Biswanath Majumder

Human Immunodeficiency Virus Type 1 Vpr Impairs Dendritic Cell Maturation and T-Cell Activation: Implications for Viral Immune Escape

ABSTRACT Antigen presentation and T-cell activation are dynamic processes involving signaling molecules present in both APCs and T cells. Effective APC function and T-cell activation can be compromised by viral immune evasion strategies, including those of human immunodeficiency virus type 1 (HIV-1). In this study, we determined the effects of HIV-1 Vpr on one of the initial target of the virus, dendritic cells (DC), by investigating DC maturation, cytokine profiling, and CD8-specific T-cell stimulation function followed by a second signal. Vpr impaired the expression of CD80, CD83, and CD86 at the transcriptional level without altering normal cellular transcription. Cytokine profiling indicated that the presence of Vpr inhibited production of interleukin 12 (IL-12) and upregulated IL-10, whereas IL-6 and IL-1β were unaltered. Furthermore, DC infected with HIV-1 vpr+ significantly reduced the activation of antigen-specific memory and recall cytotoxic-T-lymphocyte responses. Taken together, these results indicate that HIV-1 Vpr may in part be responsible for HIV-1 immune evasion by inhibiting the maturation of costimulatory molecules and cytokines essential for immune activation.

HIV-1 Mediated Immune Pathogenesis: Spotlight on the Role of Viral Protein R (VPR)

HIV-1 exploits the cellular machinery to replicate in the host cells by targeting a wide range of host factors at different stages of its life cycle. HIV-1 also induces detrimental effects in the infected and uninfected bystander cells resulting in dysregulation including interference in immune effector functions. The latter is specifically linked to the immune evasion strategies of the virus. In addition to the essential roles of structural proteins (Gag, Pol & Env), HIV-1 encoded auxiliary proteins such as Nef, Vif, Vpu, and Vpr through their interaction with the host cellular partners facilitate viral replication and dissemination. HIV-1 Vpr, a virion-associated molecule, has been implicated to play a role in the early events in virus life cycle. Vpr is a pleiotropic protein that exerts a range of effects including inhibition of cell proliferation, induction of apoptosis and modulation of a number of immune molecules. These functions could be in part responsible for Vpr induced immune evasion and virus replication. Appreciating this view is the genetic variation in vpr gene reflected in the form of polymorphisms at the amino acid level that may contribute to the potential CTL escape of the virus. It is likely that Vpr mediated dysregulation of host immune response contributes, in part, to the progression of disease. This review focuses on the recent advances regarding HIV-1 Vpr mediated immunopathogenesis and the mechanistic insight from in vitro and in vivo studies.

Dendritic Cells Infected with vpr-Positive Human Immunodeficiency Virus Type 1 Induce CD8+ T-Cell Apoptosis via Upregulation of Tumor Necrosis Factor Alpha

ABSTRACT Human immunodeficiency virus type 1 (HIV-1) viral protein R (Vpr) plays a crucial role in viral replication and pathogenesis by inducing cell cycle arrest, apoptosis, translocation of preintegration complex, potentiation of glucocorticoid action, impairment of dendritic cell (DC) maturation, and T-cell activation. Recent studies involving the direct effects of Vpr on DCs and T cells indicated that HIV-1 containing Vpr selectively impairs phenotypic maturation, cytokine network, and antigen presentation in DCs and dysregulates costimulatory molecules and cytokine production in T cells. Here, we have further investigated the indirect effect of HIV-1 Vpr+ virus-infected DCs on the bystander CD8+ T-cell population. Our results indicate that HIV-1 Vpr+ virus-infected DCs dysregulate CD8+ T-cell proliferation and induce apoptosis. Vpr-containing virus-infected DC-mediated CD8+ T-cell killing occurred in part through enhanced tumor necrosis factor alpha production by infected DCs and subsequent induction of death receptor signaling and activation of the caspase 8-dependent pathway in CD8+ T cells. Collectively, these results provide evidence that Vpr could be one of the important contributors to the host immune escape by HIV-1 through its ability to dysregulate both directly and indirectly the DC biology and T-cell functions.

