Role of Chromatin Remodeling in Circadian Regulation

DESCRIPTION (provided by applicant): Circadian regulation in Neurospora crassa is controlled in part by a negative feedback loop where the Frequency (FRQ) protein inhibits its own transcription through a direct interaction with the White Collar complex (WCC). Currently there is no information on chromatin organization at the frequency (frq) gene promoter and what, if any type of chromatin remodeling occurs during the circadian cycle. I propose to examine the role of chromatin remodeling in circadian regulation through three specific aims. First, I plan to map the nucleosome positions at the frq promoter over the circadian cycle and in response to light induction using ligation-mediated polymerase chain reaction (LM-PCR). Relative changes in nucleosome positions that occur will be confirmed by DNase I protection and restriction site accessibility assays. Secondly, I plan to determine the covalent modifications that occur on the amino-terminal tail domains of the histones using the chromatin immunoprecipitation (ChiP) assays. Finally, I will order the histone modifications that occur at the frq locus to develop a circadian regulated 'histone code'. The completion of these aims will further both the circadian biology and chromatin remodeling fields.