TAT ASSOCIATED KINASE REGULATION OF HIV GENE EXPRESSION

描述：(改编自《调查人员摘要》)TAT刺激基因 提高RNA聚合酶II高效表达能力的研究 经历转录延伸。最近的研究表明，TAT可以 结合一种能磷酸化C端结构域(CTD)的细胞激酶 RNA聚合酶II。研究人员已经纯化了激酶复合体，并 证明了它含有新的成分以及以前的 多蛋白转录因子Tf II H的特征成分。 野生型，而不是突变的TAT蛋白，极大地增强了 激酶复合体使RNA聚合酶II CTD磷酸化，并添加 用HIV-1 LTR对体外转录的激酶进行显著检测 刺激TAT激活，表明TAT相关激酶复合体是 参与HIV-1 TAT激活的关键细胞因子。这个 研究人员建议研究TAT与 一种激酶复合体增加了HIV-1基因的表达。具体目标包括： 确定激酶复合体的哪些细胞成分能够 与TAT直接关联。测定新成分的功能 TAT相关复合体的体外和体内检测。测定 TAT是否促进了激酶复合体与RNA的结合 聚合酶II和TAT是否改变TF II H的酶活性。 建立一种重组的体外转录系统，其中TAT 激活依赖于TAT相关的激酶的添加和 进一步确定该激酶在体内调节HIV-1的作用 基因表达。

DESCRIPTION: (Adapted From Investigator's Abstract) Tat stimulates gene expression by increasing the ability of RNA polymerase II to efficiently undergo transcriptional elongation. Recent studies suggest that tat can bind a cellular kinase that phosphorylates the C-terminal domain (CTD) of RNA polymerase II. The investigator has purified the kinase complex and demonstrated that it contains novel components as well as previously characterized components of the multi-protein transcription factor, TF II H. Wildtype, but not mutant tat proteins, greatly enhance the ability of the kinase complex to phosphorylate the RNA polymerase II CTD and addition of the kinase to in vitro transcription assays with the HIV-1 LTR markedly stimulated tat activation, suggesting that tat-associated kinase complex is a critical cellular factor involved in tat activation of HIV-1. The investigator proposes to examine the mechanism by which tat association with a kinase complex increases HIV-1 gene expression. Specific aims include: Determine which cellular components of the kinase complex are able to directly associate with tat. Assay the function of novel components of the tat-associated complex using both in vitro and in vivo assays. Determine whether tat facilitates association of the kinase complex with RNA polymerase II and whether tat alters enzymatic activities of TF II H. Develop a reconstituted in vitro transcription system in which tat activation is dependent on the addition of the tat-associated kinase and further to determine the role of this kinase on regulating in vivo HIV-1 gene expression.