Role of Multimerization in H3.3/H4 deposition by the HIRA complex

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Project Summary The goal of this project is to characterize, with molecular detail, the role that multimerization of the HIRA histone chaperone complex plays in depositing histone H3.3/H4 onto DNA. In order to fit roughly two meters worth of DNA into a ten-micron cell nucleus, histones are used as scaffolding proteins to condense linear genetic material into the higher order structures of chromatin. Epigenetic regulatory machinery is essential to ensure that the chromatin landscape is well maintained to allow for certain gene expressions to be readily activated or inactivated to regulate both cell fate and cell stability. The HIRA histone chaperone complex is responsible for delivering histone H3.3/H4 onto nascent DNA in areas of active gene transcription and is therefore implicated in chromatin changes during development, in maintenance of terminally differentiated cell types, and in aging. The strategies I propose will first provide understanding of the molecular basis of the interaction between the C-terminal trimerization domain of HIRA and CABIN1 and then will probe the relevance of these interactions in vitro by investigating the effect that mutants have on the histone deposition process using a nucleosome assembly assay and in cells by investigating the effect that mutants have on complex localization and histone deposition within the crowded nuclear landscape. The proposed study will deepen our understanding of a ubiquitous epigenetic mechanism essential for regulating the chromatin environment of healthy human cells.