scholarly journals Shared Mechanisms Govern HIV Transcriptional Suppression in Circulating CD103+ and Gut CD4+ T Cells

2020 ◽  
Vol 95 (2) ◽  
pp. e01331-20
Author(s):  
Steven A. Yukl ◽  
Shahzada Khan ◽  
Tsui-Hua Chen ◽  
Martin Trapecar ◽  
Frank Wu ◽  
...  
Keyword(s):  
T Cells ◽  
Hiv Infection ◽  
Cd4 T Cells ◽  
Molecular Mechanisms ◽  
Hiv Latency ◽  
Expression Levels ◽  
Hiv Rna ◽  
Infected Cells ◽  
Latent Hiv ◽  
Main Barrier

ABSTRACTLatent HIV infection is the main barrier to cure, and most HIV-infected cells reside in the gut, where distinct but unknown mechanisms may promote viral latency. Transforming growth factor β (TGF-β), which induces the expression of CD103 on tissue-resident memory T cells, has been implicated in HIV latency. Using CD103 as a surrogate marker to identify cells that have undergone TGF-β signaling, we compared the HIV RNA/DNA contents and cellular transcriptomes of CD103+ and CD103− CD4 T cells from the blood and rectum of HIV-negative (HIV−) and antiretroviral therapy (ART)-suppressed HIV-positive (HIV+) individuals. Like gut CD4+ T cells, circulating CD103+ cells harbored more HIV DNA than did CD103− cells but transcribed less HIV RNA per provirus. Circulating CD103+ cells also shared a gene expression profile that is closer to that of gut CD4 T cells than to that of circulating CD103− cells, with significantly lower expression levels of ribosomal proteins and transcriptional and translational pathways associated with HIV expression but higher expression levels of a subset of genes implicated in suppressing HIV transcription. These findings suggest that blood CD103+ CD4 T cells can serve as a model to study the molecular mechanisms of HIV latency in the gut and reveal new cellular factors that may contribute to HIV latency.IMPORTANCE The ability of HIV to establish a reversibly silent, “latent” infection is widely regarded as the main barrier to curing HIV. Most HIV-infected cells reside in tissues such as the gut, but it is unclear what mechanisms maintain HIV latency in the blood or gut. We found that circulating CD103+ CD4+ T cells are enriched for HIV-infected cells in a latent-like state. Using RNA sequencing (RNA-seq), we found that CD103+ T cells share a cellular transcriptome that more closely resembles that of CD4+ T cells from the gut, suggesting that they are homing to or from the gut. We also identified the cellular genes whose expression distinguishes gut CD4+ or circulating CD103+ T cells from circulating CD103− T cells, including some genes that have been implicated in HIV expression. These genes may contribute to latent HIV infection in the gut and may serve as new targets for therapies aimed at curing HIV.

scholarly journals Epigenomic characterization of latent HIV infection identifies latency regulating transcription factors

2021 ◽  
Vol 17 (2) ◽  
pp. e1009346
Author(s):  
Stuart R. Jefferys ◽  
Samuel D. Burgos ◽  
Jackson J. Peterson ◽  
Sara R. Selitsky ◽  
Anne-Marie W. Turner ◽  
...  
Keyword(s):  
Gene Expression ◽  
T Cells ◽  
Transcription Factors ◽  
Hiv Infection ◽  
Cd4 T Cells ◽  
Viral Gene Expression ◽  
Viral Gene ◽  
Hiv Latency ◽  
Latently Infected ◽  
Infected Cells

Transcriptional silencing of HIV in CD4 T cells generates a reservoir of latently infected cells that can reseed infection after interruption of therapy. As such, these cells represent the principal barrier to curing HIV infection, but little is known about their characteristics. To further our understanding of the molecular mechanisms of latency, we characterized a primary cell model of HIV latency in which infected cells adopt heterogeneous transcriptional fates. In this model, we observed that latency is a stable, heritable state that is transmitted through cell division. Using Assay of Transposon-Accessible Chromatin sequencing (ATACseq) we found that latently infected cells exhibit greatly reduced proviral accessibility, indicating the presence of chromatin-based structural barriers to viral gene expression. By quantifying the activity of host cell transcription factors, we observe elevated activity of Forkhead and Kruppel-like factor transcription factors (TFs), and reduced activity of AP-1, RUNX and GATA TFs in latently infected cells. Interestingly, latency reversing agents with different mechanisms of action caused distinct patterns of chromatin reopening across the provirus. We observe that binding sites for the chromatin insulator CTCF are highly enriched in the differentially open chromatin of infected CD4 T cells. Furthermore, depletion of CTCF inhibited HIV latency, identifying this factor as playing a key role in the initiation or enforcement of latency. These data indicate that HIV latency develops preferentially in cells with a distinct pattern of TF activity that promotes a closed proviral structure and inhibits viral gene expression. Furthermore, these findings identify CTCF as a novel regulator of HIV latency.

