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 TNF? signal to endothelial chromatin, inducing coordinated genomic BRD4 redistribution to de novo super-enhancers that help drive transcription of the pro-inflammatory NF?B 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 TNF?-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 TNF?/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.
PI: Plutzky, J We have uncovered that a family of proteins known as BETs help control which genes are turned on and off in the endothelial cells that line the inside of blood vessels and inflammatory cells known as T cells. Through this action, BETs help control large programs of gene expression involved in normal endothelial cell and T cell function, how these cells interact with each other, and processes like inflammation and new blood vessel growth.
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