Infection with Vpr-Positive Human Immunodeficiency Virus Type 1 Impairs NK Cell Function Indirectly through Cytokine Dysregulation of Infected Target Cells

ABSTRACT Human immunodeficiency virus type 1 (HIV-1) infection has been implicated in impairing various aspects of NK cell function in viremic condition, and several viral factors contribute to these defects. Here, we evaluated the effect of HIV-1 Vpr on NK cell cytolytic function and cytokine (gamma interferon [IFN-γ]) production in the context of infection and exposure. Our data indicate that NK cells derived from a peripheral blood mononuclear cell culture infected in vitro with HIV-1 vpr(+) virus or exposed to recombinant Vpr protein exhibited reduced target cell killing in conjunction with diminished expression of CD107a and reduced IFN-γ production compared to their Vpr-negative counterparts. This Vpr-induced NK cell defect is in part through differential regulation of interleukin-12 and transforming growth factor β production by the infected target cells and concomitant activation of Smad3 signaling pathway. Collectively, these results illustrate the ability of Vpr to impair NK cell-mediated innate immune functions indirectly by dysregulating multiple cytokines in the infected target cells, thus increasing disease severity and affecting the final outcome in HIV-1 infection.

Availability of a diversely Avid CD8+ T cell repertoire specific for the subdominant HLA-A2-restricted HIV-1 gag p2419-27 epitope

HLA-A2-restricted CTL responses to immunodominant HIV-1 epitopes do not appear to be very effective in the control of viral replication in vivo. In this study, we studied human CD8+ T cell responses to the subdominant HLA-A2-restricted epitope TV9 (Gag p2419–27, TLNAWVKVV) to explore the possibility of increasing its immune recognition. We confirmed in a cohort of 313 patients, infected by clade B or clade C viruses, that TV9 is rarely recognized. Of interest, the functional sensitivity of the TV9 response can be relatively high. The potential T cell repertoires for TV9 and the characteristics of constituent clonotypes were assessed by ex vivo priming of circulating CD8+ T cells from healthy seronegative donors. TV9-specific CTLs capable of suppressing viral replication in vitro were readily generated, suggesting that the cognate T cell repertoire is not limiting. However, these cultures contained multiple discrete populations with a range of binding avidities for the TV9 tetramer and correspondingly distinct functional dependencies on the CD8 coreceptor. The lack of dominant clonotypes was not affected by the stage of maturation of the priming dendritic cells. Cultures primed by dendritic cells transduced to present endogenous TV9 were also incapable of clonal maturation. Thus, a diffuse TCR repertoire appeared to be an intrinsic characteristic of TV9-specific responses. These data indicate that subdominance is not a function of poor immunogenicity, cognate TCR repertoire availability, or the potential avidity properties thereof, but rather suggest that useful responses to this epitope are suppressed by competing CD8+ T cell populations during HIV-1 infection.

Molecular and functional characterization of a novel splice variant of ANKHD1 that lacks the KH domain and its role in cell survival and apoptosisc

Multiple ankyrin repeat motif‐containing proteins play an important role in protein–protein interactions. ANKHD1 proteins are known to possess multiple ankyrin repeat domains and a single KH domain with no known function. Using yeast two‐hybrid system analysis, we identified a novel splice variant of ANKHD1. This splice variant of ANKHD1, which we designated as HIV‐1 Vpr‐binding ankyrin repeat protein (VBARP), does not contain the signature KH domain, and codes for only a single ankyrin repeat motif. We characterized VBARP by molecular and functional analysis, revealing that VBARP is ubiquitously expressed in different tissues as well as cell lines of different lineage. In addition, blast searches indicated that orthologs and homologs to VBARP exist in different phyla, suggesting that VBARP might be evolutionarily conserved, and thus may be involved in basic cellular function(s). Furthermore, biochemical analysis revealed the presence of two VBARP isoforms coding for 69 and 49 kDa polypeptides, respectively, that are primarily localized in the cytoplasm. Functional analysis using short interfering RNA approaches indicate that this gene product is essential for cell survival through its regulation of caspases. Taken together, these results indicate that VBARP is a novel splice variant of ANKHD1 and may play a role in cellular apoptosis (antiapoptotic) and cell survival pathway(s).

Attenuated nef DNA vaccine construct induces cellular immune response: role in HIV-1 multiprotein vaccine.

HIV-1 positive patients generate Nef-specific CTL response, indicating that Nef is a potent immunogen. However, Nef is also known to down regulate the expression of CD4 and MHC-I molecules, thereby protecting virally infected target cells. We compared the immunogenicity of non-functional nef vaccine constructs to wild type functional nef as potential immunogen. Mice were immunized with different nef constructs and assessed for their ability to induce cellular immune responses. Evaluation of T cell immune responses in mice showed that non-functional nef vaccine constructs are capable of inducing a significant T cell immune response measured by IFN-gamma ELISPOT. Further epitope mapping studies indicate that one of our attenuated constructs, Nef R-38, has multiple CTL epitopes spanning throughout the gene. Our results indicate that functionally attenuated Nef antigen might be a better candidate for future multiprotein HIV-1 vaccine.