Deciphering the Role of CPSF6 in HIV Infection

解读 CPSF6 在 HIV 感染中的作用

基本信息

批准号：
10646402
负责人：
Judd F Hultquist
金额：
$ 39.52万
依托单位：
NORTHWESTERN UNIVERSITY AT CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-07-13 至 2026-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10646402
关键词：
3&apos Untranslated Regions Agonist Binding CD4 Positive T Lymphocytes CRISPR/Cas technology Capsid Cell Line Cell Survival Cells Chemicals Cleavage And Polyadenylation Specificity Factor Code Complex Cyclophilin A Cytoplasm Data Defect Dependence Detection Development Distal Enhancers Genes Genetic Genetic Epistasis Genetic Transcription HIV HIV Infections HIV-1 Heterochromatin Hour Immune Immune response Infection Infection Control Innate Immune Response Integration Host Factors Interferons Knock-out Laboratory Finding Life Cycle Stages Link Mammalian Cell Mediating Messenger RNA Molecular Mutation Nuclear Nuclear Translocation Phase Phenocopy Phenotype Play Poly A Polyadenylation Polyadenylation Pathway Post-Transcriptional Regulation Post-Translational Protein Processing Proteins Regulation Regulatory Pathway Reporting Role Route Saquinavir Site T-Lymphocyte Testing Therapeutic Time Viral Virus Virus Replication Work cell type cleavage factor curative treatments design in vivo innate immune function integration site knock-down lead candidate member mutant overexpression permissiveness preference prevent recruit response small molecule trafficking transcriptome virus core

项目摘要

PROJECT SUMMARY Cleavage and polyadenylation specificity factor 6 (CPSF6) is an HIV host factor recruited to incoming viral cores during the early stage of the lifecycle. The interaction between HIV capsid (CA) and CPSF6 is known to dictate the cellular determinants of nuclear translocation and influence integration site preference, but its impact on overall viral infectivity is unclear. While overexpression of cytoplasmic CPSF6 has been shown to restrict viral replication, knock-down of CPSF6 or disruption of the CA-CPSF6 interaction has been reported to have a broad range of infectivity phenotypes. Recently, our lab found that knock-out of CPSF6 in primary CD4+ T cells dramatically increases HIV replication with minimal impact on cell viability. This increase in replication correlates with decreased induction of interferon-stimulated genes, contrary to prior reports that suggest CPSF6 acts to shield the virus from immune recognition. In this proposal, we are testing the overall hypothesis that CPSF6 plays a critical role in regulating the innate immune response to HIV infection and that the virus recruits CPSF6 in part to circumvent this response. This hypothesis will be tested in three aims that broadly seek to understand the mechanism by which loss of CPSF6 dampens the immune response to infection, the impact of HIV infection on CPSF6 function, and the potential role of CPSF6 regulatory pathways in controlling infection. In Aim 1, we will test the hypothesis that loss of CPSF6 acts to dampen the immune response directly by induction of alternative polyadenylation (APA) or indirectly by allowing enhanced recruitment of Cyclophilin A (CYPA) to protect the core from restriction by the antiviral factor TRIM5. CPSF6 normally acts as a member of the CFIm cleavage factor complex to direct polyadenylation to distal sites of the 3' untranslated region (UTR). Inhibition of CFIm activity triggers APA, which has been previously implicated in the regulation of the innate immune response, and could explain the observed phenotype. Alternatively, there is evidence to suggest that CPSF6 and CYPA compete for core binding and loss of CYPA binding has been previously linked to enhanced restriction and innate sensing. In Aim 2, we will test the hypothesis that CPSF6 recruitment by incoming viral cores can alter overall CFIm function and induce APA. Regardless if this is linked to dampening of the immune response above, it is well established that other viruses hijack the APA pathway to enhance their replication, though this hasn't been explored during HIV infection. Finally, in Aim 3, we will test the hypothesis that perturbation of the CPSF6 regulatory network can control viral infectivity and the immune response to infection. CPSF6 activity is regulated by post-translational modification and nuclear-cytoplasmic shuttling. Truncation mutants of CPSF6 that force cytoplasmic localization have been shown to restrict HIV infection, and we will test if we can mimic that effect by genetic and/or chemical perturbation of its endogenous regulatory mechanisms. Altogether, this work explores a newly described phenotype for a well-known HIV host factor, CPSF6, and seeks to provide a better understanding of viral manipulation of the innate immune response by post-transcriptional regulation.

项目摘要切割和聚腺苷酸化特异性因子6（CPSF6）是招募到传入病毒核心的HIV宿主因子在生命周期的早期。已知HIV CAPSID（CA）和CPSF6之间的相互作用决定了核易位和影响整合现场偏好的细胞决定剂，但其影响总体病毒感染尚不清楚。虽然已证明细胞质CPSF6的过表达限制病毒据报道，复制，CPSF6的敲击或CA-CPSF6相互作用的破坏具有广泛的感染表型范围。最近，我们的实验室发现，在初级CD4+ T细胞中CPSF6的敲除动态增加HIV复制，对细胞活力的影响最小。复制的增加与随着干扰素刺激的基因的诱导降低，与表明CPSF6作用为的先前报道形成了鲜明对比保护病毒免疫识别。在此提案中，我们正在测试CPSF6的总体假设在调查先天免疫对艾滋病毒感染的反应中起着至关重要的作用，该病毒报告了CPSF6 部分是为了避免这种响应。该假设将在三个目标中进行检验 CPSF6丧失会抑制对感染的免疫反应的机制，HIV感染的影响在CPSF6功能上，以及CPSF6调节途径在控制感染中的潜在作用。在AIM 1中，我们将检验以下假设：CPSF6的丢失是通过诱导直接使免疫反应的替代聚腺苷酸化（APA）或间接通过允许增强环素A（CYPA）的募集到保护核心免受抗病毒因子TRIM5的限制。 CPSF6通常充当CFIM的成员裂解因子复合物将聚腺苷酸化直达3'未翻译区（UTR）的远端位点。抑制 CFIM活动触发APA，该APA先前已在先天免疫的调节中实施反应，可以解释观察到的表型。或者，有证据表明CPSF6 CYPA竞争核心结合和CYPA结合的丧失，以前已与增强的限制有关和天生的感官。在AIM 2中，我们将检验以下假设：CPSF6通过传入病毒核的募集可以改变总体CFIM函数并诱导APA。不管这是否与免疫反应的衰减有关上面，有很好的确定，其他病毒劫持了APA途径以增强其复制，尽管这是在艾滋病毒感染期间尚未探索。最后，在AIM 3中，我们将检验以下假设。 CPSF6调节网络可以控制病毒感染和对感染的免疫响应。 CPSF6活动是受翻译后修饰和核质班级穿梭的调节。 CPSF6的截断突变体已经证明，力细胞质定位限制了HIV感染，我们将测试是否可以模仿其内源性调节机制的遗传和/或化学扰动作用。总共，这项工作探索众所周知的艾滋病毒宿主因子CPSF6的新描述的表型，并试图提供更好的通过转录后调节对先天免疫响应的病毒操纵的理解。