scholarly journals Biogenesis of P-TEFb in CD4+ T cells to reverse HIV latency is mediated by protein kinase C (PKC)-independent signaling pathways

2021 ◽  
Author(s):  
Uri Mbonye ◽  
Konstantin Leskov ◽  
Meenakshi Shukla ◽  
Saba Valadkhan ◽  
Jonathan Karn
Keyword(s):  
T Cells ◽  
Protein Kinase C ◽  
T Cell ◽  
Protein Kinase ◽  
Signaling Pathways ◽  
Cd4 T Cells ◽  
Hiv Latency ◽  
Kinase C ◽  
Latent Hiv ◽  
Memory Cd4 T Cells

The switch between HIV latency and productive transcription is regulated by an auto-feedback mechanism initiated by the viral trans-activator Tat, which functions to recruit the host transcription elongation factor P-TEFb to proviral HIV. A heterodimeric complex of CDK9 and one of three cyclin T subunits, P-TEFb is expressed at vanishingly low levels in resting memory CD4 + T cells and cellular mechanisms controlling its availability are central to regulation of the emergence of HIV from latency. Using a well-characterized primary T-cell model of HIV latency alongside healthy donor memory CD4 + T cells, we characterized specific T-cell receptor (TCR) signaling pathways that regulate the generation of transcriptionally active P-TEFb, defined as the coordinate expression of cyclin T1 and phospho-Ser175 CDK9. Protein kinase C (PKC) agonists, such as ingenol and prostratin, stimulated active P-TEFb expression and reactivated latent HIV with minimal cytotoxicity, even in the absence of intracellular calcium mobilization with an ionophore. Unexpectedly, inhibition-based experiments demonstrated that PKC agonists and TCR-mobilized diacylglycerol signal through MAP kinases ERK1/2 rather than through PKC to effect the reactivation of both P-TEFb and latent HIV. Single-cell and bulk RNA-seq analyses revealed that of the four known isoforms of the Ras guanine nucleotide exchange factor RasGRP, RasGRP1 is by far the predominantly expressed diacylglycerol-dependent isoform in CD4 + T cells. RasGRP1 should therefore mediate the activation of ERK1/2 via Ras-Raf signaling upon TCR co-stimulation or PKC agonist challenge. Combined inhibition of the PI3K-mTORC2-AKT-mTORC1 pathway and the ERK1/2 activator MEK prior to TCR co-stimulation abrogated active P-TEFb expression and substantially suppressed latent HIV reactivation. Therefore, contrary to prevailing models, the coordinate reactivation of P-TEFb and latent HIV in primary T cells following either TCR co-stimulation or PKC agonist challenge is independent of PKC but rather involves two complementary signaling arms of the TCR cascade, namely, RasGRP1-Ras-Raf-MEK-ERK1/2 and PI3K-mTORC2-AKT-mTORC1.

scholarly journals Endogenous factors enhance HIV infection of tissue naive CD4 T cells by stimulating high molecular mass APOBEC3G complex formation

2006 ◽  
Vol 203 (4) ◽  
pp. 865-870 ◽  
Author(s):  
Jason F. Kreisberg ◽  
Wes Yonemoto ◽  
Warner C. Greene
Keyword(s):  
T Cells ◽  
Hiv Infection ◽  
Molecular Mass ◽  
Cd4 T Cells ◽  
Molecular Mechanisms ◽  
High Molecular Mass ◽  
Lymphoid Tissues ◽  
Soluble Factors ◽  
Endogenous Factors ◽  
Memory Cd4 T Cells

