Abstract 9108: Identification and Validation of Protein Modules Driving Chromatin Remodeling in Cardiac Hypertrophy and Failure
Chromatin remodeling is a necessary precursor to global changes in gene expression-however, the mechanisms for genome-wide changes in chromatin state are poorly understood. In the heart, diseases like hypertrophy and failure are well known to be driven by large gene expression changes that enact a fetal-like phenotype, particularly in the areas of metabolism, calcium handling and contraction. Previous work has investigated classes of histone modifying enzymes that alter chromatin accessibility during cardiac hypertrophy. What remains unknown is how changes in histone modifications and transcription factor activation are interfaced at the level of nucleosome remodeling. To discover modules of proteins involved in nucleosome remodeling in heart failure, quantitative proteomics was used to measure chromatin-associated proteins during different stages of heart disease in a mouse model of pressure overload (transverse aortic constriction). Out of 733 proteins detected, 366 of which showed quantitative changes during disease, unsupervised clustering was used to reveal modules of proteins with statistically similar (p<0.01) changes in expression. These modules were prioritized using gene ontology-based bioinformatics: a candidate module, containing 63 proteins, showed significant increase in all members during hypertrophy and further increased during failure. To investigate the role of this module in cardiac phenotype, a high-throughput loss of function screen was carried out using morpholino-based knockdown in zebrafish. Candidates were further prioritized based on their predicted involvement in nucleosome insertion/eviction. Among the initial proteins investigated were Ncl, Nap1l4 and Rcc1. Loss of Ncl led to defects in cardiac function and morphology exhibited by severe edema, regurgitation and dysregulation of chamber size and patterning. In addition, siRNA-mediated knockdown was used to investigate the function of individual members in neonatal rat ventricular myocytes. This investigation reports on a novel discovery method in a clinically-relevant model of heart failure, interfaced with a rapid phenotypic screening tool in zebrafish to identify and physiologically characterize novel chromatin remodelers in the heart.
- © 2011 by American Heart Association, Inc.