Effects of HIV Rev on Host Cell Gene Expression

HIV Rev 对宿主细胞基因表达的影响

• 批准号: 10673153
• 负责人: MARIE-LOUISE HAMMARSKJOLD
• 金额: $ 16.15万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10673153
• 关键词: Affect; Algorithms; Alternative Splicing; Binding; Bioinformatics; CD4 Positive T Lymphocytes; Cell physiology; Cells; Coupled; Cytoplasm; Data Analyses; Development; Electrophoretic Mobility Shift Assay; Elements; Endogenous Retroviruses; Gene Expression; Genes; HIV; HIV Infections; Human; Immune response; In Vitro; Infection; Introns; Libraries; Mediating; Messenger RNA; Methods; Natural Immunity; Paper; Peripheral; Play; Post-Transcriptional Regulation; Process; Protein Isoforms; RNA; RNA Sequences; RNA Splicing; Response Elements; Retroviral Vector; Retroviridae; Role; Structure; System; T-Lymphocyte; Testing; Tissue-Specific Gene Expression; Transcript; Translating; Translations; Untranslated RNA; Viral; Virus Replication; bioinformatics tool; cell growth regulation; cellular transduction; cis acting element; experimental study; in vivo; mRNA Expression; novel; novel therapeutics; posttranscriptional; response; rev Protein; shape analysis; transcriptome; transcriptome sequencing; vector; viral RNA

It is well established that all retroviruses have evolved processes to overcome the usual regulatory mechanisms of the cell that control or restrict the nucleocytoplasmic export of RNA until splicing is complete. A hallmark of every retroviral transcriptome is that viral mRNA with at least one retained intron reaches the cytoplasm. In the case of HIV infected cells, many different viral RNAs with one or two retained introns appear in the cytoplasm, and it is the binding of the Rev protein to a cis-acting element, the Rev Response Element (RRE), present in all of these RNAs, that facilitates this process. This proposal will explore the hypothesis that, in addition to facilitating the export and expression of viral RNAs with retained introns, Rev interacts with RNA elements present in bona fide cellular genes and human endogenous retroviruses, to overcome the mechanisms that normally control and restrict the export of RNA isoforms with retained introns. Novel RNA isoforms, whose expression has been dysregulated by Rev, may function to change the milieu of the infected cell to affect innate immunity and/or promote viral replication, either directly, or by being translated into a novel protein isoforms. The proposal has 2 specific aims: In Aim #1 we will identify cellular RNAs that functionally interact with Rev. This will be accomplished through the use of a novel vector-trap system. This will allow the isolation of cellular sequences that function like RREs (cRREs) and identify genes with potential to be regulated by Rev. In Aim #2 we will perform RNASeq on SupT1 cells transduced with retroviral vectors that express Rev or both Tat and Rev and identify genes that have novel RNA isoforms expressed in the cytoplasm. At the completion of this R21, we expect to have determined whether HIV Rev can induce the cytoplasmic expression of novel RNA species at the post-transcriptional level through interaction with cRREs. Further experiments, beyond the scope of this two year proposal, will analyze how this novel Rev-mediated post-transcriptional regulation of cellular RNAs influences HIV replication and cellular functions.

众所周知，所有逆转录病毒都有进化过程来克服通常的监管 细胞控制或限制 RNA 核胞质输出直至剪接完成的机制。一个 每个逆转录病毒转录组的标志是至少有一个保留内含子的病毒 mRNA 到达 细胞质。对于 HIV 感染的细胞，会出现许多带有一两个保留内含子的不同病毒 RNA 在细胞质中，它是 Rev 蛋白与顺式作用元件（Rev 响应元件）的结合 (RRE)，存在于所有这些 RNA 中，促进了这一过程。该提案将探讨以下假设： 除了促进保留内含子的病毒 RNA 的输出和表达外，Rev 还与 RNA 相互作用 存在于真正的细胞基因和人内源性逆转录病毒中的元素，以克服 通常控制和限制带有保留内含子的 RNA 同工型输出的机制。新型RNA 亚型的表达已被 Rev 失调，可能会改变感染者的环境 细胞直接影响先天免疫和/或促进病毒复制，或通过转化为新的 蛋白质亚型。 该提案有 2 个具体目标： 在目标 #1 中，我们将鉴定与 Rev 功能性相互作用的细胞 RNA。这将得以实现 通过使用新型载体陷阱系统。这将允许分离具有功能的细胞序列 像 RRE (cRRE) 一样，并识别有可能受到 Rev 调控的基因。在目标 #2 中，我们将执行 用表达 Rev 或同时表达 Tat 和 Rev 的逆转录病毒载体转导的 SupT1 细胞上的 RNASeq 并识别 具有在细胞质中表达的新RNA亚型的基因。 完成 R21 后，我们希望确定 HIV Rev 是否​​可以诱导 通过与 cRRE 相互作用，在转录后水平上表达新型 RNA 种类的细胞质。 超出这两年提案范围的进一步实验将分析这种新颖的 Rev 介导的 细胞 RNA 的转录后调控影响 HIV 复制和细胞功能。