit-name-given-names">B</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Fatty acids and cancer-amplified Zdhhc19 promote Stat3 activation through S-Palmitoylation</span>. <abbr class="cit-jnl-abbrev">Nature</abbr> <span class="cit-pub-date">2019</span>;<span class="cit-vol">573</span>:<span class="cit-fpage">139</span>–<span class="cit-lpage">43</span>. <a href="http://dx.doi.org/10.1038/s41586-019-1511-x">doi:10.1038/s41586-019-1511-x</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DNature%26rft.volume%253D573%26rft.spage%253D139%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-5-2" title="View reference 31 in text" id="ref-31">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.31" data-doi="10.1038/s41589-021-00785-8"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Remsberg</span> <span class="cit-name-given-names">JR</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Suciu</span> <span class="cit-name-given-names">RM</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Zambetti</span> <span class="cit-name-given-names">NA</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Cravatt, Abhd17 regulation of plasma membrane Palmitoylation and N-Ras-dependent cancer growth</span>. <abbr class="cit-jnl-abbrev">Nat Chem Biol</abbr> <span class="cit-pub-date">2021</span>;<span class="cit-vol">17</span>:<span class="cit-fpage">856</span>–<span class="cit-lpage">64</span>. <a href="http://dx.doi.org/10.1038/s41589-021-00785-8">doi:10.1038/s41589-021-00785-8</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DNat%2BChem%2BBiol%26rft.volume%253D17%26rft.spage%253D856%26rft_id%253Dinfo%253Adoi%252F10.1038%252Fs41589-021-00785-8%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1038/s41589-021-00785-8&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-5-2" title="View reference 32 in text" id="ref-32">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.32" data-doi="10.1126/science.aau6391"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Lu</span> <span class="cit-name-given-names">Y</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Zheng</span> <span class="cit-name-given-names">Y</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Coyaud</span> <span class="cit-name-given-names">É</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Palmitoylation of Nod1 and Nod2 is required for bacterial sensing</span>. <abbr class="cit-jnl-abbrev">Science</abbr> <span class="cit-pub-date">2019</span>;<span class="cit-vol">366</span>:<span class="cit-fpage">460</span>–<span class="cit-lpage">7</span>. <a href="http://dx.doi.org/10.1126/science.aau6391">doi:10.1126/science.aau6391</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DScience%26rft.stitle%253DScience%26rft.aulast%253DLu%26rft.auinit1%253DY.%26rft.volume%253D366%26rft.issue%253D6464%26rft.spage%253D460%26rft.epage%253D467%26rft.atitle%253DPalmitoylation%2Bof%2BNOD1%2Band%2BNOD2%2Bis%2Brequired%2Bfor%2Bbacterial%2Bsensing%26rft_id%253Dinfo%253Adoi%252F10.1126%252Fscience.aau6391%26rft_id%253Dinfo%253Apmid%252F31649195%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/ijlink/YTozOntzOjQ6InBhdGgiO3M6MTQ6Ii9sb29rdXAvaWpsaW5rIjtzOjU6InF1ZXJ5IjthOjQ6e3M6ODoibGlua1R5cGUiO3M6NDoiQUJTVCI7czoxMToiam91cm5hbENvZGUiO3M6Mzoic2NpIjtzOjU6InJlc2lkIjtzOjEyOiIzNjYvNjQ2NC80NjAiO3M6NDoiYXRvbSI7czoyMjoiL2d1dGpubC83My83LzExNTYuYXRvbSI7fXM6ODoiZnJhZ21lbnQiO3M6MDoiIjt9" class="cit-ref-sprinkles cit-ref-sprinkles-ijlink"><span><span class="cit-reflinks-abstract">Abstract</span><span class="cit-sep cit-reflinks-variant-name-sep">/</span><span class="cit-reflinks-full-text"><span class="free-full-text">FREE </span>Full Text</span></span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-33-1" title="View reference 33 in text" id="ref-33">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.33"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Ge</span> <span class="cit-name-given-names">C</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Tan</span> <span class="cit-name-given-names">J</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Dai</span> <span class="cit-name-given-names">X</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Hepatocyte phosphatase Dusp22 mitigates NASH-HCC progression by targeting FAK</span>. <abbr class="cit-jnl-abbrev">Nat Commun</abbr> <span class="cit-pub-date">2022</span>;<span class="cit-vol">13</span>:<span