Abstract

Forty years after the identification of the Human Immunodeficiency Virus (HIV) as the causal agent of the AIDS pandemic, HIV infection is still a major global public health issue. The development of antiretroviral treatments (ART) enabled the control of HIV infection and considerably improved the life expectancy of people living with HIV. Nevertheless, HIV infection remains incurable due to the persistence of viral reservoirs mainly composed of resting CD4+ T lymphocytes harboring silent but replication-competent integrated proviruses. With no effective cure targeting latently infected cells, they represent the main barrier for HIV eradication as the virus can be reactivated following treatment escape, leading to a rapid viral rebound. One of the major challenges to develop a cure is to better characterize the molecular mechanisms that control HIV expression. In latently infected cells, HIV silencing is the result of multifactorial transcriptional blocks among which RNAP-II proximal pausing near Transcription Start Site (TSS) at the HIV 5’LTR promoter is a rate-limiting regulatory checkpoint. If paused RNAPII complexes do not resume to productive elongation, Premature Transcription Termination (PTT) can occur and lead to the accumulation of short abortive viral RNAs. Recent studies revealed that PTT is a widespread mechanism that attenuate both protein coding and non-coding transcription. PTT involves several termination pathways and is often associated to the recognition of cryptic-polyadenylation signals (PAS) by the cleavage and polyadenylation complex (CPA). Interestingly, as integrated HIV provirus contains a duplicated LTR region at each end of its genome, the PAS that regulates the 3’ end processing of viral mRNA is also located in the 5’LTR and must be occluded during active transcription. However, the role of the proximal PAS in repressing HIV transcription in latently infected cells remains to be explored. Previous studies have shown that depletion of the CPA complex subunit PCF11 reactivates HIV expression in latently infected cells. Still, little is known about the role of the CPA complex in this process.

The main aim of my thesis was to characterize the contribution of CPA factors to HIV-1 PTT in latently infected cells.

We first evaluated the contribution of the CPA complex in the regulation of LTR-driven basal transcription and identified that only PCF11 inhibits HIV-1 expression. As our results indicated that PCF11 represses HIV-1 for the most part independently of the PAS-motif and the other CPA factors, we further investigated whether PCF11 could associate with other non-canonical termination factors. Interestingly, we found that PCF11 strongly interacts with WDR82, a protein that was recently shown to enforce early termination of non-coding RNAs in human cells. Then, we addressed the role of this newly identified nuclear complex on HIV-1 repression in cellular models of HIV-1 latency and showed that PCF11 and WDR82 are required for the maintenance of HIV-1 post-integration latency. PCF11 and WDR82 are specifically recruited at a promoter-proximal region downstream of the viral promoter in an interdependent manner and act on the same pathway to silence HIV-1 transcription.

These findings highlight the key role of PCF11/WDR82 as a premature transcription termination complex silencing HIV-1 expression in latently infected cells.

Key-words: HIV-1 – Latency – Transcription – Termination – PCF11 – WDR82