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Chromosomal integration of genome-intact HIV-1 sequences into the host genome creates a reservoir of virally infected cells that persists throughout life, necessitating indefinite antiretroviral suppression therapy. During effective antiviral treatment, the majority of these proviruses remain transcriptionally silent, but mechanisms responsible for viral latency are insufficiently clear. Here, we used matched integration site and proviral sequencing (MIP-Seq), an experimental approach involving multiple displacement amplification of individual proviral species, followed by near- full-length HIV-1 next-generation sequencing and corresponding chromosomal integration site analysis to selectively map the chromosomal positions of intact and defective proviruses in 3 HIV-1-infected individuals undergoing long-term antiretroviral therapy. Simultaneously, chromatin accessibility and gene expression in autologous CD4* T cells were analyzed by assays for transposase-accessible chromatin using sequencing (ATAC-Seq) and RNA-Seq. We observed that in comparison to proviruses with defective sequences, intact HIV-1 proviruses were enriched for non-genic chromosomal positions and more frequently showed an opposite orientation relative to host genes. In addition, intact HIV-1 proviruses were preferentially integrated in either relative proximity to or increased distance from active transcriptional start sites and to accessible chromatin regions. These studies strongly suggest selection of intact proviruses with features of deeper viral latency during prolonged antiretroviral therapy, and may be informative for targeting the genome-intact viral reservoir., Introduction HIV-1 integration into host chromosomes, catalyzed by the viral enzyme integrase, allows the virus to persist in the human organism for the life span of the infected cell and [...]

HIV-1 causes a chronic, incurable disease due to its persistence in [CD4.sup.+]T cells that contain replication -competent provirus, but exhibit little or no active viral gene expression and effectively resist combination antiretroviral therapy (cART). These latently infected T cells represent an extremely small proportion of all circulating [CD.sup.+]T cells but possess a remarkable long-term stability and typically persist throughout life, for reasons that are not fully understood. Here we performed massive single-genome, near-full-length next-generation sequencing of HIV-1 DNA derived from unfractionated peripheral blood mononuclear cells, exvivo-isolated [CD4.sup.+]T cells, and subsets of functionally polarized memory [CD4.sup.+]T cells. This approach identified multiple sets of independent, near-full-length proviral sequences from cART-treated individuals that were completely identical, consistent with clonal expansion of [CD4.sup.+]T cells harboring intact HIV-1. Intact, near-full-genome HIV-1 DNA sequences that were derived from such donally expanded [CD4.sup.+] T cells constituted 62% of all analyzed genomeintact sequences in memory [CD4.sup.+] T cells, were preferentially observed in Th1-polarized cells, were longitudinally detected overa duration of up to 5 years, and were fully replication- and infection-competent. Together, these data suggest that clonal proliferation of Th1-polarized [CD4.sup.+]T cells encoding for intact HIV-1 represents a driving force for stabilizing the pool of latently infected [CD4.sup.+]T cells., Introduction A distinct characteristic of HIV-1 infection is the establishment of a long-lived reservoir of virally infected [CD4.sup.+] T cells that encode for, but do not actively express, replication-competent HIV-1 [...]

Elite controllers represent a unique group of HIV-1-infected persons with undetectable HIV-1 replication in the absence of antiretroviral therapy. However, the mechanisms contributing to effective viral immune defense in these patients remain unclear. Here, we show that compared with HIV-1 progressors and HIV-1-negative persons, [CD4.sup.+] T cells from elite controllers are less susceptible to HIV-1 infection. This partial resistance to HIV-1 infection involved less effective reverse transcription and mRNA transcription from proviral DNA and was associated with strong and selective upregulation of the cyclin-dependent kinase inhibitor p21 (also known as cip-1 and waf-1). Experimental blockade of p21 in [CD4.sup.+] T cells from elite controllers resulted in a marked increase of viral reverse transcripts and mRNA production and led to higher enzymatic activities of cyclin-dependent kinase 9 (CDK9), which serves as a transcriptional coactivator of HIV-1 gene expression. This suggests that p21 acts as a barrier against HIV-1 infection in [CD4.sup.+] T cells from elite controllers by inhibiting a cyclin-dependent kinase required for effective HIV-1 replication. These data demonstrate a mechanism of host resistance to HIV-1 in elite controllers and may open novel perspectives for clinical strategies to prevent or treat HIV-1 infection., Introduction HIV-1 infection leads to progressively rising viremia, loss of [CD4.sup.+] T cell counts, and clinical symptoms of immunodeficiency in the vast majority of untreated individuals; however, a small proportion [...]

Elite controllers represent a unique group of HIV-1-infected persons with undetectable HIV-1 replication in the absence of antiretroviral therapy. However, the mechanisms contributing to effective viral immune defense in these patients remain unclear. Here, we show that compared with HIV-1 progressors and HIV-1-negative persons, CD4+ T cells from elite controllers are less susceptible to HIV-1 infection. This partial resistance to HIV-1 infection involved less effective reverse transcription and mRNA transcription from proviral DNA and was associated with strong and selective upregulation of the cyclin-dependent kinase inhibitor p21 (also known as cip-1 and waf-1). Experimental blockade of p21 in CD4+ T cells from elite controllers resulted in a marked increase of viral reverse transcripts and mRNA production and led to higher enzymatic activities of cyclin-dependent kinase 9 (CDK9), which serves as a transcriptional coactivator of HIV- 1 gene expression. This suggests that p21 acts as a barrier against HIV-1 infection in CD4+ T cells from elite controllers by inhibiting a cycin-dependent kinase required for effective HIV-1 replication. These data demonstrate a mechanism of host resistance to HIV-1 in elite controllers and may open novel perspectives for clinical strategies to prevent or treat H1V-1 infection. [ABSTRACT FROM AUTHOR]