class="cit-elocation-id">5945</span>. <a href="http://dx.doi.org/10.1038/s41467-022-33493-5">doi:10.1038/s41467-022-33493-5</a> </cite></div><div class="cit-extra"></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-33-1" title="View reference 34 in text" id="ref-34">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.34"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Li</span> <span class="cit-name-given-names">Y</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Xu</span> <span class="cit-name-given-names">J</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Lu</span> <span class="cit-name-given-names">Y</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Drak2 aggravates Nonalcoholic fatty liver disease progression through Srsf6-associated RNA alternative splicing</span>. <abbr class="cit-jnl-abbrev">Cell Metabolism</abbr> <span class="cit-pub-date">2021</span>;<span class="cit-vol">33</span>:<span class="cit-fpage">2004</span>–<span class="cit-lpage">2020</span>. <a href="http://dx.doi.org/10.1016/j.cmet.2021.09.008">doi:10.1016/j.cmet.2021.09.008</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DCell%2BMetabolism%26rft.volume%253D33%26rft.spage%253D2004%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-35-1" title="View reference 35 in text" id="ref-35">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.35" data-doi="10.1074/jbc.273.25.15830"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Duncan</span> <span class="cit-name-given-names">JA</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Gilman</span> <span class="cit-name-given-names">AG</span> </span> </li></ol><cite>. <span class="cit-article-title">A cytoplasmic Acyl-protein Thioesterase that removes Palmitate from G protein alpha subunits and P21(RAS)</span>. <abbr class="cit-jnl-abbrev">J Biol Chem</abbr> <span class="cit-pub-date">1998</span>;<span class="cit-vol">273</span>:<span class="cit-fpage">15830</span>–<span class="cit-lpage">7</span>. <a href="http://dx.doi.org/10.1074/jbc.273.25.15830">doi:10.1074/jbc.273.25.15830</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DJournal%2Bof%2BBiological%2BChemistry%26rft.stitle%253DJ%2BBiol%2BChem%26rft.aulast%253DDuncan%26rft.auinit1%253DJ.%2BA.%26rft.volume%253D273%26rft.issue%253D25%26rft.spage%253D15830%26rft.epage%253D15837%26rft.atitle%253DA%2BCytoplasmic%2BAcyl-Protein%2BThioesterase%2BThat%2BRemoves%2BPalmitate%2Bfrom%2BG%2BProtein%2B%257Balpha%257D%2BSubunits%2Band%2Bp21RAS%26rft_id%253Dinfo%253Adoi%252F10.1074%252Fjbc.273.25.15830%26rft_id%253Dinfo%253Apmid%252F9624183%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/ijlink/YTozOntzOjQ6InBhdGgiO3M6MTQ6Ii9sb29rdXAvaWpsaW5rIjtzOjU6InF1ZXJ5IjthOjQ6e3M6ODoibGlua1R5cGUiO3M6NDoiQUJTVCI7czoxMToiam91cm5hbENvZGUiO3M6MzoiamJjIjtzOjU6InJlc2lkIjtzOjEyOiIyNzMvMjUvMTU4MzAiO3M6NDoiYXRvbSI7czoyMjoiL2d1dGpubC83My83LzExNTYuYXRvbSI7fXM6ODoiZnJhZ21lbnQiO3M6MDoiIjt9" class="cit-ref-sprinkles cit-ref-sprinkles-ijlink"><span><span class="cit-reflinks-abstract">Abstract</span><span class="cit-sep cit-reflinks-variant-name-sep">/</span><span class="cit-reflinks-full-text"><span class="free-full-text">FREE </span>Full Text</span></span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-35-1" title="View reference 36 in text" id="ref-36">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.36"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Tomatis</span> <span class="cit-name-given-names">VM</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Trenchi</span> <span class="cit-name-given-names">A</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Gomez</span> <span class="cit-name-given-names">GA</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Acyl-protein Thioesterase 2 Catalyzes the Deacylation of peripheral membrane-associated GAP-43</span>. <abbr class="cit-jnl-abbrev">PLoS One</abbr> <span class="cit-pub-date">2010</span>;<span class="cit-vol">5</span>:<span class="cit-elocation-id">e15045</span>. <a href="http://dx.doi.org/10.1371/journal.pone.0015045">doi:10.1371/journal.pone.0015045</a> </cite></div><div class="cit-extra"></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-37-1" title="View reference 37 in text" id="ref-37">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.37" data-doi="10.1038/nrc2222"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Menendez</span> <span class="cit-name-given-names">JA</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Lupu</span> <span class="cit-name-given-names">R</span> </span> </li></ol><cite>. <span class="cit-article-title">Fatty acid synthase and the Lipogenic phenotype in cancer pathogenesis</span>. <abbr class="cit-jnl-abbrev">Nat Rev Cancer</abbr> <span class="cit-pub-date">2007</span>;<span class="cit-vol">7</span>:<span class="cit-fpage">763</span>–<span class="cit-lpage">77</span>. <a href="http://dx.doi.org/10.1038/nrc2222">doi:10.1038/nrc2222</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DNature%2Breviews.