摘要
目的探究丝氨酸/精氨酸富集剪接因子1(serine/arginine-rich splicing factor 1,SRSF1)对乙型肝炎病毒(hepatitis B virus,HBV)复制的影响和机制。方法通过Southern blot、Northern blot、ELISA实验在不同的HBV复制细胞模型中检测SRSF1对HBV复制的影响;通过荧光定量PCR实验、荧光素酶报告系统以及ChIP实验在HepG2细胞中研究SRSF1对HBV核心启动子/增强子活性的影响。利用Western blot和泛素化检测实验探索SRSF1与P53的关系,同时在不同的HBV复制细胞模型中验证P53对HBV复制的影响,之后明确P53在SRSF1抑制HBV中的作用。结果过表达SRSF1可以在多种HBV复制细胞模型中显著抑制HBV DNA和HBV RNA,降低HBeAg和HBsAg的分泌水平(P<0.0001)。SRSF1还可以抑制HBV核心启动子活性,尽管这种抑制是通过间接机制调控的。此外,SRSF1通过保护P53免受泛素化和随后的蛋白酶体降解从而增强P53的稳定性。同时,过表达P53或敲低P53对HBV的调控作用也在不同的细胞模型中得到了验证(P<0.0001)。结论剪接因子SRSF1的过表达可以显著抑制HBV的复制,且该抑制效应在多种肝癌细胞模型中均有重现性,这一抑制效应由其增强P53的稳定性来介导。
Objective To investigate the effect and underlying mechanism of serine and arginine rich splicing factor 1(SRSF1)on the replication of hepatitis B virus(HBV).Methods The effects of SRSF1 on HBV replication were investigated in different HBV replicating cell models by Southern blotting,Northern blotting and ELISA.Quantitative PCR,luciferase reporter assay and chromatin immunoprecipitation(ChIP)were applied to determine the effects of SRSF1 on the activities of HBV core promoter/enhancer in HepG2 cells.The relationship between SRSF1 and P53 was explored with Western blotting and ubiquitination assay.The effects of P53 on HBV replication were verified in different HBV replicating cell models,and the role of P53 in SRSF1 inhibition of HBV was clarified.Results Overexpression of SRSF1 significantly inhibited HBV DNA and HBV RNA and reduced HBsAg and HBeAg secretion levels in a variety of HBV replicative cell models(P<0.0001).SRSF1 also inhibited the activity of HBV core promoter,although this inhibition was regulated by indirect mechanisms.In addition,SRSF1 enhanced P53 stability by protecting P53 from ubiquitination and subsequent proteasomal degradation.Meanwhile,the regulatory effect of overexpression or knockdown of P53 on HBV was validated in different cell models(P<0.0001).Conclusion Overexpression of splicing factor SRSF1 significantly inhibits HBV replication in a variety of cell models,and this inhibitory effect is mediated by its enhancement of P53 stability.
作者
刘佳俊
龙少媛
胡接力
崔静
LIU Jiajun;LONG Shaoyuan;HU Jieli;CUI Jing(Key Laboratory of Molecular Biology for Infectious Diseases of Ministry of Education,Chongqing Medical University,Chongqing;First Affiliated Hospital,Air Force Medical University,Xi’an,Shaanxi,China)
出处
《陆军军医大学学报》
北大核心
2025年第13期1475-1483,共9页
Journal of Army Medical University
基金
国家自然科学基金面上项目(82470634)
重庆市教委科学技术研究重点项目(KJZD-K202200409)。
