Dubrovsky, L. | bioRxiv Science

bioRxiv2022

Extracellular vesicles carrying HIV-1 Nef induce trained immunity in myeloid cells

SummaryPersistent inflammation is a hallmark of HIV infection and is not reversed after suppression of viral replication by anti-retroviral therapy (ART). One explanation for chronic inflammation in ART-treated HIV-infected individuals is hyperreactivity of the myeloid cells due to a phenomenon called trained immunity. Here, we demonstrate that human monocyte derived macrophages originating from monocytes initially treated with extracellular vesicles containing HIV-1 protein Nef (exNef), but differentiating in the absence of exNef, released increased levels of pro-inflammatory cytokines after lipopolysaccharide stimulation. This effect was associated with epigenetic changes related to inflammation and cholesterol metabolism pathways, upregulation of the lipid rafts, and was blocked by methyl-{beta}-cyclodextrin, statin, and inhibitor of activity of lipid raft-associated receptor IGF1R. Bone marrow-derived macrophages from exNef-injected mice had higher abundance of lipid rafts and produced elevated levels of TNF. These phenomena are consistent with trained immunity and may contribute to persistent inflammation and co-morbidities in HIV-infected individuals.

immunology↗

bioRxiv2017

DNA-PK facilitates HIV transcription by regulating the activity of RNA polymerase II and the recruitment of transcription machinery at HIV LTR

Despite the use of highly effective antiretroviral therapy (HAART), the presence of latent or transcriptionally silent proviruses prevents cure and eradication of HIV infection. These transcriptionally silent proviruses are well protected from both the immune system and HAART regimens. Thus, in order to tackle the problem of latent HIV reservoirs, it is a prerequisite to define all the pathways that regulate HIV transcription. We have previously reported that DNA-PK facilitates HIV transcription by interacting with the RNA polymerase II (RNAP II) complex recruited at HIV LTR. To extend those studies further, here we demonstrate that DNA-PK promotes HIV transcription by supporting it at several stages, including initiation, pause-release and elongation. We discovered that DNA-PK increases phosphorylation of RNAP II C-terminal domain (CTD) at serine 5 (Ser5) and serine 2 (Ser2) by both directly catalyzing and by augmenting the recruitment of P-TEFb at HIV LTR. We found that DNA-PK facilitates the establishment of euchromatin structure at HIV LTR, which further supports HIV gene expression. DNA-PK inhibition or knockdown leads to the severe impairment of HIV gene expression and conversion of euchromatin to heterochromatin at HIV LTR. It also profoundly restricts HIV replication and reactivation of latent provirus. DNA-PK promotes the recruitment of TRIM28 at LTR and facilitates the release of paused RNAP II through TRIM28 phosphorylation. The results were reproduced in cell lines belonging to both lymphoid and myeloid lineages and were confirmed in primary CD4+ T cells and peripheral blood mononuclear cells (PBMCs) from HIV-infected patients.\n\nIMPORTANCEOur results reveal the important role of DNA-PK in supporting HIV transcription, replication and latent proviral reactivation. Intriguingly, this study sheds light on an important pathway that affects HIV gene expression. These findings provide strong rationale for developing and using transcriptional inhibitors, such as DNA-PK inhibitors, as supplement to HAART regimens in order to further enhance their effectiveness and to suppress toxicity due to HIV proteins.

molecular biology↗