空军军医大学学报2024，Vol.45Issue(7)：732-739,8.DOI:10.13276/j.issn.2097-1656.2024.07.003
乙型肝炎病毒大表面蛋白诱导内质网应激调控p27 IRES翻译活性的机制研究
Regulation mechanism of p27 IRES translation activity under large HBV surface protein-induced endoplasmic reticulum stress
摘要
Abstract
Objective To investigate the activity changes of internal ribosome entry site(IRES)in tumor suppressor gene p27 5'UTR and its regulatory mechanism under endoplasmic reticulum stress induced by large HBV surface protein(LHB).Methods Dual-luciferase reporter plasmid was constructed,and dual-luciferase reporter assay was used to detect the activity of IRES in p27 5'UTR.qRT-PCR and Western blotting were used to detect the expression levels of unfolded protein response-related molecules induced by LHB.RNA pull-down,mass spectrometry and immunoprecipitation were used to screen and verify the potential proteins that bound to p27 5'UTR.The knock-down cell models of PCBP2 and ANXA2 were established with siRNA transfection,and changes in the translation activity of p27 IRES in PCBP2 and ANXA2 knock-down cells were detected by dual-luciferase reporter assay.Results A dual-luciferase reporter plasmid was constructed to detect the activity of p27 IRES.LHB could induce endoplasmic reticulum stress and significantly increased the translation activity of p27 IRES(P＜0.01).Overexpression of LHB activated PERK pathway and increased the phosphorylation of eIF2α.The phosphorylation level of RERK and eIF2α was decreased by using the specific PERK pathway inhibitor GSK2606414,and the IRES translation activity of p27 5'UTR was significantly decreased(P＜0.01,P＜0.05).Meanwhile,phosphatase inhibitor Sal003 significantly increased the phosphorylation level of eIF2α,which further enhanced the activity of p27 IRES(P＜0.01,P＜0.05).Finally,we found that PCBP2 and ANXA2 as potential IRES trans-acting factors,played a positive role in enhancing the activity of p27 IRES.Conclusion LHB-induced endoplasmic reticulum stress enhances the translation activity of p27 IRES through eIF2α phosphorylation.IRES trans-acting factors PCBP2 and ANXA2 positively regulate the translation activity of p27 IRES.关键词
乙型肝炎病毒大表面蛋白/p27 5'UTR/内质网应激/内部核糖体进入位点/反式作用因子Key words
large HBV surface protein/p27 5'UTR/endoplasmic reticulum stress/internal ribosome entry sites/IRES trans-acting factor分类
医药卫生引用本文复制引用
郭亦笑,邵杰,尉丁,陈志南,边惠洁..乙型肝炎病毒大表面蛋白诱导内质网应激调控p27 IRES翻译活性的机制研究[J].空军军医大学学报,2024,45(7):732-739,8.基金项目
国家自然科学基金(82130084) （82130084）
