REGULATORY MECHANISMS OF LINKER HISTONES AND THEIR POST-TRANSLATIONAL MODIFICATIONS PROJECT SUMMARY Chromatin structural dynamics control the accessibility of DNA to complexes that regulate transcription, repair DNA damage and replicate DNA. A ubiquitous regulator of chromatin dynamics is the linker histone H1. In metazoan somatic cells, linker histones are highly abundant with almost one linker histone per nucleosome, and regulate gene expression during development and within somatic cells. Changes in linker histone expression and their post-translational modifications are strongly associated with disease, including ovarian cancer, while linker histone mutations are linked to follicular lymphoma. Amazingly, in spite of H1 abundance and health relevance, this key chromatin architectural protein and its modifications remain poorly understood because of the lack of quantitative tools to investigate H1 function. Recently, we developed ensemble and single molecule fluorescence based tools to investigate linker histone and chromatin interactions, and a hybrid ligation method to prepare for the first time homogenously modified linker histones. Using these new methods we recently discovered: (i) Nucleosomes remain dynamic while H1 is bound, directly challenging the fundamental idea that H1 holds nucleosomes and chromatin in compact static structures. (ii) A single post-translational modification, acetylation of histone H3 at lysine 56, abolishes the influence of H1 on transcription factor binding within the nucleosome. These findings have inspired us to investigate the hypothesis that H1 functions as a dynamic regulator of chromatin dynamics and accessibility. We will leverage our new tools and findings to investigate the following Specific Aims: (1) Elucidate how H1 remains dynamics while regulating DNA accessibility within chromatin. (2) Determine the influence of histone H3 and H4 PTMs on H1 targeting and regulation of chromatin dynamics and accessibility. (3) Determine the influence of H1 PTMs on H1 targeting and its regulation of chromatin dynamics and accessibility. Together these studies will provide a foundation for understanding linker histone regulation of nucleosome and chromatin dynamics to control accessibility to complexes that transcribe, repair and replicate chromatin.
Regulation of gene expression and genomic stability by epigenetic factors such as histone modifications, chromatin architectural proteins such as the linker histone H1 and chromatin dynamics have become a dominant field in cancer epigenetics because of the reversible nature of epigenetic alterations. Recent advances in understanding epigenetic factors have led to the emergence of epigenetic therapy and the recent FDA approval of multiple epigenetic drugs for cancer treatment. This proposal uses a combination of advanced protein chemistry and single molecule approaches to reveal the molecular mechanisms behind the function of chromatin epigenetic regulators. These studies will provide molecular information for understanding drug resistance, tumorigenesis, and for designing new cancer drugs and therapies.
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