Implications of HIV-1 Nef for “Shock and Kill” Strategies to Eliminate Latent Viral Reservoirs

Finding a cure for HIV is challenging because the virus is able to integrate itself into the host cell genome and establish a silent state, called latency, allowing it to evade antiviral drugs and the immune system. Various “shock and kill” strategies are being explored in attempts to eliminate latent HIV reservoirs. The goal of these approaches is to reactivate latent viruses (“shock”), thereby exposing them to clearance by viral cytopathic effects or immune-mediated responses (“kill”). To date, there has been limited clinical success using these methods. In this review, we highlight various functions of the HIV accessory protein Nef and discuss their double-edged effects that may contribute to the limited effectiveness of current “shock and kill” methods to eradicate latent HIV reservoirs in treated individuals

No evidence for selection of HIV-1 with enhanced Gag-Protease or Nef function among breakthrough infections in the CAPRISA 004 tenofovir microbicide trial

BACKGROUND: Use of antiretroviral-based microbicides for HIV-1 prophylaxis could introduce a transmission barrier that inadvertently facilitates the selection of fitter viral variants among incident infections. To investigate this, we assessed the in vitro function of gag-protease and nef sequences from participants who acquired HIV-1 during the CAPRISA 004 1% tenofovir microbicide gel trial. Methods and RESULTS: We isolated the earliest available gag-protease and nef gene sequences from 83 individuals and examined their in vitro function using recombinant viral replication capacity assays and surface protein downregulation assays, respectively. No major phylogenetic clustering and no significant differences in gag-protease or nef function were observed in participants who received tenofovir gel versus placebo gel prophylaxis. CONCLUSION: Results indicate that the partial protective effects of 1% tenofovir gel use in the CAPRISA 004 trial were not offset by selection of transmitted/early HIV-1 variants with enhanced Gag-Protease or Nef fitness

Attenuation Of Multiple Nef Functions In HIV-1 Elite Controllers

Background Impaired HIV-1 Gag, Pol, and Env function has been described in elite controllers (EC) who spontaneously suppress plasma viremia to < 50 RNA copies/mL; however, activity of the accessory protein Nef remains incompletely characterized. We examined the ability of 91 Nef clones, isolated from plasma of 45 EC and 46 chronic progressors (CP), to down-regulate HLA class I and CD4, up-regulate HLA class II invariant chain (CD74), enhance viral infectivity, and stimulate viral replication in PBMC. Results In general, EC Nef clones were functional; however, all five activities were significantly lower in EC compared to CP. Nef clones from HLA-B*57-expressing EC exhibited poorer CD4 down-regulation function compared to those from non-B*57 EC, and the number of EC-specific B*57-associated Nef polymorphisms correlated inversely with 4 of 5 Nef functions in these individuals. Conclusion Results indicate that decreased HIV-1 Nef function, due in part to host immune selection pressures, may be a hallmark of the EC phenotype

Relative Resistance of HLA-B to Downregulation by Naturally Occurring HIV-1 Nef Sequences

ABSTRACT HIV-1 Nef binds to the cytoplasmic region of HLA-A and HLA-B and downregulates these molecules from the surface of virus-infected cells, thus evading immune detection by CD8+ T cells. Polymorphic residues within the HLA cytoplasmic region may affect Nef’s downregulation activity. However, the effects of HLA polymorphisms on recognition by primary Nef isolates remain elusive, as do the specific Nef regions responsible for downregulation of HLA-A versus HLA-B. Here, we examined 46 Nef clones isolated from chronically HIV-1 subtype B-infected subjects for their ability to downregulate various HLA-A, HLA-B, and HLA-C molecules on the surface of virus-infected cells. Overall, HLA-B exhibited greater resistance to Nef-mediated downregulation than HLA-A, regardless of the cell type examined. As expected, no Nef clone downregulated HLA-C. Importantly, the differential abilities of patient-derived Nef clones to downregulate HLA-A and HLA-B correlated inversely with the sensitivities of HIV-infected target cells to recognition by effector cells expressing an HIV-1 Gag-specific T cell receptor. Nef codon function analysis implicated amino acid variation at position 202 (Nef-202) in differentially affecting the ability to downregulate HLA-A and HLA-B, an observation that was subsequently confirmed by experiments using Nef mutants constructed by site-directed mutagenesis. The in silico and mutagenesis analyses further suggested that Nef-202 may interact with the C-terminal Cys-Lys-Val residues of HLA-A, which are absent in HLA-B. Taken together, the results show that natural polymorphisms within Nef modulate its interaction with natural polymorphisms in the HLA cytoplasmic tails, thereby affecting the efficiency of HLA downregulation and consequent recognition by HIV-specific T cells. These results thus extend our understanding of this complex pathway of retroviral immune evasion

