Do the B2 and Alu ncRNAs Repress Host Cell mRNA Transcription During HSV-1 Infect

B2 和 Alu ncRNA 在 HSV-1 感染期间抑制宿主细胞 mRNA 转录吗

• 批准号: 8503359
• 负责人: James Goodrich
• 金额: $ 7.63万
• 依托单位: UNIVERSITY OF COLORADO
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-12-21 至 2014-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8503359
• 关键词: Address; Affect; Antibodies; Applications Grants; Binding; Biological Assay; Biological Process; Cells; ChIP-seq; Chromatin; DNA; Disease; Functional RNA; Future; Gene Expression; Genes; Genetic Transcription; Genome; Goals; Heat-Shock Response; Herpesvirus 1; Hour; Human; Human Genome; Infection; Laboratories; Life; Malignant Neoplasms; Mammalian Cell; Measures; Mediating; Messenger RNA; Modeling; Mus; Names; Oligonucleotides; Pilot Projects; Polymerase; Precipitation; Process; Proteins; RNA; RNA Polymerase II; RNA Polymerase III; Repression; Research; Short Interspersed Nucleotide Elements; Techniques; Testing; Transact; Transcription Repressor/Corepressor; Viral Genes; Virus; Virus Diseases; Work; base; chromatin immunoprecipitation; gene repression; genome-wide; insight; interest; knock-down; mouse genome; programs; promoter; public health relevance; response

DESCRIPTION (provided by applicant): Herpes simplex virus type-1 (HSV-1) causes changes in host cell transcriptional programs during the hours after it infects mammalian cells. The transcription of protein-encoding genes by RNA polymerase II (Pol II) is largely repressed after HSV-1 infection, however the mechanism of this repression is not fully understood. It is hypothesized that specific non-coding RNAs (ncRNAs) that are upregulated during viral infection act as global, transacting repressors of Pol II transcription after HSV-1 infection. The specific ncRNAs proposed to act as repressors are mouse B2 RNA and human Alu RNA, which are transcribed by RNA polymerase III from short interspersed elements (SINEs) that litter the mouse and human genomes, respectively. This hypothesis is based on prior studies investigating the global repression of mRNA transcription that occurs during heat shock; B2 and Alu RNAs were found to directly repress Pol II transcription after heat shock in mouse and human cells, respectively. The pilot study described here seeks to determine whether B2 and Alu RNAs function in the repression of Pol II transcription after HSV-1 infection. The proposal has three highly focused Specific Aims. 1) Identify the Pol II-transcribed host cell genes that are repressed after HSV-1 infection of mouse and human cells by measuring changes in Pol II occupancy across the genomes before and after viral infection. 2) Determine whether B2 RNA and Alu RNA contribute to repression of host cell Pol II transcription after HSV-1 infection by knocking down the ncRNAs and determining the effect on Pol II occupancy across the genome before and after infection. 3) Determine whether B2 and Alu RNAs associate with Pol II and with the promoters of repressed genes in response to HSV-1 infection using two established cell-based assays. The proposed research will determine whether B2 and Alu RNAs mediate HSV-1 induced repression of host cell transcription. In testing this hypothesis, a genome-wide understanding of how HSV-1 infection reprograms Pol II occupancy on mRNA genes will be obtained, as well as an understanding of how HSV-1 affects the activity of B2 RNA and Alu RNA. The results of these pilot studies will enable a new line of research on the interplay between HSV-1 infection, SINE ncRNA transcription and function, host cell transcription, and transcription of viral genes, which will be the focus of a future R01 grant application. Lastly, since B2 RNA and Alu RNA levels are known to increase after infection by other viruses and in response to other biological processes, our studies could provide general insight into how these ncRNAs control transcription in multiple situations.

描述（由申请人提供）：1型单纯疱疹病毒（HSV-1）在感染哺乳动物细胞后数小时内引起宿主细胞转录程序的变化。HSV-1感染后，RNA聚合酶II（Pol II）对蛋白编码基因的转录在很大程度上受到抑制，但这种抑制的机制尚未完全了解。假设在病毒感染期间上调的特异性非编码RNA（ncRNA）在HSV-1感染后充当Pol II转录的全局反式作用阻遏物。被提议作为阻遏物的特异性ncRNA是小鼠B2 RNA和人Alu RNA，它们由RNA聚合酶III从分别散布在小鼠和人基因组中的短散布元件（西内斯）转录。这一假设是基于先前的研究，研究在热休克期间发生的mRNA转录的整体抑制; B2和Alu RNA被发现分别在小鼠和人细胞中热休克后直接抑制Pol II转录。这里描述的试点研究旨在确定B2和Alu RNA是否在HSV-1感染后抑制Pol II转录中起作用。该提案有三个高度集中的具体目标。1)鉴定Pol II转录的宿主细胞基因， 通过测量病毒感染前后基因组中Pol II占有率的变化，在HSV-1感染小鼠和人细胞后抑制。2)通过敲低ncRNA并确定感染前后对整个基因组Pol II占用率的影响，确定B2 RNA和Alu RNA是否有助于HSV-1感染后宿主细胞Pol II转录的抑制。3)使用两种已建立的基于细胞的检测方法，确定B2和Alu RNA是否与Pol II和受抑制基因的启动子相关，以响应HSV-1感染。这项研究将确定B2和Alu RNA是否介导HSV-1诱导的宿主细胞转录抑制。在检验这一假设时，将获得对HSV-1感染如何重新编程mRNA基因上的Pol II占用的全基因组理解，以及对HSV-1如何影响B2 RNA和Alu RNA活性的理解。这些试点研究的结果将使一个新的研究线之间的相互作用HSV-1感染，SINE ncRNA转录和功能，宿主细胞转录和病毒基因的转录，这将是未来的R 01赠款申请的重点。最后，由于已知B2 RNA和Alu RNA水平在被其他病毒感染后会增加，并响应其他生物过程，我们的研究可以提供这些ncRNA如何在多种情况下控制转录的一般见解。