Human immunodeficiency virus (HIV) can infect resting CD4 T cells residing in lymphoid tissues but not those circulating in peripheral blood. The molecular mechanisms producing this difference remain unknown. We explored the potential role of the tissue microenvironment and its influence on the action of the antiviral factor APOBEC3G (A3G) in regulating permissivity to HIV infection. We found that endogenous IL-2 and -15 play a key role in rendering resident naive CD4 T cells susceptible to HIV infection. Infection of memory CD4 T cells also requires endogenous soluble factors, but not IL-2 or -15. A3G is found in a high molecular mass complex in HIV infection–permissive, tissue-resident naive CD4 T cells but resides in a low molecular mass form in nonpermissive, blood-derived naive CD4 T cells. Upon treatment with endogenous soluble factors, these cells become permissive for HIV infection, as low molecular mass A3G is induced to assemble into high molecular mass complexes. These findings suggest that in lymphoid tissues, endogenous soluble factors, likely including IL-2 and -15 and others, stimulate the formation of high molecular mass A3G complexes in tissue-resident naive CD4 T cells, thereby relieving the potent postentry restriction block for HIV infection conferred by low molecular mass A3G.

Asymptomatic HIV infection is characterized by rapid turnover of HIV RNA in plasma and lymph nodes but not of latently infected lymph-node CD4+ T cells

AIDS ◽  
1997 ◽  
Vol 11 (9) ◽  
pp. 1103-1110 ◽  
Author(s):  
Hans-Jürgen Stellbrink ◽  
Jan van Lunzen ◽  
Frank T. Hufert ◽  
Günter Fröschle ◽  
Guido Wolf-Vorbeck ◽  
...  
Keyword(s):  
T Cells ◽  
Lymph Node ◽  
Hiv Infection ◽  
Lymph Nodes ◽  
Cd4 T Cells ◽  
Rapid Turnover ◽  
Hiv Rna ◽  
Latently Infected ◽  
Infected Lymph Node ◽  
Asymptomatic Hiv

scholarly journals HIV Populations Shift After Tuberculosis Associated Immune Reconstitution Inflammatory Syndrome: Implications For HIV Remission

2020 ◽  
Author(s):  
Camille Lange ◽  
Maura Manion ◽  
Natalie Lindo ◽  
Robert Gorelick ◽  
Ana Ortega-Villa ◽  
...  
Keyword(s):  
T Cells ◽  
Immune Reconstitution Inflammatory Syndrome ◽  
Cd4 T Cells ◽  
Immune Reconstitution ◽  
Long Term Effects ◽  
Genetic Characteristics ◽  
Hiv Rna ◽  
Hiv Dna ◽  
Infected Cells ◽  
Inflammatory Syndrome

Abstract Tuberculosis associated immune reconstitution inflammatory syndrome (TB-IRIS) is a serious complication of starting combination antiretroviral therapy (cART). TB-IRIS emerges early after cART initiation and is characterized by rapid expansions of TB-specific CD4+ T cells and high levels of inflammatory mediators driven by CD4+ T cells. The effects of TB-IRIS on HIV populations are unknown, but could result in profound expansion and elimination of HIV infected cells via cellular activation and acute inflammation. We investigated immediate and long-term effects of TB-IRIS on HIV infected cells with and without TB-IRIS. We measured plasma HIV RNA, cell-associated HIV RNA and HIV DNA levels and compared genetic characteristics of HIV populations after prolonged cART. We found that TB-IRIS was associated with more diverse HIV DNA populations and HIV reservoirs after IRIS were distinct from pre-therapy populations, suggesting that TB-IRIS can shape the HIV reservoir with detrimental implications for HIV remission strategies.

scholarly journals Clonal Expansion of Infected CD4+ T Cells in People Living with HIV

Viruses ◽  
2021 ◽  
Vol 13 (10) ◽  
pp. 2078
Author(s):  
John M. Coffin ◽  
Stephen H. Hughes
Keyword(s):  
T Cells ◽  
Cd4 T Cells ◽  
Large Fraction ◽  
Latent Reservoir ◽  
People Living With Hiv ◽  
Resting Cells ◽  
Hiv Rna ◽  
Art Initiation ◽  
Infected Cells ◽  
Living With Hiv

