Concerted recruitment of the mediator complex explains STAT-NFkB interaction in the antimicrobial response of macrophages

Macrophages respond to microbial invaders such as the Gram-positive intracellular bacterium Listeria monocytogenes (Lm) with a complex signaling network and the establishment of specialized gene expression profiles. The Nos2 gene encoding inducible nitric oxide synthase belongs to a group of antimicrobial genes whose transcription is regulated by functional cooperation between a type I IFN receptor->ISGF3 (STAT1/STAT2/IRF9) pathway and the NFκB pathway. We have recently shown that NFκB stimulates recruitment of the RNA polymerase II (Pol II) kinases CDK7/TFIIH and CDK9/pTEFb to the Nos2 promoter whereas ISGF3 is required for the binding of the Pol II enzyme. Recruitment of CDK7, but not of CDK9 was BET-protein (Brd4) dependent. Here we investigate whether the kinase module of the mediator functions as an alternative platform for pTEFb recruitment. Mediator is a multi-subunit protein complex bridging transcription factors with Pol II, initiation and elongation factors. We used ChIP-Seq analyses to explore the group of genes that are, like Nos2, subject to dual-regulation by both NFκB and ISGF3 in Lm-infected cells. In this context, we examined the role of STAT and NFκB pathways in the modification of promoter chromatin and the modification of Pol II. We report an essential role for different portions of the mediator complex for NFκB-dependent deposition of pTEFb and for ISGF3-dependent recruitment of Pol II.