Dynamics of DNA-histone interactions

Project Summary The overarching goal of my research program is to elucidate the roles of dynamic DNA-histone interactions in the nucleosome in regulating gene accessibility from single-molecule approaches. The nucleosome is the fundamental packing unit of genes in eukaryotes and plays important roles in gene regulation. Errors in gene regulation often lead to developmental failure and lethal diseases such as various types of cancer. The nucleosome is made of ~147 bp ds-DNA with an octameric histone protein core. DNA and histones are targets for various chromatin modifications that are often critically implicated in gene regulation mechanisms. We reported several important changes in the structure and structural dynamics of the nucleosome induced by various chromatin modifications and histone variants such as DNA methylation, histone acetylations, SUMOylation, CENP-A, and H2A.Z in the context of their roles in gene regulation. As these changes are heterogeneous, unsynchronized, and/or complex, they are often impossible to resolve with ensemble-averaging biochemical, genetic, and static structural biology tools. We develop and improve single- molecule fluorescence methods and systems, mainly based on FRET, FCS, and colocalization at an optical super-resolution to address these problems. We will continue pushing the boundaries of these experimental systems and methods to support our future research that will center around three synergistic themes: how (1) chromatin modifications and histone variants, (2) spontaneous molecular motions and interactions, and (3) active and passive molecular machines and enzymes affect DNA-histone dynamics in nucleosomes and nucleosome arrays and how the effects are implicated in gene regulation mechanisms. We recently started evolving our mostly nucleosome-focused experimental systems by combining various enzymes that act on the nucleosome and expanding them to include nucleosome arrays. Based on these systems, our research will continue largely in three topical areas in the next five years: (1) nucleosome dynamics during its interaction with complex enzymes such as RNA polymerase II and Dot1L, a key player in H3K79 methylation-dependent leukemia pathogenesis, (2) the effects of chromatin modifications on the structure and dynamics of the nucleosome and their implications in nucleosome-enzyme interactions, (3) conformations and dynamics of nucleosome arrays and the effects of chromatin modifications on nucleosome dynamics in nucleosome arrays. Investigations in these areas will help understand the fundamental molecular processes that regulate nucleosome dynamics and gene accessibility during nucleosome-enzyme interactions in a nucleosome and a nucleosome-array context and at depth and clarity afforded by our highly-refined tractable single-molecule systems.