Characterization of HIV-1 vpu Gene from Suppressed Viremic Older Individuals with HIV on Long-Term Antiretroviral Therapy.

INTRODUCTION/OBJECTIVE: Although advancements in antiretroviral therapy (ART) have been able to control HIV infection in people with HIV (HIV+) to achieve suppressed viral loads and recovered CD4 T cell counts, the virus persists in latent reservoirs. Characterizing the molecular, biological, and immunological features of HIV-1 genes from these HIV+ virologically controlled individuals is essential for the development of eradication strategies. The HIV-1 accessory gene, vpu, plays important roles in the release of progeny virions and modulation of innate immune signaling, contributing to viral pathogenesis and persistence in the host. In this study, we analyzed the vpu sequences from 21 predominantly older HIV+ individuals with long-term ART-mediated viral suppression (mostly undetectable viral load) and mostly restored CD4 T cell counts.

METHODS: Genomic DNA was isolated from peripheral blood mononuclear cells (PBMC) extracted from blood samples from 21 HIV+ individuals. Amplification of the vpu gene was performed by polymerase chain reaction (PCR), followed by cloning utilizing a TOPO vector. Bioinformatics analysis of the vpu nucleotide sequences was conducted.

RESULTS: Phylogenetic analysis of 288 vpu sequences revealed that the vpu sequences from each of the 21 HIV+ individuals were distinctly separated from one another and formed individual clusters within their respective subtrees. These sequences showed a low degree of genetic variability, reduced genetic diversity estimates, and most patient sequences did not display evidence of positive selective pressure. In 82% of the deduced Vpu amino acid sequences, the open reading frames were intact, with most sequences demonstrating conservation of functional domains necessary for Vpu activity involving virion release, BST-2 degradation, oligomerization, and CD4 degradation. Deduced amino acid sequences exhibited increased variability in the previously identified CTL epitope within Vpu, suggesting the presence of escape mutants.

DISCUSSION: Our findings demonstrate that the coding sequences and functional domains of the HIV-1 vpu gene from HIV+ PMBC DNA on long-term ART and improved CD4 counts were highly conserved with low genetic diversity and increased variability in the CTL epitope, sug-gesting a role for Vpu in viral persistence.

CONCLUSION: The vpu sequences from 21 older HIV+ individuals on long-term ART and suppressed viremia exhibited a low degree of genetic diversity, conserved functional domains, and variability within the CTL epitope, suggesting a role in viral persistence in these HIV+ individuals. These findings may help develop new treatments and curative strategies.