Control of histone ubiquitylation during the cell cycle

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PROJECT SUMMARY The site-specific ubiquitylation of histone proteins can dictate whether a gene gets expressed or silenced, thus, this type of modification is indispensable for normal development and cellular homeostasis. By contrast, the mis-regulation of histone ubiquitylation can result in developmental lethality, neurodegeneration, and aging. ~1–2% of histone H2B is mono-ubiquitylated (H2Bub1) during interphase, ranking this protein alongside some of the most ubiquitylated by abundance. H2Bub1 has been shown to promote gene expression through multiple mechanisms. Strikingly, H2Bub1 disappears from metaphase chromosomes and reaccumulates as cells exit from mitosis, but considering recent reports, which challenge the long-held view that histones function to condense chromatin into higher-order structures in vivo, why the state of H2B ubiquitylation needs to be reset during each cell cycle is incompletely understood. Thus, there is a pressing need to elucidate why and how H2Bub1 is stripped from mitotic chromosomes. Multiple deubiquitylases (DUBs) have been shown to catalyze the removal of ubiquitin from H2Bub1, nonetheless, it remains to be seen how they cooperate with one another, if there are yet unidentified DUBs specific for H2Bub1, the genomic sites they regulate, and when they function during the cell cycle. Our working hypothesis posits that each H2Bub1-specific DUB targets overlapping but distinct genomic regions to ensure the rapid clearance of H2Bub1 on mitotic chromosomes. To address this hypothesis, we will use a multidisciplinary approach, integrating classical biochemical approaches with global measurements. More specifically, we propose to identify the key DUBs that target H2Bub1, to elucidate the extent of their redundancy, to characterize their mechanisms of action, and to examine how they influence gene expression throughout the cell cycle. Though this work aims to uncover the purpose of histone deubiquitylation during mitosis, we also expect to gain new insights into the basic principles of ubiquitin- dependent gene regulation, including mechanisms of spatiotemporal control, cooperativity and interplay between DUBs, and crosstalk with other chromatin-modifying enzymes. Finally, our work will have major implications in how dividing cells maintain their transcriptional memory to ensure the faithful propagation of cellular identity to future generations, and this study will have a positive impact by providing a fresh context for understanding ubiquitin-dependent disorders and may reveal how best to develop therapeutic strategies to combat these diseases.