Nuclear RNA surveillance complexes silence HIV-1 transcription

Abstract

Expression from the HIV-1 LTR can be repressed in a small population of cells, which contributes to the latent reservoir. The factors mediating this repression have not been clearly elucidated. We have identified a network of nuclear RNA surveillance factors that act as effectors of HIV-1 silencing. RRP6, MTR4, ZCCHC8 and ZFC3H1 physically associate with the HIV-1 TAR region and repress transcriptional output and recruitment of RNAPII to the LTR. Knock-down of these factors in J-Lat cells increased the number of GFP-positive cells, with a concomitant increase in histone marks associated with transcriptional activation. Loss of these factors increased HIV-1 expression from infected PBMCs and led to reactivation of HIV-1 from latently infected PBMCs. These findings identify a network of novel transcriptional repressors that control HIV-1 expression and which could open new avenues for therapeutic intervention.

Chromatin-associated MRN complex protects highly transcribing genes from genomic instability

“…1A). To validate the interaction of DDR factors with actively transcribed HIV-1, we performed chromatin immunoprecipitation (ChIP) in cells containing an integrated HIV-1 LTR linked to a luciferase reporter, as described previously (12). Tat These data indicated that DDR factors such as MDC1 and MRN may be closely linked with active transcription.…”

“…Tandem affinity purification of Flag-HA MDC1 was performed, as described previously (12)(13)(14) ( Fig. 1B).…”

“…Production of shRNA-expressing lentiviral particles was performed as described previously (12) using plasmids expressing shRNAs targeting MRE11 (Sigma mission shRNA TRCN000338391), NBS1 (Sigma mission shRNA TRCN0000288622) or a non-targeting control (obtained through (Addgene, plasmid 1864), as shown in Table S3. For knockdown experiments, HeLa cells were transduced with lentiviral particles and harvested 4 days later, as described previously (12).…”

“…Co-immunoprecipitation was performed as described previously (12) using antibodies shown in Table S4.…”

Des complexes protéiques impliqués dans la surveillance de l’ARN nucléaire inhibent la transcription du VIH-1

“…Cette inhibition résultait de la dégradation de l'ARN viral naissant par RRP6. Plus récemment, dans un manuscrit paru dans PLOS Pathogens [7] tant de souligner que seule la déplétion concomitante des deux protéines a permis d'activer la transcription du promoteur VIH-1 à un niveau comparable à celui des cellules n'exprimant plus RRP6. En revanche, l'extinction unique de ZCCHC8 ou de ZFC3H1 n'a pas eu d'effet majeur.…”