Autophagy Induction by HIV-Tat and Methamphetamine in Primary Midbrain Neuronal Cells of Tree Shrews via the mTOR Signaling and ATG5/ATG7 Pathway

Background: Addictive stimulant drugs, such as methamphetamine (METH), increase the risk of exposure to the human immunodeficiency virus-1 (HIV-1) infection and thus predispose individuals to the development of HIV-associated neurocognitive disorders (HANDs). Previous studies have indicated that HIV-Tat (the transactivator of transcription) and METH can synergistically induce autophagy in SH-SY5Y neuroblastoma cells and that autophagy plays a pivotal role in the neuronal dysfunction in HANDs. However, the underlying mechanism of METH-and HIV-Tat-induced neuronal autophagy remains unclear.Methods: We cultured primary midbrain neuronal cells of tree shrews and treated them with METH and HIV-Tat to study the role of METH and HIV-Tat in inducing autophagy. We evaluated the effects of the single or combined treatment of METH and HIV-Tat on the protein expressions of the autophagy-related genes, including Beclin-1 and LC3B, ATG5, and ATG7 in METH and HIV-Tat-induced autophagy. In addition, the presence of autophagosomes in the METH and/or HIV-Tat treatment was revealed using transmission electron microscopy.Results: The results indicated that METH increased the protein levels of LC3B and Beclin-1, and these effects were significantly enhanced by HIV-Tat. Moreover, the results suggested that ATG5 and ATG7 were involved in the METH and HIV-Tat-induced autophagy. In addition, it was found that mTOR inhibition via pharmacological intervention could trigger autophagy and promote METH and HIV-Tat-induced autophagy.Discussion: Overall, this study contributes to the knowledge of the molecular underpinnings of METH and HIV-Tat-induced autophagy in primary midbrain neuronal cells. Our findings may facilitate the development of therapeutic strategies for METH-and HIV-Tat-induced autophagy in HANDs

No evidence for selection of HIV-1 with enhanced gag-protease or nef function among breakthrough infections in the CAPRISA 004 tenofovir microbicide trial.

CAPRISA, 2013.Background: Use of antiretroviral-based microbicides for HIV-1 prophylaxis could introduce a transmission barrier that inadvertently facilitates the selection of fitter viral variants among incident infections. To investigate this, we assessed the in vitro function of gag-protease and nef sequences from participants who acquired HIV-1 during the CAPRISA 004 1% tenofovir microbicide gel trial. Methods and Results: We isolated the earliest available gag-protease and nef gene sequences from 83 individuals and examined their in vitro function using recombinant viral replication capacity assays and surface protein down regulation assays, respectively. No major phylogenetic clustering and no significant differences in gag-protease or nef function were observed in participants who received tenofovir gel versus placebo gel prophylaxis. Conclusion: Results indicate that the partial protective effects of 1% tenofovir gel use in the CAPRISA 004 trial were not offset by selection of transmitted/early HIV-1 variants with enhanced Gag-Protease or Nef fitness

Nef-mediated down-regulation of CD4 and HLA class I in HIV-1 subtype C infection: association with disease progression and influence of immune pressure.

CAPRISA, 2014.Abstract available in pdf