%2BCancer%26rft.stitle%253DNat%2BRev%2BCancer%26rft.aulast%253DMenendez%26rft.auinit1%253DJ.%2BA.%26rft.volume%253D7%26rft.issue%253D10%26rft.spage%253D763%26rft.epage%253D777%26rft.atitle%253DFatty%2Bacid%2Bsynthase%2Band%2Bthe%2Blipogenic%2Bphenotype%2Bin%2Bcancer%2Bpathogenesis.%26rft_id%253Dinfo%253Adoi%252F10.1038%252Fnrc2222%26rft_id%253Dinfo%253Apmid%252F17882277%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1038/nrc2222&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a><a href="/lookup/external-ref?access_num=17882277&link_type=MED&atom=%2Fgutjnl%2F73%2F7%2F1156.atom" class="cit-ref-sprinkles cit-ref-sprinkles-medline"><span>PubMed</span></a><a href="/lookup/external-ref?access_num=000249691700013&link_type=ISI" class="cit-ref-sprinkles cit-ref-sprinkles-newisilink cit-ref-sprinkles-webofscience"><span>Web of Science</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-38-1" title="View reference 38 in text" id="ref-38">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.38"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Wang</span> <span class="cit-name-given-names">H</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Zhou</span> <span class="cit-name-given-names">Y</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Xu</span> <span class="cit-name-given-names">H</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Therapeutic efficacy of FASN inhibition in Preclinical models of HCC</span>. <abbr class="cit-jnl-abbrev">Hepatology</abbr> <span class="cit-pub-date">2022</span>;<span class="cit-vol">76</span>:<span class="cit-fpage">951</span>–<span class="cit-lpage">66</span>. <a href="http://dx.doi.org/10.1002/hep.32359">doi:10.1002/hep.32359</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DHepatology%26rft.volume%253D76%26rft.spage%253D951%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-39-1" title="View reference 39 in text" id="ref-39">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.39" data-doi="10.1016/j.cmet.2018.07.019"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Bruning</span> <span class="cit-name-given-names">U</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Morales-Rodriguez</span> <span class="cit-name-given-names">F</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Kalucka</span> <span class="cit-name-given-names">J</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Impairment of angiogenesis by fatty acid synthase inhibition involves mTOR Malonylation</span>. <abbr class="cit-jnl-abbrev">Cell Metab</abbr> <span class="cit-pub-date">2018</span>;<span class="cit-vol">28</span>:<span class="cit-fpage">866</span>–<span class="cit-lpage">880</span>.<span class="cit-elocation-id">S1550-4131(18)30462-5</span>. <a href="http://dx.doi.org/10.1016/j.cmet.2018.07.019">doi:10.1016/j.cmet.2018.07.019</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DCell%2BMetab%26rft.volume%253D28%26rft.spage%253D866%26rft_id%253Dinfo%253Adoi%252F10.1016%252Fj.cmet.2018.07.019%26rft_id%253Dinfo%253Apmid%252F30146486%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1016/j.cmet.2018.07.019&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a><a href="/lookup/external-ref?access_num=30146486&link_type=MED&atom=%2Fgutjnl%2F73%2F7%2F1156.atom" class="cit-ref-sprinkles cit-ref-sprinkles-medline"><span>PubMed</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-40-1" title="View reference 40 in text" id="ref-40">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.40"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Yao</span> <span class="cit-name-given-names">H</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Lan</span> <span class="cit-name-given-names">J</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Li</span> <span class="cit-name-given-names">C</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Inhibiting PD-L1 