Friday, May 16, 2025
5/16/2025
A novel mechanism regulating genome-wide mRNA expression in hypoxic lung disease - Project Summary/Abstract Hypoxia, a key contributor to multiple lung diseases, evokes functional and structural responses in target cells by modulating expression of many hundreds of genes. Much is known about regulation of the hypoxic transcriptional response at the single gene level. Induction begins with the reactive oxygen species (ROS)- mediated accumulation hypoxia-inducible transcription factors (Hifs) which, along with coactivators, assemble into multiprotein complexes on hypoxia response elements (HREs) located in gene promoters, enhancers, and perhaps intergenic regions to activate transcription. Far less is known, however, about orchestration of the transcriptional response to hypoxia on a genomic level, with many incongruities between hypoxic exposure, Hif- HRE interactions, and local chromatin restructuring highlighting unresolved complexities of the regulatory apparatus. This proposal addresses the prospect that hypoxia activates a pathway operating in concert with the Hifs to govern the hypoxic transcriptome. Our foundational discoveries along with rapidly accumulating evidence from other fields converge on the concept that ROS generated in hypoxia to initiate Hif accumulation also cause widespread oxidation of guanine to 8-oxoganine (8-oxoG) in distinct motifs nested within DNA regulatory sequences. Oxidized guanines then serve as a novel epigenetic mark by recruiting bifunctional enzymes comprising the DNA Base oxidation and repair pathway (BER). Along with repairing oxidized bases, two of these BER components, specifically OGG1 and Ref-1APE1, direct deployment of canonical enzymes modifying histone acetylation and methylation, thereby governing chromatin accessibility. We call this pathway “BRACR” for Base oxidation and Repair Activated Chromatin Restructuring, and here we propose to explore the novel concept that BRACR is a fundamental component of the gene regulatory apparatus in hypoxia, acting to license HREs and other response elements for transcription factor occupancy by modulating chromatin accessibility. Using cultured human pulmonary arterial cells and intact mice, we will test the hypotheses that: (1) the genome-wide deployment of BRACR in hypoxia requires 8-oxoG formation but not Hifs; (2) BRACR licenses genes for hypoxic regulation through 8-oxoG-mediated engagement of the BER enzymes Ogg1 and Ref-1/APE 1; and, (3) BRACR activation in lung vascular cells drives hypoxic pulmonary hypertension in intact mice. These studies will be transformative. They will define a novel mechanism regulating the hypoxic transcriptome on a genome-wide level, identify new pharmacologic targets to treat hypoxic lung diseases, and because 8-oxoG is mutagenic, may point to a link between transcriptional signaling and somatic mutations that drive malignant and non-malignant pulmonary disorders.
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