O-GlcNAcylation Coordinates Glutaminolysis by Regulating the Stability and Membrane Trafficking of ASCT2 in Hepatic Stellate Cells  

作　　者：Feixia Wang Li Chen Biyun Zhang Zhanghao Li Min Shen Li Tang Zili Zhang Jiangjuan Shao Feng Zhang Shizhong Zheng Shanzhong Tan 

机构地区：[1]Department of Integrated TCM and Western Medicine,Nanjing Hospital Affiliated to Nanjing University of Chinese Medicine,Nanjing,Jiangsu,China [2]Jiangsu Key Laboratory for Pharmacology and Safety Evaluation of Chinese Materia Medica,Nan-jing University of Chinese Medicine,Nanjing,Jiangsu,China [3]Department of Nuclear Medicine,Jiangsu Province Hospital of Chinese Medicine,Affiliated Hospital of Nanjing University of Chinese Medicine,Nanjing,Jiangsu,China

出　　处：《Journal of Clinical and Translational Hepatology》2022年第6期1107-1116,共10页临床与转化肝病杂志（英文版）

基　　金：the Leading Talent Project of Ji-angsu Province Traditional Chinese Medicine(SLJ0216);the National Natural Science Foundation of China(82073914,81870423);the Major Project of the Natural Science Research of Jiangsu Higher Education Institutions(19KJA310005);the Postgraduate Research&Practice Innovation Program of Jiangsu Province,Grant/Award Number:KYCX20_1493;the Joint Project of Jiangsu Key Laboratory for Pharmacol-ogy and Safety Evaluation of Chinese Materia Medica and Yangtze River Pharmaceutical(JKLPSE202005)。

摘　　要：Background and Aims:Recognition of excessive activa-tion of hepatic stellate cells(HSCs)in liver fibrosis prompt-ed us to investigate the regulatory mechanisms of HSCs.We aimed to examine the role of O-GlcNAcylation modifica-tion of alanine,serine,cysteine transporter 2(ASCT2)in HSCs and liver fibrosis.Methods:The expression of O-Glc-NAcylation modification in fibrotic mice livers and activated HSCs was analyzed by western blotting.Immunoprecipita-tion was used to assess the interaction of ASCT2 and O-Glc-NAc transferase(OGT).In addition,ASCT2 protein stability was assayed after cycloheximide(CHX)treatment.The O-GlcNAcylation site of ASCT2 was predicted and mutated by site-directed mutagenesis.Real-time PCR,immunofluores-cence,kit determinations and Seahorse assays were used to clarify the effect of ASCT2 O-GlcNAcylation on HSC glu-taminolysis and HSC activation.Western blotting,immuno-chemistry,and immunohistofluorescence were used to ana-lyze the effect of ASCT2 O-GlcNAcylation in vivo.Results:We observed significantly increased O-GlcNAcylation modi-fication of ASCT2.ASCT2 was found to interact with OGT to regulate ASCT2 stability.We predicted and confirmed that O-GlcNAcylation of ASCT2 at Thr122 site resulted in HSCs activation.We found Thr122 O-GlcNAcylation of ASCT2 me-diated membrane trafficking of glutamine transport and attenuated HSC glutaminolysis.Finally,we validated the expression and function of ASCT2 O-GlcNAcylation after in-jection of AAV8-ASCT2 shRNA in CCl4-induced liver fibrosis mice in vivo.Conclusions:Thr122 O-GlcNAcylation regu-lation of ASCT2 resulted in stability and membrane traf-ficking-mediated glutaminolysis in HSCs and liver fibrosis.Further studies are required to assess its role as a putative therapeutic target.

关 键 词：ASCT2 O-GLCNACYLATION Stability TRAFFICKING Liver fibrosis 

分 类 号：R575.2[医药卫生—消化系统]