Palmitoylation enhances T-cell immune responses against tumours</span>. <abbr class="cit-jnl-abbrev">Nat Biomed Eng</abbr> <span class="cit-pub-date">2019</span>;<span class="cit-vol">3</span>:<span class="cit-fpage">414</span>. <a href="http://dx.doi.org/10.1038/s41551-019-0402-7">doi:10.1038/s41551-019-0402-7</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DNat%2BBiomed%2BEng%26rft.volume%253D3%26rft.spage%253D414%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-41-1" title="View reference 41 in text" id="ref-41">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.41"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Xu</span> <span class="cit-name-given-names">Z</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Xu</span> <span class="cit-name-given-names">M</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Liu</span> <span class="cit-name-given-names">P</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">The Mtorc2-Akt1 Cascade is crucial for C-Myc to promote Hepatocarcinogenesis in mice and humans</span>. <abbr class="cit-jnl-abbrev">Hepatology</abbr> <span class="cit-pub-date">2019</span>;<span class="cit-vol">70</span>:<span class="cit-fpage">1600</span>–<span class="cit-lpage">13</span>. <a href="http://dx.doi.org/10.1002/hep.30697">doi:10.1002/hep.30697</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DHepatology%26rft.volume%253D70%26rft.spage%253D1600%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-42-1" title="View reference 42 in text" id="ref-42">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.42" data-doi="10.1038/s41568-021-00388-4"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Hoy</span> <span class="cit-name-given-names">AJ</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Nagarajan</span> <span class="cit-name-given-names">SR</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Butler</span> <span class="cit-name-given-names">LM</span> </span> </li></ol><cite>. <span class="cit-article-title">Tumour fatty acid metabolism in the context of therapy resistance and obesity</span>. <abbr class="cit-jnl-abbrev">Nat Rev Cancer</abbr> <span class="cit-pub-date">2021</span>;<span class="cit-vol">21</span>:<span class="cit-fpage">753</span>–<span class="cit-lpage">66</span>. <a href="http://dx.doi.org/10.1038/s41568-021-00388-4">doi:10.1038/s41568-021-00388-4</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DNat%2BRev%2BCancer%26rft.volume%253D21%26rft.spage%253D753%26rft_id%253Dinfo%253Adoi%252F10.1038%252Fs41568-021-00388-4%26rft_id%253Dinfo%253Apmid%252F34417571%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/external-ref?access_num=10.1038/s41568-021-00388-4&link_type=DOI" class="cit-ref-sprinkles cit-ref-sprinkles-doi cit-ref-sprinkles-crossref"><span>CrossRef</span></a><a href="/lookup/external-ref?access_num=34417571&link_type=MED&atom=%2Fgutjnl%2F73%2F7%2F1156.atom" class="cit-ref-sprinkles cit-ref-sprinkles-medline"><span>PubMed</span></a></div></div></li><li><a class="rev-xref-ref" href="#xref-ref-43-1" title="View reference 43 in text" id="ref-43">↵</a><div class="cit ref-cit ref-journal" id="cit-73.7.1156.43" data-doi="10.1136/gutjnl-2018-317581"><div class="cit-metadata"><ol class="cit-auth-list"><li> </li><li><span class="cit-auth"> <span class="cit-name-surname">Che</span> <span class="cit-name-given-names">L</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Chi</span> <span class="cit-name-given-names">W</span> </span>, </li><li><span class="cit-auth"> <span class="cit-name-surname">Qiao</span> <span class="cit-name-given-names">Y</span> </span>, <span class="cit-etal">et al</span> </li></ol><cite>. <span class="cit-article-title">Cholesterol biosynthesis supports the growth of Hepatocarcinoma lesions depleted of fatty acid synthase in mice and humans</span>. <abbr class="cit-jnl-abbrev">Gut</abbr> <span class="cit-pub-date">2020</span>;<span class="cit-vol">69</span>:<span class="cit-fpage">177</span>–<span class="cit-lpage">86</span>. <a href="http://dx.doi.org/10.1136/gutjnl-2018-317581">doi:10.1136/gutjnl-2018-317581</a> </cite></div><div class="cit-extra"><a href="{openurl}?query=rft.jtitle%253DGut%26rft_id%253Dinfo%253Adoi%252F10.1136%252Fgutjnl-2018-317581%26rft_id%253Dinfo%253Apmid%252F30954949%26rft.genre%253Darticle%26rft_val_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Ajournal%26ctx_ver%253DZ39.88-2004%26url_ver%253DZ39.88-2004%26url_ctx_fmt%253Dinfo%253Aofi%252Ffmt%253Akev%253Amtx%253Actx" class="cit-ref-sprinkles