REGULATION OF HIV GENE EXPRESSION BY TAT PEPTIDES

描述：(改编自申请者摘要)长期目标 本课题旨在了解TAT的生化机制。 (反式激活剂)动作，并利用TAT功能的基本知识 设计HIV复制的拮抗剂。首先，为了理解TAT的功能， 研究人员建议将重点放在一种新的人类细胞蛋白质上，这种蛋白质含有36 KDA，他已经证明：(1)与TAT紧密结合，(2)与TAT形成络合物 TAT在体外，以及(3)当导入TAT时，增强TAT的反式激活 啮齿动物细胞。作者将进一步描述这个36 kDa的蛋白质， 描绘与其结合的TAT识别序列，确定细胞 物种和细胞类型的表达，克隆其cDNA并研究其在TAT中的作用 激活。其次，在了解TAT吸收的研究中，作者 将进一步鉴定他发现的一种90 kDa的细胞表面蛋白 在体内和体外与TAT结合。他建议分析 这种90 kDa的蛋白质通过交联性研究和TAT细胞摄取 Scatchard分析，描绘用于结合的TAT识别序列 90 kDa，并通过氨基酸序列分析确定90 KDA是一种新的蛋白质。如果有必要，他将分离出编码的cdna克隆。 90 kDa，用于进一步研究。第三，进一步发展TAT的研究 HIV复制的多肽拮抗剂，研究人员将利用 (1)细胞TAT识别序列作图研究结果 摄取，(2)焦油结合，和(3)36 kDa结合 将被细胞摄取并阻断HIV的小TAT多肽 复制。作为一个长期目标，有希望的TAT多肽的类似物 将采用模拟多肽技术合成，产率稳定且更多 高效的TAT拮抗剂。第四，探索TAT多肽的用途 将外源多肽运送到细胞内的序列，作者 建议分析细胞摄取、核运输和生物 含有TAT“递送”序列的融合多肽的功能。他 建议首先检查116个氨基酸的HIV Rev(病毒粒子的调节因子 蛋白质表达[以前的ART/TRS])与TAT融合的蛋白质。

DESCRIPTION: (Adapted from applicant's abstract) The long-term goals of this project are to understand the biochemical mechanism of Tat (transactivator) action and to utilize basic knowledge of Tat function to design antagonists of HIV replication. First, to understand Tat function, the investigator proposes to focus on a new human cellular protein of 36 kDa that he has shown to (1) bind tightly to Tat, (2) form a complex with Tat in vitro, and (3) potentiate Tat transactivation when introduced into rodent cells. The author will further characterize this 36 kDa protein, delineate the Tat recognition sequences for binding it, determine cell species and cell type expression, clone its cDNA and study its role in Tat transactivation. Second, in studies to understand Tat uptake, the author will further characterize a 90 kDa cell surface protein that he has found to bind to Tat in vivo and in vitro. He proposes to analyze the role of this 90 kDa protein in Tat cellular uptake by cross-linking studies and Scatchard analysis, to delineate the Tat recognition sequences for binding 90 kDa, and to determine by amino acid sequence analysis whether the 90 kDa is a new protein. If warranted, he will isolate a cDNA clone encoding 90 kDa for further studies. Third, in studies to develop further Tat peptide antagonists of HIV replication, the investigator will utilize the results of mapping studies of Tat recognition sequences for (1) cell uptake, (2) TAR binding, and (3) 36 kDa binding in attempts to design small Tat peptides that will be taken up by cells and block HIV replication. As a long-range goal, analogues of promising Tat peptides will be synthesized by peptide mimetic technology to yield stable and more efficient Tat antagonists. Fourth, to explore the use of Tat peptide sequences to transport foreign polypeptides into cells, the author proposes to analyze the cellular uptake, nuclear transport, and biological function of fusion polypeptides containing Tat "delivery" sequences. He proposes to first examine the 116 amino acid HIV Rev (regulator of virion protein expression [formerly art/trs]) protein fused to Tat.