PhD

dissertationTo coordinate behavior and physiology with changes in the environment, nearly all organisms employ a circadian clock. In most cases, the circadian clock maintains a period of almost, but not exactly, 24 hours. To keep pace with a 24-hour circadian dark/light cycle, the clock can be reset (or entrained) in response to external environmental cues, most notably light. In mice, the master circadian clock is located within the suprachiasmatic nucleus and consists of a complex network of interconnected transcription-translation feedback loops in which regulatory proteins experience daily oscillations in abundance. Transcription of the negative regulators Period1 and 2 (denoted mPer1 and mPer2) is driven by the CLOCK/BMAL1 heterodimeric transcription factor. After reaching a threshold level, the mPER proteins translocate to the nucleus and inhibit their own transcription. The mPER proteins are then phosphorylated and subsequently degraded, thereby relieving repression of transcription. The clock is said to be reset and is now poised for another cycle of transcription, translation and transcription repression. Although diverse lines of evidence have linked the serine/threonine protein kinase CKI? to mPER protein phosphorylation and stability, the mechanism by which CKI? regulates circadian rhythm is largely unknown. This report further defines the role of CKI? within the mammalian circadian clock. First CKI? binds and phosphorylates mPER1 and mPER2 and regulates rnPER1 subcellular localization. Second, CKI? phosphorylates the circadian transcription factor BMALl and increases its transactivation activity towards a circadian promoter. Third, CKI? directly regulates mPER2 stability by promoting binding of the ubiquitin ligase (?-TrCP to mPER2. Fourth, mPER2 net phosphorylation and stability depends on the coordinate activity of CKI? and an as of yet identified protein phosphatase