When faced with chronic stress, the heart enters a compensatory hypertrophic stage; without intervention it eventually succumbs to decompensation marked by a dilated left ventricular chamber and decreased ejection fraction. While the morphological cardiac remodeling that occurs during the progression of heart failure is well characterized, the exact molecular cause for this gradual switch to failure is not known. In addition to the numerous alterations in signaling pathways, a conserved switch in the transcriptome, known as the fetal gene program, occurs during hypertrophy as a protective effort to sustain contractility by reverting to fetal isoforms of metabolic, contractile and calcium handling genes. We hypothesize that the reproducible, coordinated reprogramming of gene expression is orchestrated by a change in chromatin structure that enables pathologic gene expression. To determine the proteins involved in repackaging chromatin during cardiac pathology, we performed quantitative proteomic analyses of nuclear proteins in a mouse model of pressure overload hypertrophy and failure. Among the hundreds of proteins we measured on chromatin, my subsequent analyses have focused on two candidates that had the potential to alter gene expression by directly affecting chromatin packing. The first was Nucleolin, a major component of the nucleolus where it mediates ribosomal biogenesis. Using isolated myocytes and the developing zebrafish embryo, we uncovered a role for Nucleolin to regulate cardiac looping, with its effect on hypertrophy context dependent, such that in isolated myoctyes knockdown can promote pathologic gene expression, but loss of Nucleolin during development does not alter myocyte size, instead affecting differentiation along the cardiac lineage. The second protein I functionally validated was High mobility group protein B2 (HMGB2), a non-histone chromatin structural protein that increases 3-fold in our proteomic analyses. We show that HMGB2 is necessary for ribosomal RNA transcription and is enriched in the nucleolus in hypertrophy; however, overexpression of HMGB2 shuts down transcription globally by compacting DNA. Furthermore, we find HMGB2 knockdown alters the chromatin environment of individual gene promoters in the same manner as hypertrophic agonist signaling in isolated myocytes. Finally, we find that the effect of HMGB2 abundance on the expression of individual genes can be partially explained by the chromatin context, and specifically identify a novel relationship between HMGB2 and CTCF. These studies add to the growing body of work characterizing chromatin remodeling in hypertrophy, and demonstrate that this remodeling extends outside of gene bodies and promoters. Finally, this work begins to uncover what features of chromatin are responsible for tailoring the effects of ubiquitous chromatin proteins toward a cell-type specific outcome.

The underlying mechanisms by which cell identity is achieved in a cell type-specific manner during development are unknown. In this project, we examine the mechanisms through which genomic architecture is regulated by different protein factors and how these proteins in turn regulate gene expression in the cardiomyocyte. We search for cardiac chromatin structural factors that are important for the establishment of genomic architecture during differentiation. We hypothesized that these candidates would also be implicated in pathological gene expression upon the onset of heart failure. Instead, we found that the expression changes of chromatin structural genes across a panel of different mouse strains were not universal, nor did they correlate with cardiac phenotype after pathological stress. Most of our current knowledge of signaling mechanisms in the heart has stemmed from genetic manipulations in a single mouse strain. Here, we examined well-characterized regulators of cardiac phenotype and showed that the relationships between gene expression and cardiac phenotype are lost when expanding across multiple genetic backgrounds. More importantly, these data demonstrate that there is no single signature gene that drives heart disease (nor is there a single gene whose expression is a biomarker of the condition), highlighting the role of genetic variability to differentially sculpt the transcriptome in the development and progression of complex diseases. In addition, our findings demonstrate that regulation of gene expression by genetics occurs in a tissue-dependent manner. We previously identified High Mobility Group B2 as an important chromatin structural protein in the heart and showed its involvement in pathological gene expression. These studies suggested this regulation occurs by remodeling global transcriptional activity. To characterize structural organization of cellular transcription, we show that transcriptional activity is compartmentalized into stable factories in the heart that undergo functional changes in vivo in response to disease stimuli. We provide evidence of direct reorganization of genomic structure by showing that nuclear positioning of cardiac genes with respect to chromatin environments and transcription factories correlates with changes in their expression. In summary, this project explores the mechanisms of cardiac gene regulation and illustrates multiple levels of regulation, with influences from genetics and chromatin architecture.

Long-Range Control of Gene Expression covers the current progress in understanding the mechanisms for genomic control of gene expression, which has grown considerably in the last few years as insight into genome organization and chromatin regulation has advanced. Discusses the evolution of cis-regulatory sequences in drosophila Includes information on genomic imprinting and imprinting defects in humans Includes a chapter on epigenetic gene regulation in cancer

This volume focuses on the relevance of epigenetic mechanisms in autoimmune disease. It provides new directions for future research in autoimmune disease.

HiC-Pro is an optimized and flexible pipeline for processing Hi-C data from raw reads to normalized contact maps. HiC-Pro maps reads, detects valid ligation products, performs quality controls and generates intra- and inter-chromosomal contact maps. It includes a fast implementation of the iterative correction method and is based on a memory-efficient data format for Hi-C contact maps. In addition, HiC-Pro can use phased genotype data to build allele-specific contact maps. We applied HiC-Pro to different Hi-C datasets, demonstrating its ability to easily process large data in a reasonable time. Source code and documentation are available at http://github.com/nservant/HiC-Pro.

Development of the heart is a complex process and can lead to serious congenital disease if the process goes awry. This book provides a detailed description of the cell lineages involved in heart development and how their migration and morphogenesis are controlled. It also examines the genetic and environmental bases for congenital heart disease and how model systems are revealing more about the processes involved. Topics covered in this essential volume include: - Anatomy of a Developing Heart - Genetic and Epigenetic Control of Heart Development - Development of the Cardiac Conduction System - Genetic Basis of Human Congenital Heart Disease - In Vivo and In Vitro Genetic Models of Congenital Heart Disease