Histone Nuclear Import and Chromatin Assembly

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DESCRIPTION (provided by applicant): The long term goal of this project is to understand how the functions of nuclear transport factors and histone chaperones are coordinated, and regulate the assembly of histones into specific chromatin domains. We will determine how histone chaperones dictate the movement of histones in the cell and thereby serve as general regulators of histone function. Every cell cycle the entire DNA content of a cell is rapidly replicated and repackaged with newly synthesized histones to form nucleosomes. In addition histones are exchanged with those already in chromatin, to allow the addition of variant histones, and to allow passage of polymerases and DNA repair machineries. Histones are imported into the nucleus by specific members of the karyopherin family. Histone chaperones are also key players in histone metabolism that bind histones shortly after synthesis and escort them to the nucleosome. This project focuses on the role of histone chaperones in histone import and export, and the significant question of whether histone chaperones regulate the movement of histones in the cell. This project builds from previous observations that the yeast histone chaperone Nap1 p is a histone nuclear import factor. Specifically, we will test the hypothesis that specific histone chaperones for H3 and H4, such as Asflp, regulate histone nuclear import and export. The hypothesis that the histone chaperone Naplp acts as an H2A.Z donor and acceptor, and regulates the cellular localization of H2A.Z will be tested. Lastly the hypothesis will be tested that distinct reversible phosphorylation events regulate cycles of Naplp function, allowing it to perform distinct role in the nucleus and cytoplasm. Analysis of phosphorylation sites in the context of the recent crystal structure will allow an understanding of the mechanism of Naplp regulation. The correct assembly of nucleosomes is critical for maintaining genomic stability in all cells. The dysfunction of proteins involved in nucleosome assembly and remodeling pathways is also coincident with human disease and cancer. This highlights the importance of dissecting the function of histone chaperones in yeast as a model organism, and will give us useful information on the role of histone chaperones in human physiology and human disease.