Mécanismes moléculaires de la réplication préférentielle du VIH-1 dans les cellules à polarisation Th1Th17 versus Th1 : rôle de PPARG dans la régulation négative de la réplication virale

Human CD4+ T cells are heterogeneous in terms of permissiveness to infection by the human immunodeficiency virus type 1 (HIV-1). Our laboratory previously demonstrated that Th1 cells (CXCR3+CCR6- phenotype) are relatively resistant to infection, whereas Th1Th17 cells (CXCR3+CCR6+ phenotype) are highly permissive to HIV-1. HIV replication depends on several cellular restriction or permissiveness factors acting at different stages of the viral life cycle. However, despite several advances, our knowledge on signaling pathways involved in HIV replication is still limited. The main objective of this MSc degree project is to characterize the molecular mechanisms of permissiveness and resistance to HIV in Th1Th17 and Th1 cells respectively. This thesis is divided into four parts, aiming at : (i) the identification of canonical pathways and biological functions differentially regulated in Th1Th17 vs Th1 cells through the analysis of their whole genome transcriptome; (ii) the validation of differential expression of relevant genes identified by microarrays at mRNA and protein levels; (iii) the characterization of the functional role of some of these factors (i.e. PPARG, AhR) on HIV replication in Th1Th17 versus Th1 cells; and (iv) the identification of the level at which these factors interfere with the HIV replication cycle. Our analysis of the large sets of microarray data by Gene Set Enrichment Analysis and Ingenuity Pathway Analysis indicate that Th1Th17 compared to Th1 cells are more susceptible to cell activation and apoptosis, promote superior inflammation and express at low levels genes related to the proteosomal degradation. These differences in the regulation of various biological functions and pathways can partly explain the susceptibility to HIV infection in these cells. We then confirmed the differential expression of some genes of interest in Th1Th17 (CXCR6, PPARG, ARNTL, CTSH, PTPN13, MAP3K4) versus Th1 (SERPINB6, PTK2) cells at mRNA and protein levels. Finally, we demonstrated the role of the transcription factors PPARG and AhR in the regulation of HIV replication. The activation of PPARG by rosiglitazone induces an important decrease in HIV replication in CD4+ T cells, while AhR activation by its exogenous ligands TCDD and FICZ promotes viral replication. We propose that the PPARG pathway acts as a negative regulator of HIV replication in these cells by interfering with Th17 polarization and probably by inhibiting the transcriptional activity of NF-kB. The role of nuclear versus cytoplasmic AhR appears diametrically opposed, since RNA interference against AhR is also associated with a significant increase in HIV replication. It is thus possible that the cytoplasmic form of AhR, known for its E3 ubiquitine ligase activity, is involved in proteasomal degradation of the viral particles. The mechanism by which the nuclear versus cytoplasmic form of AhR interferes with viral replication is being studied in the laboratory. This study represents the first characterization of the differential expression of genes in the entire genome of CD4+ T subpopulations permissive (Th1Th17) versus resistant (Th1) to infection by HIV. Ours results identify new molecular targets for therapeutic strategies to limit HIV replication in primary CD4+ T lymphocytes.

Keyphrases: Th1Th17, Th1, PPARG, AhR, HIV-1, CD4+ T cells, cDNAmicroarrays