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Regulation of HIV-1 RNA Processing

HIV-1 RNA 加工的调控

基本信息

批准号：
7151161
负责人：
MASSIMO CAPUTI
金额：
$ 28.69万
依托单位：
FLORIDA ATLANTIC UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2002
资助国家：
美国
起止时间：
2002-12-15 至 2009-11-30
项目状态：
已结题

项目摘要

The integrated HIV-1 proviral genome is transcribed by the host transcription machinery into a single 9.2 kb primary transcript. To express the nine different gene products required for viral replication, HIV has developed a number of strategies to regulate splicing and to circumvent cellular mechanisms restricting unspliced RNA in the cytoplasm. The regulatory protein Rev promotes transport of the incompletely spliced viral RNAs to the cytoplasm, binding to the Rev Responsive Element (RRE) on the viral RNA. Mechanisms regulating the appearance of unspliced and spliced viral RNAs, and their transport to the cytoplasm, involve interactions between viral RNA sequences and host cell factors. The characterization of these viral/host interactions could allow the design of novel therapeutic compounds. hnRNP H proteins have been shown to be necessary for HIV-1 RNA splicing in a series of in vitro experiments. They may also regulate RNA export and trafficking in infected cells. The role of single members of the hnRNP H protein family in HIV-1 RNA processing in vivo will be investigated by modulating hnRNP H protein expression in cells transfected with HIV-1-derived plasmids, bearing mutations in regulatory sequences. Several viral sequences that regulate viral RNA splicing in vitro have been identified. These will be tested to determine whether they are necessary for the regulation of viral RNA splicing in vivo. Preliminary data indicates that viral sequences interact with the RRE-Rev complex and, possibly, have a role in the Rev-dependent viral RNA transport pathway. These interactions will be characterized by a combination of in vivo and in vitro assays. Transcription and splicing have been shown to be closely coupled in other systems. Preliminary data indicates that the viral promoter influences splicing of viral substrates in vitro. Possible coupling of HIV-1 transcription and splicing will be investigated, using both a coupled transcription-splicing assay and transient transfection experiments.

整合的HIV-1前病毒基因组由宿主转录机制转录成一个转录因子。单个9.2kb初级转录物。为了表达病毒复制所需的九种不同的基因产物， HIV已经发展了许多策略来调节剪接和规避细胞机制限制细胞质中未剪接的RNA。调节蛋白Rev可促进蛋白质的转运，不完全剪接的病毒RNA到细胞质，结合到细胞质上的Rev应答元件（RRE）。病毒RNA调节未剪接和剪接的病毒RNA的出现的机制，以及它们的转运到细胞质，涉及病毒RNA序列和宿主细胞因子之间的相互作用。的这些病毒/宿主相互作用的表征可以允许设计新的治疗化合物。 hnRNP H蛋白在一系列体外实验中显示为HIV-1 RNA剪接所必需。实验它们还可能调节受感染细胞的RNA输出和运输。单身的作用 hnRNP H蛋白家族成员在体内HIV-1 RNA加工中的作用将通过在用HIV-1衍生质粒转染的细胞中调节hnRNP H蛋白表达，调控序列中的突变。几种在体外调节病毒RNA剪接的病毒序列具有被识别。这些将被测试，以确定它们是否是必要的调节病毒体内RNA剪接。初步数据表明，病毒序列与RRE-Rev复合物相互作用并可能在Rev依赖性病毒RNA转运途径中发挥作用。这些互动将是其特征在于体内和体外测定的组合。转录和剪接已经被证明与其他系统紧密结合。初步数据表明，病毒启动子影响体外病毒底物的剪接。HIV-1转录和剪接的可能偶联将是研究，使用耦合转录剪接测定和瞬时转染实验。