STAT1对hsp90α基因表达的负调控作用  

Role of STAT1 on the Regulation the Human hsp90α Gene Expression

作　　者：陈雪松[1] 吴宁华[1] 沈珝琲[1] 

机构地区：[1]中国医学科学院 中国协和医科大学 基础医学研究所医学分子生物学国家重点实验室,北京100005

出　　处：《中国医学科学院学报》2001年第4期356-360,共5页Acta Academiae Medicinae Sinicae

基　　金：国家自然科学基金重点项目(39930050)资助

摘　　要：目的 探讨转录因子STAT1对hsp90α 基因表达的调控作用。方法 将STAT1表达质粒转染Jurkat细胞，利用竞争性RT-PCR定量检测其内源性hsp90α 基因mRNA水平。共转染STAT1表达质粒和hsp90α基因上游调控区不同长度的片段驱动的氯霉素乙酰转移酶（CAT）报告基因质粒，利用定量RT-PCR检测CAT报告基因转录水平。用Western印迹分析方法检测42℃ 1 h热激前后Jurkat细胞中STAT1酪氨酸磷酸化状态，用电泳迁移率变更分析（EMSA）检测STAT1与hsp90α 5′基因上游调控区中含有STAT1特异结合元件的双链寡核苷酸片段特异性结合程度。结果 STAT1既可抑制Jurkat细胞hsp90α基因的表达，又能抑制hsp90α基因5′上游调控区驱动的CAT报告基因活性。热激以前Jurkat细胞中的STAT1 701位酪氨酸已有一定程度的磷酸化，热激以后其磷酸化程度明显升高，而且STAT1与hsp90α基因 5′上游调控区中的STAT1结合元件呈特异性结合。结论 STAT1对hsp90α基因的表达具有负调控作用。Objective To investigate the role of STAT1 on the regulation of human hsp90α gene expression． Methods We first transfected Jurkat cells with the STAT1 expression construct and analysised the expression of hsp90α gene expression via quantitative RT-PCR system． Then we co-transfected the STAT1 expression construct and the CAT reporter gene driven by different length of 5′ flanking sequence of hsp90α gene． Western blot was carried out to detect the level of tyrosine phosphorylation in Jurkat cells with and without heat shock treatment(42℃ 1 h)． By electrophoretic mobility shift assays(EMSA), we evaluated the DNA binding activity of a STAT1 responsible element located in the regulatory region of hsp90α gene in Jurkat cell nuclear extracts． Results The mRNA level of hsp90α gene in Jurkat cells was decreased when transfected by STAT1 expression construct, over-ex- pression of STAT1 down-regulates the expression of CAT reporter gene with the present of a distal fragment from －1756 to －1463 within the 5′ flanking regulatory sequences of hsp90α gene． The tyro- sine phosphorylation of STAT1 was detectable in Jurkat cells and increased when subjected to heat shock． Electrophoretic mobility shift assays(EMSA)results showed that STAT1 could bind to its resp- onsible element in the regulatory region of hsp90α gene． Conclusion STAT1 could negatively regu- late the human hsp90α gene expression．

关 键 词：STA1 hsp90∝ 基因表达调控 

分 类 号：Q343.1[生物学—遗传学]