Epigenetic Control of Endothelial and T Cell States By BET Reader Proteins

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DESCRIPTION (provided by applicant): Endothelial cells (EC) must rapidly transition between distinct functional cell states - under basal conditions, where they help maintain homeostasis, and in response to inflammation, where they help coordinate systemic responses, like T cell recruitment. Such cell states are defined to a significant extent by the modulated, integrated expression of large, multiple gene transcriptional programs. The Bromodomain and Extra-Terminal (BET) family of bromodomain-containing epigenetic reader proteins (BRD2, BRD3, BRD4, collectively referred to here as BETs) orchestrate gene expression in response to specific stimuli by binding to specific acetylated lysines on N-terminal histone tails, fostering assembly of transcriptional machinery. Despite burgeoning interest in BETs and ongoing clinical BET inhibitor trials in humans, the role of BETs in ECs, T cells and EC-T cell interactions remains unexplored in terms of the endothelium and poorly understood in terms of T cell differentiation and function. We provide evidence here that BRD4 transduces the TNFα signal to endothelial chromatin, inducing coordinated genomic BRD4 redistribution to de novo super-enhancers that help drive transcription of the pro-inflammatory NFκB endothelial program. Moreover, these newly formed inflammatory regulatory regions accumulate at the expense of immediately decommissioned BRD4 super-enhancers previously active in quiescent ECs. Consistent with these findings, BET inhibition limits leukocyte responses to TNFα-activated ECs in vitro, ex vivo, and in vivo. This data frames our central hypothesis under study here: BET action and redeployment dynamically governs global transcriptional programs in ECs and T cells at rest and after inflammatory cytokine stimulation, thus controlling cell states and functional responses in inflammation and angiogenesis. Aim 1 will test the hypothesis that the demonstrated Brd4-controlled endothelial pro- inflammatory program varies as a function of the kinetics of BRD4-mediated responses, distinct EC types, induction of previously unrecognized TNFα/BRD4-controlled EC target genes, and the proximal cytokine stimulus. Aim 2 will test the hypothesis that BRD4 acts as a switch, controlling gene expression involved in basal EC function, and during angiogenesis under both physiologic (exercise) and pathologic (inflammatory angiogenesis in myocardial ischemia/reperfusion) conditions. Aim 3 will test the hypothesis that BETs modulate EC/T cell interactions and T effector cell differentiation through their control of transcription, limiting T cell-driven experimental autoimmune myocarditis (EAM). Together these studies will help define the role of BET epigenetic reader proteins as novel controllers of dynamic transcriptional programs in ECs, in T cells, and their interaction, yielding new insight into inflammation and angiogenesis.