155 : IRF8 and IRF1: Genomic regulators of innate immunity
IRF8 transcription factor is essential for proper macrophage and dendritic cell (DC) differentiation, activation and function. We recently identified IRF8 mutations that are associated with a novel primary myeloid immunodeficiency, coupled with mycobacterial infections susceptibility (MSMD). Interestingly, mice bearing Irf8R294C loss-of-function alleles exhibit a similar phenotype; i.e. development of a chronic myelogenous leukemia-like syndrome characterized by a severe loss of dendritic cells and expansion of immature myeloid precursors, concomitant with susceptibility to intracellular infections. Recent GWAS and eQTL mapping studies associated IRF8 locus with various inflammatory diseases. Our objective is to define transcriptional networks controlled by these important inflammatory checkpoints (IRF8 and its binding partners IRF1 and PU.1), in order to identify novel genes implicated in MSMD and in susceptibility to Mycobacterium tuberculosis infection. To identify these networks, we used a combination high-throughput sequencing techniques. Primary macrophages, prior and after IFNγ activation, were subjected to (1) RNA-seq to identify IFNγ-regulated genes and (2) chromatin immunoprecipitation (ChIP-seq) to localize IRF8, IRF1 and PU.1 genomic binding sites. The comparison of ChIP-seq results for these well-known binding partners confirmed their co-recruitment to regulatory elements. Interestingly however, they are not obligate partners as we uncovered many different binding schemes. Furthermore, IFNγ triggers de novo IRF1 DNA binding, but had limited influence on IRF8 and PU.1 that are already occupying their binding sites. The overlap of ChIP-seq and RNA-seq data revealed that IFNγ-upregulated genes are strongly associated with IRF binding sites, as opposed to repressed genes. ChIP-qPCR and RNA-seq in IRF8R294C and IRF1−/− primary macrophages confirmed the interdependence of IRF8 and IRF1 co-recruitment and their respective importance for IFNγ-regulated genes subsets. Genes identified in these transcriptional pathways represent the “core myeloid transcriptome” and are likely to play critical roles in macrophage function and may represent valuable candidates for myeloid immunodeficiencies and susceptibility to mycobacterial infections.