摘要
目的探究人肝癌细胞HepG2.2.15中Ras相关蛋白7a (Rab7a)的巯基化机制及其对乙型肝炎病毒(HBV)复制的影响。方法通过生物素转换实验检测H2S对HepG2.2.15细胞中Rab7a巯基化水平的影响并验证Rab7a的巯基化位点;通过酶联免疫法、RT-qPCR和Western blot检测巯基化的Rab7a对HBV复制标志物水平的影响。结果在HepG2.2.15细胞中H2S可增强内源性Rab7a的巯基化水平(P<0.01);NaHS(H2S速释供体)可增强外源性Rab7a的巯基化表达,但其巯基化位点突变后则不受NaHS影响(P<0.01);Rab7a巯基化后,可抑制HepG2.2.15细胞上清中HBsAg、HBeAg和HBV-DNA及细胞内HBV-cccDNA、3.5kb RNA、total RNA和HBcAg蛋白的表达(P<0.05)。结论 Rab7a的巯基化抑制HBV阳性细胞系HepG2.2.15细胞中HBV的复制水平。
Objective To investigate the mechanism of sulfhydrated Ras related protein 7 a(Rab7 a) in HepG2.2.15 cells and its effect on hepatitis B virus(HBV) replication. Methods The sulfhydrated Rab7 a and sulfhydrated sites of Rab7 a were verified by biotin switch method. The effect of the sulfhydrated Rab7 a on the level of HBV replication markers was detected by enzyme-linked immunosorbent assay, RT-qPCR and Western blot. Results In HepG2.2.15 cells, H2S enhanced the sulfhydration level of endogenous Rab7 a(P<0.01). NaHS(H2S rapid release donor) enhanced the expression of sulfhydration of exogenous Rab7 a, but the mutation of sulfhydration site was not affected by NaHS(P<0.01). In HepG2.2.15 cells, H2S enhanced the sulfhydrylation level of endogenous and exogenous Rab7 a;the sulfhydrylated Rab7 a inhibited the expression of HBsAg, HBeAg and HBV-DNA in supernatant, as well as HBV-cccDNA, 3.5 RNA, total RNA and HBcAg protein in HepG2.2.15 cells(P<0.05). Conclusions Sulfhydrated Rab7 a inhibites the replication of HBV in HBV positive cell line HepG2.2.15.
作者
宋佳茹
朱席琳
伍晓盼
刘英
SONG Jia-ru;ZHU Xi-lin;WU Xiao-pan;LIU Ying(State Key Laboratory of Medical Molecular Biology,Institute of Basic Medical Sciences CAMS,School of Basic Medicine PUMC,Beijing 100005,China)
出处
《基础医学与临床》
CSCD
2020年第1期37-40,共4页
Basic and Clinical Medicine
基金
国家重点基础研究发展计划(973计划)(2012CB519005)
