REGULATION OF TRANSCRIPTIONAL ELONGATION BY HIV1 TAT

转录延长的控制已经被认为是一种 基因调控的重要一步，但调控机制 RNA聚合酶II的伸长效率尚未得到 经过广泛研究。拟议研究的目标是 用HIV-L TAT作为研究对象阐明延长的调节机制 模型系统。TAT通过以下方式刺激伸长效率 识别定位的反式作用反应(TAR)RNA元件 在新生的病毒记录的5‘端。TAT反式激活 需要特定的细胞辅因子。申请人最近 分离和克隆了一种细胞辅因子TAT-SFI，它是 是TAT激活延伸率所必需的，并且是 相关的纤维素酶的底物。TAT-SFI与 EWS和FUS/TLS，它们是一类新的 具有RNA识别基序的假定转录因子和 常与肉瘤有关。初步研究表明 纹身可能通过招募一名员工来刺激伸长效率 含TAT-SFI及其激酶的HIV预制复合体 并通过增强伸长因子的作用， Elongin(Sill)。这项建议旨在进一步剖析其功能 通过分析序列分析TAT-SFI在TAT激活中的作用 TAT、TAT-SFI、TAT-SFI、TAT-SFI之间各种相互作用的特异性 SFI激酶和TAR，并通过作图和氨基酸 对TAT-SFI活性至关重要的残留物。一个重要的目标是 评估TAT-SFI耐人寻味的功能意义 TAT激活中的磷酸化以及分离、克隆和 确定它的激酶的特征。还将做出重大努力，致力于 TAT和TAT SFI作用机制的研究 激发延伸率和识别的效率 TAT刺激的拉长聚合酶复合体的组成部分。 最后，TAT与细胞伸长的关系 Elongin和TFIIS等因子在聚合酶激活中的作用 过程性将被研究。TAT活性的进一步分析 延伸率和细胞辅因子的作用 在这个过程中可能会揭示基因的一般机制 伸长阶段的调控。

Control of transcriptional elongation has been recognized as an important step in gene regulation, but mechanisms regulating the efficiency of elongation by RNA polymerase II have not been extensively studied. The goal of the proposed research is to elucidate mechanisms regulating elongation using HIV-l Tat as a model system. Tat stimulates the efficiency of elongation by recognizing the trans-acting-response (TAR) RNA element located at the 5' end of the nascent viral transcript. Tat trans-activation requires specific cellular cofactors. The applicant has recently isolated and cloned a cellular cofactor, Tat-SFI, which is specifically required for Tat activation of elongation and is a substrate of an associated linase. Tat-SFI is distantly related to EWS and FUS/TLS, which are members of a novel class of putative transcription factors with RNA recognition motifs and are frequently associated with sarcomas. Preliminary studies suggest that Tat may stimulate the efficiency of elongation by recruiting a preformed complex containing Tat-SFI and its kinase to the HIV promoter and by enhancing the effect of an elongation factor, Elongin (Sill). This proposal seeks to further dissect the function of Tat-SFI in Tat activation through analyzing the sequence specificity of the various interactions among Tat, Tat-SFI, the Tat- SFI kinase, and TAR, and through mapping the and amino acid residues critical for Tat-SFI activity. One important objective is to assess the intriguing functional significance of Tat-SFI phosphorylation in Tat activation and to isolate, clone, and characterize its kinase. A significant effort will also be devoted to the investigation of the mechanisms by which Tat and Tat SFI stimulate the efficiency of elongation and the identification of components of a Tat-stimulated elongating polymerase complex. Finally, the relationship between Tat and cellular elongation factors such as Elongin and TFIIS in activation of polymerase processivity will be studied. Further analysis of Tat activation of the efficiency of elongation and the role of the cellular cofactors in this process will likely reveal general mechanisms of gene regulation at the stage of elongation.