Post-transcriptional regulation of HIV-1 gene expression by the host factor MATR3

The post-transcriptional regulation of HIV-1 replication is finely controlled by both viral and host factors. Among the former, Rev is involved in the nuclear ex-port of viral intron-containing mRNAs, a group of transcripts which encodes for viral enzymes and structural proteins, but also constitute the viral genome that will be encapsidated into nascent virions. To avoid the nuclear retention of these intron-containing transcripts, Rev en-gages an alternative export route which ultimately involves the CRM-1 export factor; during this process Rev requires the concerted action of several host fac-tors but the details of its interactions are still not fully understood. To dissect this pathway a novel proteomic approach for the immunoprecipitation of the viral RNA was developed in our laboratory : we thus identified the nuclear matrix protein MATR3 as a Rev co-factor which was recruited after mRNA biogenesis during this process (Kula et al., 2011, 2013). We could assess that MATR3 acts in the post-transcriptional steps of viral replication and we could demonstrate its role as a Rev-cofactor during the nuclear export of viral mRNAs. To establish the functional role of MATR3 during acute viral infection we modu-late its levels in Jurkat cells by both knockdown or overexpression. We found that, while MATR3 depletion resulted in the drastic reduction of viral replication, its overexpression leads to enhanced viral particle production. We applied the same approach to primary PBLs and obtained a similar result concluding that MATR3 is a positive regulator of viral replication. To investigate a possible role for MATR3 in the establishment of viral latency we depleted MATR3 from J-Lat cells, a well-established model of latency. We ob-served that MATR3 depletion did not impede transcriptional reactivation of the integrated provirus upon TNF\u3b1 stimulation, but strongly impaired intracellular viral protein production and full viral rescue. This observation demonstrated that MATR3 depletion affects the post-transcriptional steps of latency reversal suggesting that this factor could play a crucial role during the maintenance of latency. Since most of the attempts done with Latency Reversal Agents (LRAs), an heterogeneous group of drugs proposed to restore viral transcription, failed to induce full reactivation of the latent provirus (Darcis et al., 2015; Spina et al., 2013) we reasoned that there could be a post-transcriptional block to full viral reactivation in latently infected cells and we suppose that MATR3 could represent a limiting factor to this process. We confirmed that MATR3 was almost undetectable in resting PBLs but could be promptly upregulated upon cellular activation. MATR3 was not induced by treatment with LRAs, such as SAHA or Romidepsin within PBL from healthy donors and HIV-infected patients. We propose that the restoration of proper MATR3 levels within latently infected cells could enhance latency reversal in LRAs-treated cells

Posttranscriptional Regulation of HIV-1 Gene Expression during Replication and Reactivation from Latency by Nuclear Matrix Protein MATR3.

Posttranscriptional regulation of HIV-1 replication is finely controlled by viral and host factors. Among the former, Rev controls the export of partially spliced and unspliced viral RNAs from the nucleus and their translation in the cytoplasm or incorporation into new virions as genomic viral RNA. To investigate the functional role of the Rev cofactor MATR3 in the context of HIV infection, we modulated its expression in Jurkat cells and primary peripheral blood lymphocytes (PBLs). We confirmed that MATR3 is a positive regulator of HIV-1 acting at a posttranscriptional level. By applying the same approach to J-lat cells, a well-established model for the study of HIV-1 latency, we observed that MATR3 depletion did not affect transcriptional reactivation of the integrated provirus, but caused a reduction of Gag production. Following these observations, we hypothesized that MATR3 could be involved in the establishment of HIV-1 posttranscriptional latency. Indeed, mechanisms acting at the posttranscriptional level have been greatly overlooked in favor of transcriptional pathways. MATR3 was almost undetectable in resting PBLs, but could be promptly upregulated upon cellular stimulation with PHA. However, HIV latency-reversing agents were poor inducers of MATR3 levels, providing a rationale for their inability to fully reactivate the virus. These data have been confirmed ex vivo in cells derived from patients under suppressive ART. Finally, in the context of MATR3-depleted J-lat cells, impaired reactivation by SAHA could be fully rescued by MATR3 reconstitution, demonstrating a direct role of MATR3 in the posttranscriptional regulation of HIV-1 latency.IMPORTANCE The life cycle of HIV-1 requires integration of a DNA copy into the genome of the host cell. Transcription of the viral genes generates RNAs that are exported to the cytoplasm with the contribution of viral and cellular factors to get translated or incorporated in the newly synthesized virions. It has been observed that highly effective antiretroviral therapy, which is able to reduce circulating virus to undetectable levels, cannot fully eradicate the virus from cellular reservoirs that harbor a transcriptionally latent provirus. Thus, persistence of latently infected cells is the major barrier to a cure for HIV-1 infection. In order to purge these reservoirs of latently infected cells, it has been proposed to activate transcription to stimulate the virus to complete its life cycle. This strategy is believed to unmask these reservoirs, making them vulnerable to the immune system. However, limited successes of this approach may indicate additional posttranscriptional restrictions that need to be overcome for full virus reactivation. In this work we identify the cellular protein MATR3 as an essential cofactor of viral RNA processing. Reactivation of HIV-1 transcription per se is not sufficient to allow completion of a full life cycle of the virus if MATR3 is depleted. Furthermore, MATR3 is poorly expressed in quiescent CD4+ T lymphocytes that are the major reservoir of latent HIV-1. Cells derived from aviremic HIV-1 patients under antiretroviral therapy didn't express MATR3, and most importantly, latency-reversing agents proposed for the rescue of latent provirus were ineffective for MATR3 upregulation. To conclude, our work identifies a cellular factor required for full HIV-1 reactivation and points to the revision of the current strategies for purging viral reservoirs that focus only on transcription.SCOPUS: ar.jinfo:eu-repo/semantics/publishe