HIV infection is not curable with current antiretroviral therapy (ART) because a small fraction of CD4+ T cells infected prior to ART initiation persists. Understanding the nature of this latent reservoir and how it is created is essential to development of potentially curative strategies. The discovery that a large fraction of the persistently infected cells in individuals on suppressive ART are members of large clones greatly changed our view of the reservoir and how it arises. Rather than being the products of infection of resting cells, as was once thought, HIV persistence is largely or entirely a consequence of infection of cells that are either expanding or are destined to expand, primarily due to antigen-driven activation. Although most of the clones carry defective proviruses, some carry intact infectious proviruses; these clones comprise the majority of the reservoir. A large majority of both the defective and the intact infectious proviruses in clones of infected cells are transcriptionally silent; however, a small fraction expresses a few copies of unspliced HIV RNA. A much smaller fraction is responsible for production of low levels of infectious virus, which can rekindle infection when ART is stopped. Further understanding of the reservoir will be needed to clarify the mechanism(s) by which provirus expression is controlled in the clones of cells that constitute the reservoir.

scholarly journals Human splice factors contribute to latent HIV infection in primary cell models and blood CD4+ T cells from ART-treated individuals

2020 ◽  
Vol 16 (11) ◽  
pp. e1009060
Author(s):  
Sara Moron-Lopez ◽  
Sushama Telwatte ◽  
Indra Sarabia ◽  
Emilie Battivelli ◽  
Mauricio Montano ◽  
...  
Keyword(s):  
T Cells ◽  
Hiv Infection ◽  
Primary Cell ◽  
Cell Models ◽  
Transcriptional Initiation ◽  
Cellular Genes ◽  
Latently Infected ◽  
Infected Cells ◽  
Latent Hiv

It is unclear what mechanisms govern latent HIV infection in vivo or in primary cell models. To investigate these questions, we compared the HIV and cellular transcription profile in three primary cell models and peripheral CD4+ T cells from HIV-infected ART-suppressed individuals using RT-ddPCR and RNA-seq. All primary cell models recapitulated the block to HIV multiple splicing seen in cells from ART-suppressed individuals, suggesting that this may be a key feature of HIV latency in primary CD4+ T cells. Blocks to HIV transcriptional initiation and elongation were observed more variably among models. A common set of 234 cellular genes, including members of the minor spliceosome pathway, was differentially expressed between unstimulated and activated cells from primary cell models and ART-suppressed individuals, suggesting these genes may play a role in the blocks to HIV transcription and splicing underlying latent infection. These genes may represent new targets for therapies designed to reactivate or silence latently-infected cells.

scholarly journals A two-color haploid genetic screen identifies novel host factors involved in HIV latency

2021 ◽  
Author(s):  
Michael D Röling ◽  
Mahsa Mollapour Sisakht ◽  
Enrico Ne ◽  
Panagiotis Moulos ◽  
Mateusz Stoszko ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Line ◽  
Cd4 T Cells ◽  
Genetic Screen ◽  
Host Factors ◽  
Hiv Latency ◽  
Novel Host ◽  
Latent Hiv ◽  
Hiv 1

