Transcriptional Elongation in NF-kB Mediated Inflammation

University Of Texas Med Br Galveston, Galveston TX

Investigators

Linked publications, trials & patents

Abstract

PROJECT SUMMARY (See instructions): Airway inflammatory diseases continue to inflict significant morbidity in the US. NF-KB/RelA is a cytosolic transcription factor that mediates the inflammatory response to cytokines, viruses, oxidative stressors, and allergens. Our studies in the past project period have demonstrated that the NF-KB/RelA activation pathway is bipartite, with one arm releasing RelA from cytoplasmic inhibition, and a separate arm inducing reactive oxygen species (ROS)-dependent, site-specific RelA Ser[276] phosphorylation, required for activating inflammation. Our preliminary studies using TNF-a as stimulus have shown that RelA Ser[276] phosphorylation is a switch required for RelA association with a complex containing the cyclin-dependent kinase (CDK)-9 and bromodomain-4 (Brd4) proteins. CDK9 phosphorylates the C-terminus of RNA Polymerase II, promoting entry into transcriptional elongation mode that produces full-length mRNA transcripts. During this next grant period P1 seeks to understand the mechanism(s) by which RSV modulates NF-KB signaling in the epithelial cell. Our central hypothesis is that NF-KB /RelA Ser[276] phosphorylation couples to an acetyl (Ac) RelA- Brd4-CDK9 pathway that mediates transcriptional elongation as a mechanism for inflammatory gene expression, leading to airway inflammation. Aim 1 will test the hypothesis that RelA Ser[310] phosphorylation is required for RelA Lys[310] acetylation and Brd4-CDK9 recruitment to promote inflammatory gene expression in response to RSV and 8-oxoG in cultured airway cells. Aim 2 will test the hypothesis that CDK9 mediates RNA Pol II C-terminal domain (CTD) phosphorylation, clearance and transcriptional elongation in NF-KB-dependent gene expression in cultured airway cells. Aim 3 will test the hypothesis that NF-KB-induced CDK9 recruitment mediates transcriptional elongation in RSV-induced mucosal inflammation in a mouse model in vivo. This project is highly synergistic with Projects 2-4, informing hypothesis testing and shared experimental work, and relies upon the support of Cores A and B. PI is innovative because it will for the first time establish the mechanism for the RelA phosphorylation/acetylation-transcriptional elongation pathway in RSV and 8-oxoGinduced inflammatory gene expression, and establish this pathway as a central mediator of airway inflammation. These discoveries will lay the foundation for new approaches to selective NF-KB inhibition, for example using CDK9 inhibitors.