B-104 Presence of HIV-1 RNA in extracellular vesicles from HIV-1 cART-treated cells

Abstract

HIV-1 is a complex retrovirus that produces chronic infection affecting approximately 40 million people worldwide and 1.1 million new infections each year. We have previously shown that during infection, EVs contain a non-coding HIV-1 RNA, which has been shown to elicit responses in recipient cells (1–3). First, we investigated the effect of exosomes derived from uninfected cells on latent HIV-1–infected cells and found that there is an abundant level of short noncoding RNA in infected cells, arguing that a true “transcriptional latency” may not exist in vivo since cells are constantly in contact with exosomes from uninfected cells. A possible mechanism for this finding revealed that the exosomes cause an increase in RNA Polymerase II loading onto the DNA within the infected cells; this was possibly driven by the presence of specific kinases in the exosomes, some of which were unique to T-cell-derived and myeloid-derived exosomes. Subsequently transcribed viral transcripts, which include TAR and a novel RNA termed “TAR-gag”, can then be packaged into exosomes and potentially be exported to neighboring uninfected cells. TAR-gag is a long, noncoding RNA that is 615 bases long, contains the TAR element, and terminates within the p17 region of HIV-1 gag (1). In the presence of transcription inhibitors F07#13 (a Tat peptide mimetic) and CR8#13 (an ATP-binding analog of Cdk9), TAR-gag is capable of binding to SWI/SNF components in the nucleus, including the mSin3A/HDAC-1 complex, and serves as a scaffolding RNA. Additionally, TAR-gag can recruit suppressive factors and RNA-binding proteins to the HIV-1 promoter, resulting in transcriptional gene silencing (TGS) (2). In our most recent data, we found altered levels of genomic RNA within EVs while other RNAs, such as TAR RNA, remained unaffected by the addition of cART treatment in both cell lines, primary macrophages, and patient biofluids. Furthermore, we determined possible mechanisms involved in the selective packaging of HIV-1 RNA products into EVs specifically, an increase in EV-associated RNA-binding protein hnRNPA2/B1, and a “reader” for methylated RNA.