Host factors regulating HIV latency and reactivation

调节HIV潜伏期和再激活的宿主因素

• 批准号: 10516096
• 负责人: Susana T Valente
• 金额: $ 48.3万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-11-01 至 2026-10-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10516096
• 关键词: Acceleration; Affect; Affinity Chromatography; Automobile Driving; Binding; Binding Proteins; Biological; CD4 Positive T Lymphocytes; Cell model; Cells; Cellular Metabolic Process; ChIP-seq; Chromatin; Chromatin Remodeling Factor; Chromatin Structure; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; Dependence; ERCC3 gene; Elements; Environment; Epigenetic Process; Generic Drugs; Genes; Genetic Transcription; Goals; HIV; HIV Genome; HIV-1; Heterochromatin; Impairment; In Vitro; Individual; Integration Host Factors; Interruption; Knock-out; Knowledge; Link; Mass Spectrum Analysis; Mediating; Messenger RNA; Modification; Molecular; Molecular Conformation; Nucleosomes; Pathway interactions; Pharmaceutical Preparations; Positioning Attribute; Post-Translational Protein Processing; Productivity; Proliferating; Proteins; Proviruses; RNA Polymerase II; RNA Splicing; Reporting; Reproducibility; Residual state; Role; Spironolactone; Testing; Transactivation; Transcription Coactivator; Translational Protein Modification; Viral; Viral Proteins; Viremia; acrosome stabilizing factor; antiretroviral therapy; cortistatin; detection limit; epigenetic silencing; follow-up; in vivo; in vivo Model; inhibitor; insight; knock-down; novel; overexpression; prohibitin; promoter; protein complex; reactivation from latency; recruit; small hairpin RNA; small molecule; targeted agent; transcription factor; transcription factor TFIIH; transcriptomics; viral rebound

Abstract Our central premise is that HIV transcriptional inhibitors can be used as latency promoting agents (LPAs) in block-and-lock functional cure approaches, aimed at reducing residual viremia during antiretroviral therapy (ART) and limiting viral rebound upon treatment interruption (TI). This hypothesis came about with the discovery of the Tat inhibitor, didehydro-Cortistatin A (dCA). In in vitro and in vivo models of HIV latency combining ART with dCA accelerated HIV-1 suppression to below the limit of detection and blocked viral rebound upon TI or stimulation with latency reversing agents (LRAs). The transcriptional shutdown by dCA resulted in the heterochromatinization and loss of RNAPII at the HIV promoter. We recently reported on another LPA, the generic drug Spironolactone (SP). Interestingly this drug degrades the host XPB subunit of the general transcription factor TFIIH inhibiting HIV transcription in primary cells from infected individuals and blocking viral reactivation from latency without affecting cellular transcriptomics7. This study is important because it highlights that a host factor can be targeted to silence HIV without affecting cellular viability. In sum, the more knowledge we gain on the interconnectivity between Tat, transcription factors regulatory mechanisms and how these affect nucleosomes positioning around the HIV promoter, the more we can leverage transcriptional regulators as antiviral targets. To identify novel host factors regulating HIV transcription we used a chromatin affinity purification approach together with mass spectrometry (ChAP-MS). We identified p32 (ASF/SF2 splicing factor-associated protein) and FUBP3 (Far upstream element binding protein 3) in productively infected ART treated promoters but not in dCA-treated where HIV was silenced; and PHB2 (prohibitin-2) was enriched in dCA treated promoters but reduced in ART only treated ones. These results suggest that p32 and FUBP3 are HIV transcriptional activators while PHB2 is a HIV silencing factor. Preliminary studies confirmed their transcriptional activity on HIV. Here we propose a detailed molecular mechanistic study on how p32 and FUBP3 enhance and how PHB2 inhibits HIV transcription. Understanding their role in pre-initiation complex (PIC) formation, initiation, elongation and nucleosome organization at the latent HIV promoter will be key for developing successful strategies for durable HIV silencing. Strong effects of one or more of these factors will prompt the search for small-molecule modulators. Our overarching goal is to durably silence latent HIV proviruses by driving each into stable heterochromatin where they will remain “locked.” We propose the following aims: Aim 1: Define the roles of p32, FUBP3 and PHB2 in HIV transcription and chromatin structure. Aim 2: Identify protein complexes associated with p32, FUBP3 and PHB2 and study interaction with Tat. Aim 3: Investigate the “block-and-lock” approach using single or combinations of factors and their dependence on Tat/TAR circuitry.

摘要