Elucidation of host-retrovirus interaction by analyzing chromatin dynamics

通过分析染色质动力学阐明宿主-逆转录病毒相互作用

基本信息

批准号：
16209016
负责人：
IBA Hideo
金额：
$ 31.12万
依托单位：
The University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2006
项目状态：
已结题

项目摘要

For the establishment of new virology after the post-genome era, it is important to elucidate molecular basis of epigenetics and to understand infection mechanisms by analyzing dynamics of host and viral chromatin. The goal of this project is to elucidate molecular mechanisms of early infection, latent infection of retrovirus and lentiviruses through viral transcriptional initiation, maintenance, gene silencing and reactivation, which involves chromatin remodeling factor, SWI/SNF complex. During these three years of this project, we obtained following results.1.We clearly demonstrated that Brm subunit of human SWI/SNF complex is essential for stable and prolonged transcription drived by MLV-LTR or HIV-LTR in the absence of tat. In human tumor cell lines that are deficient in Brm expression, the expression of MLV- or HIV- based vectors are rapidly silenced in a stochastic manner.2.Using proteomics, we have identified several proteins that substoichiometorically interact with SWI/SNF complex. Among them, we scrutinized p54^<nrb>, because this protein is suggested to be involve in transcription, splicing and RNA editing and also because it can bind to a specific region if HIV RNA. Our analysis clearly indicated that p54^<nrb> and SWI/SNF complex translocates several promoter regions and they also bind to the downstream of the promoter or its RNA transcripts, affecting splicing patterns.In Brm deficient cell lines, therefore, some exons were tend to be excluded. We are now analyzing whether p54^<nrb> is also affect the splicing of HIV that produces several subgenomic RNA.

对于后基因组时代后新病毒学的建立，阐明表观遗传学的分子基础并通过分析宿主和病毒染色质的动态来理解感染机制具有重要意义。该项目的目标是阐明逆转录病毒和慢病毒通过病毒转录起始、维持、基因沉默和重新激活的早期感染、潜伏感染的分子机制，其中涉及染色质重塑因子、SWI/SNF复合物。在这个项目的这三年里，我们获得了以下结果： 1.我们清楚地证明了人类SWI/SNF复合物的Brm亚基对于在tat缺失的情况下由MLV-LTR或HIV-LTR驱动的稳定和延长的转录至关重要。在Brm表达缺陷的人类肿瘤细胞系中，基于MLV或HIV的载体的表达以随机方式迅速沉默。2.利用蛋白质组学，我们已经鉴定了几种与SWI/SNF复合物以亚化学计量相互作用的蛋白质。其中，我们仔细研究了p54^<nrb>，因为该蛋白被认为参与转录、剪接和RNA编辑，还因为它可以与HIV RNA的特定区域结合。我们的分析清楚地表明，p54^<nrb> 和 SWI/SNF 复合体易位了多个启动子区域，并且它们还与启动子或其 RNA 转录本的下游结合，影响剪接模式。因此，在 Brm 缺陷细胞系中，一些外显子倾向于被排除在外。我们现在正在分析 p54^<nrb> 是否也影响产生多个亚基因组 RNA 的 HIV 剪接。