Although highly active retroviral therapy (HAART) dramatically reduces viral load and restores CD4 T-cell counts in HIV-infected individuals, its success is hampered by the fact that HIV can establish a latent proviral reservoir within long-lived resting memory CD4 lymphocytes. The molecular underpinnings of HIV latency involve multiple host factors and pathways actively repressing HIV transcription and maintaining the proviral genome in a transcriptionally inert state. Many host factors have been implicated based on genome-wide screens in cell-line based models of latency. But the exact role of these candidate genes in HIV latency occurring within resting primary CD4 T cells, remain unknown. The evaluation of these genes in resting primary CD4 T cells is hampered by the fact that shRNAs cannot be introduced into these cells using lentiviral vectors. In this collaboration among three CARE investigators (Warner C. Greene, Jonathan Karn, Vincente Planelles), we now propose to utilize a novel technology based on HIV-2 Vpx to render resting CD4 T cells permissive to lentivirus infection. This technology will permit efficient knockdown and evaluation of the candidate regulatory genes in resting primary CD4 T cells. This approach first involves pretreating cells with virion like particles (VLP) encapsulating Vpx to remove anti-viral reverse transcription block mediated by SAMHD1 followed by shRNA lentiviral transduction. A number of potential positive and negative regulators of HIV latency identified in CARE-sponsored studies will be systematically tested including the genome organizer and transcription repressor CTCF, select receptor subunits in the immunosuppressive PDCD1 and pro-inflammatory interleukin IL22RA pathways, and a putative Tat cofactor termed TRIM32. The evaluation of these cellular genes in primary CD4 T cell models of HIV latency formed in memory CD4 T cells, Th17 CD4 T cells and Treg CD4 T cells promises to provide key insights into the role of these genes in HIV latency. These studies could lay the foundation for finding small molecules that when combined will promote effective purging of the latent reservoir in patients while not producing a general state of cellular activation.

Public Health Relevance

Although highly active retroviral therapy (HAART) can dramatically extend the life expectancies of HIVinfected individuals, these drugs cannot cure patients because of the presence of a latent reservoir of druginsensitive virus. We propose testing the involvement of various gene candidates in primary CD4 T cells using a novel Vpx-based technology to introduce lentiviral encoded shRNAs into these cells. These studies promise to provide key insights into the molecular underpinnings of HIV latency that will inform the search for small molecule cocktails capable of effectively purging latent HIV proviruses.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Program--Cooperative Agreements (U19)
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Special Emphasis Panel (ZAI1-JBS-A (M1))
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University of North Carolina Chapel Hill
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Davis, Zachary B; Cogswell, Andrew; Scott, Hamish et al. (2016) A Conserved HIV-1-Derived Peptide Presented by HLA-E Renders Infected T-cells Highly Susceptible to Attack by NKG2A/CD94-Bearing Natural Killer Cells. PLoS Pathog 12:e1005421
Imaz, Arkaitz; Martinez-Picado, Javier; Niubó, Jordi et al. (2016) HIV-1-RNA Decay and Dolutegravir Concentrations in Semen of Patients Starting a First Antiretroviral Regimen. J Infect Dis 214:1512-1519
Honeycutt, Jenna B; Wahl, Angela; Baker, Caroline et al. (2016) Macrophages sustain HIV replication in vivo independently of T cells. J Clin Invest 126:1353-66
Tokarev, Andrey; Stoneham, Charlotte; Lewinski, Mary K et al. (2016) Pharmacologic Inhibition of Nedd8 Activation Enzyme Exposes CD4-Induced Epitopes within Env on Cells Expressing HIV-1. J Virol 90:2486-502
Victor Garcia, J (2016) Humanized mice for HIV and AIDS research. Curr Opin Virol 19:56-64
Smith, Davey M; Nakazawa, Masato; Freeman, Michael L et al. (2016) Asymptomatic CMV Replication During Early Human Immunodeficiency Virus (HIV) Infection Is Associated With Lower CD4/CD8 Ratio During HIV Treatment. Clin Infect Dis 63:1517-1524
Gianella, Sara; Anderson, Christy M; Var, Susanna R et al. (2016) Replication of Human Herpesviruses Is Associated with Higher HIV DNA Levels during Antiretroviral Therapy Started at Early Phases of HIV Infection. J Virol 90:3944-52
Garcia, J Victor (2016) In vivo platforms for analysis of HIV persistence and eradication. J Clin Invest 126:424-31
Lee, Sook-Kyung; Zhou, Shuntai; Baldoni, Pedro L et al. (2016) Quantification of the Latent HIV-1 Reservoir Using Ultra Deep Sequencing and Primer ID In A Viral Outgrowth Assay. J Acquir Immune Defic Syndr :
Wagner, Gabriel A; Chaillon, Antoine; Liu, Siqi et al. (2016) HIV-associated neurocognitive disorder is associated with HIV-1 dual infection. AIDS 30:2591-2597

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