cit-ref-sprinkles-openurl cit-ref-sprinkles-open-url"><span>OpenUrl</span></a><a href="/lookup/ijlink/YTozOntzOjQ6InBhdGgiO3M6MTQ6Ii9sb29rdXAvaWpsaW5rIjtzOjU6InF1ZXJ5IjthOjQ6e3M6ODoibGlua1R5cGUiO3M6NDoiQUJTVCI7czoxMToiam91cm5hbENvZGUiO3M6NjoiZ3V0am5sIjtzOjU6InJlc2lkIjtzOjg6IjY5LzEvMTc3IjtzOjQ6ImF0b20iO3M6MjI6Ii9ndXRqbmwvNzMvNy8xMTU2LmF0b20iO31zOjg6ImZyYWdtZW50IjtzOjA6IiI7fQ==" class="cit-ref-sprinkles cit-ref-sprinkles-ijlink"><span><span class="cit-reflinks-abstract">Abstract</span><span class="cit-sep cit-reflinks-variant-name-sep">/</span><span class="cit-reflinks-full-text"><span class="free-full-text">FREE </span>Full Text</span></span></a></div></div></li></ol></div><span class="highwire-journal-article-marker-end"></span></div><span class="related-urls"></span></div></div> </div> </div> <div class="panel-separator"></div><div class="panel-pane pane-highwire-article-data-supp" id="supplementary-materials"> <h2 class="pane-title">Supplementary materials</h2> <div class="pane-content"> <div class="item-list"><ul><li class="first last"><div class="highwire-markup"><div xmlns="http://www.w3.org/1999/xhtml" id="content-block" xmlns:xhtml="http://www.w3.org/1999/xhtml"><div><span class="highwire-journal-article-marker-start"></span><div class="auto-clean"><span style="font-family: Verdana,Arial,Helvetica,sans-serif; font-size: 83.33%"> <h2>Supplementary Data</h2> <p>This web only file has been produced by the BMJ Publishing Group from an electronic file supplied by the author(s) and has not been edited for content. </p> <ul><li><a href="/highwire/filestream/258919/field_highwire_adjunct_files/0/gutjnl-2023-330826supp001_data_supplement.pdf" class="rewritten" data-icon-position="" data-hide-link-title="0">Data supplement 1</a> </li></ul> </span> </div><span class="highwire-journal-article-marker-end"></span></div><span id="related-urls"></span></div></div></li> </ul></div> </div> </div> <div class="panel-separator"></div><div class="panel-pane pane-highwire-markup footnotes" > <div class="pane-content"> <div class="highwire-markup"><div xmlns="http://www.w3.org/1999/xhtml" class="content-block-markup" xmlns:xhtml="http://www.w3.org/1999/xhtml"><div xmlns:xhtml="http://www.w3.org/1999/xhtml" class="section fn-group" id="fn-group-1"><h2>Footnotes</h2><ul><li class="fn-other" id="fn-1"><p id="p-48"><span class="fn-label">Correction notice</span> This article has been corrected since it published Online First. The ORCID ID's for authors have been corrected.</p></li><li class="fn-other" id="fn-2"><p id="p-49"><span class="fn-label">Contributors</span> Guarantor: JG. Conception and design: JG, LB and MK. Development of methodology: LB, ZZ, JC, YF, JG and YS. Acquisition of data (provided animals, acquired and managed patients, provided facilities, etc): LB, ZZ, JC, YF, HW, XZ, XW. Analysis and interpretation of data (eg, statistical analysis, biostatistics, computational analysis): LB, ZZ, JC, QJ, BG. Writing the manuscript and revision: JG, LB. Administrative, technical, or material support (ie, reporting or organising data, constructing databases): LB, ZZ, JC, LW, XZ, WX, KH, WW, MK. Study supervision: JG, MK. Approved manuscript: all authors.</p></li><li class="fn-other" id="fn-3"><p id="p-50"><span class="fn-label">Funding</span> This work was supported in part by National Key Research and Development Program of China (2023YFC3402100 to J.G), National Nature Science Foundation of China (32070767 to J.G, 82302911 to L.B) and Guangdong Basic and Applied Basic Research Foundation (2022A1515220004 to J.G).</p></li><li class="fn-conflict" id="fn-4"><p id="p-51"><span class="fn-label">Competing interests</span> WW is a co-founder and consultant for the ReKindle Therapeutics.</p></li><li class="fn-other" id="fn-5"><p id="p-52"><span class="fn-label">Patient and public involvement</span> Patients and/or the public were not involved in the design, or conduct, or reporting, or dissemination plans of this research.</p></li><li class="fn-other" id="fn-6"><p id="p-53"><span class="fn-label">Provenance and peer review</span> Not commissioned; externally peer reviewed.</p></li><li class="fn-other" id="fn-7"><p id="p-54"><span class="fn-label">Supplemental material</span> This content has been supplied by the author(s). 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