AbstractTo identify novel host factors as putative targets to reverse HIV latency, we performed an insertional mutagenesis genetic screen in a latently HIV-1-infected pseudo-haploid KBM7 cell line (Hap-Lat). Following mutagenesis, insertions were mapped to the genome and bioinformatic analysis resulted in the identification of 69 candidate host genes involved in maintaining HIV-1 latency. A select set of candidate genes was functionally validated using shRNA mediated depletion in latent HIV-1 infected J-Lat A2 and 11.1 T cell lines. We confirmed ADK, CHD9, CMSS1, EVI2B, EXOSC8, FAM19A, GRIK5, IRF2BP2, NF1, and USP15 as novel host factors involved in the maintenance of HIV latency. Chromatin immunoprecipitation assays indicated that CHD9, a Chromodomain Helicase DNA-binding protein, maintains HIV latency via direct association with the HIV 5’LTR, and its depletion results in increased histone acetylation at the HIV-1 promoter, concomitant with HIV-1 latency reversal. FDA-approved inhibitors 5-Iodotubercidin, Trametinib, and Topiramate, targeting ADK, NF1, and GRIK5, respectively were characterized for their latency reversal potential. While 5-Iodotubercidin exhibited significant cytotoxicity in both J-Lat and primary CD4+ T cells, Trametinib reversed latency in J-Lat cells but not in latently HIV-1-infected primary CD4+ T cells. Crucially, Topiramate reversed latency in cell-line models and latently infected primary CD4+ T cells, without inducing T cell activation or significant toxicity. Thus, using an adaptation of a haploid forward genetic screen, we identified novel and druggable host factors contributing to HIV-1 latency.ImportanceA reservoir of latent HIV-1-infected cells persists in the presence of combination antiretroviral therapy (cART), representing a major obstacle for viral eradication. Reactivation of the latent HIV-1 provirus is part of curative strategies which aim to promote clearance of the infected cells. Using a two-color haploid screen, we identified 69 candidate genes as latency maintaining host factors and functionally validated a subset of 10 of those in additional T-cell based cell line models of HIV-1 latency. We further demonstrated that CHD9 is associated with HIV-1’s promoter, the 5’LTR while this association is lost upon reactivation. Additionally, we characterized the latency reversal potential of FDA compounds targeting ADK, NF1, and GRIK5 and identify the GRIK5 inhibitor Topiramate as a viable latency reversal agent with clinical potential.

scholarly journals Telomere and ATM Dynamics in CD4 T-Cell Depletion in Active and Virus-Suppressed HIV Infections

2020 ◽  
Vol 94 (22) ◽  
Author(s):  
Sushant Khanal ◽  
Qiyuan Tang ◽  
Dechao Cao ◽  
Juan Zhao ◽  
Lam Nhat Nguyen ◽  
...  
Keyword(s):  
Dna Damage ◽  
T Cells ◽  
T Cell ◽  
Hiv Infection ◽  
Cell Apoptosis ◽  
Cd4 T Cells ◽  
Molecular Mechanisms ◽  
Cd4 T Cell ◽  
Cell Aging

ABSTRACT CD4 T-cell depletion is a hallmark of HIV/AIDS, but the underlying mechanism is still unclear. We have recently shown that ataxia-telangiectasia-mutated (ATM) deficiency in CD4 T cells accelerates DNA damage, telomere erosion, and cell apoptosis in HIV-infected individuals on antiretroviral therapy (ART). Whether these alterations in ART-treated HIV subjects occur in vitro in HIV-infected CD4 T cells remains unknown. In this study, we employed a cellular model of HIV infection to characterize the mechanisms underlying CD4 T-cell destruction by analyzing the telomeric DNA damage response (DDR) and cellular apoptosis in highly permissive SupT1 cells, followed by the validation of our observations in primary CD4 T cells with active or drug-suppressed HIV infection. Specifically, we established an in vitro HIV T-cell culture system with viral replication and raltegravir (RAL; an integrase inhibitor) suppression, mimicking active and ART-controlled HIV infection in vivo. We demonstrated that HIV-induced, telomeric DDR plays a pivotal role in triggering telomere erosion, premature T-cell aging, and CD4 T-cell apoptosis or depletion via dysregulation of the PI3K/ATM pathways. This in vitro model provides a new tool to investigate HIV pathogenesis, and our results shed new light on the molecular mechanisms of telomeric DDR and CD4 T-cell homeostasis during HIV infection. IMPORTANCE The hallmark of HIV infection is a gradual depletion of CD4 T cells, with a progressive decline of host immunity. How CD4 T cells are depleted in individuals with active and virus-suppressed HIV infection remains unclear. In this study, we employed a cellular model of HIV infection to characterize the mechanisms underlying CD4 T-cell destruction by analyzing the chromosome end (telomere) DNA damage response (DDR) and cellular apoptosis in a T-cell line (highly permissive SupT1 cells), as well as in primary CD4 T cells with active or drug-suppressed HIV infection. We demonstrated that HIV-induced telomeric DDR plays a critical role in inducing telomere loss, premature cell aging, and CD4 T-cell apoptosis or depletion via dysregulation of the PI3K/ATM pathways. This study sheds new light on the molecular mechanisms of telomeric DDR and its role in CD4 T-cell homeostasis during HIV infection.

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