Mechanisms Regulating Chromatin Organization and Epigenetic Memory - SUMMARY Upon fertilization, the zygotic genome is dramatically remodeled to achieve a transient totipotent state that is necessary to lay the foundation for embryonic development and differentiation. Activation of the genome requires choreographed contacts between enhancers and promoters. Mutations in the genes that encode the machinery required to orchestrate this remodeling and contact are causative of numerous developmental syndromes or embryonic lethality. Despite its unequivocal importance for child development, reproduction and fertility, the relationship between chromatin structure and enhancer-promoter communication that underpins correct developmental programming remains poorly understood, particularly in vertebrates. In addition, the factors and motifs that mediate these structures and gene regulatory communication, including their specific functions, have not been extensively studied. A major barrier to understanding these relationships in the dynamic system of a developing vertebrate has been a dearth of suitable approaches that can achieve genome-wide and high-resolution information given the limitations in input material. To overcome these limitations, in our preliminary studies we have optimized techniques to detect novel regulatory factors and chromatin structures with unprecedented resolution in zebrafish embryos. In addition, our recently developed expansion microscopy technique now allows us to address fundamental questions about how enhancers and promoters interact and transmit information at nucleosome resolution. The confluence of development of these techniques now allows us to investigate the molecular mechanisms that regulate genome activation and gene expression in a dynamic vertebrate system. Using these approaches, we aim to understand how specific factors contribute to the establishment of the 3D genome architecture as well as investigate a mechanism for explaining how transient genomic contacts can propagate longer-term regulatory information (Aim 1), and identify the repertoire of regulatory elements that mediate enhancer-promoter interactions as well as the factors and their motifs that regulate gene expression (Aim 2). By defining the regulatory grammar and mechanisms guiding genome activation in the zebrafish embryo, this work will fill critical gaps in our knowledge of gene regulation in a dynamic developing system, and provide valuable insights with broad implications for developmental biology, epigenetics, and reproductive medicine.
