The long term goal of this project is to understand the molecular mechanism of circadian clocks in eukaryotic cells. Circadian clocks are fundamental cellular processes by which almost all organisms use to keep time, and they control a wide variety of cellular, physiological, and behavioral activities. The importance of circadian clock in human physiology and mental health is evident from its ubiquitous influence on a wide range of cellular and physiological processes, including sleep/wake and body temperature cycles, endocrine functions, drug tolerance and resistance, and jet lag. The malfunction of the clock is known to be associated with several forms of human psychiatric illness and sleep disorders. Despite their importance, however, clues to the underlying molecular mechanism are only just beginning to emerge. To achieve our goal, we are focusing our studies in the model system Neurospora crassa, which has one of the best understood clock systems. In Neurospora, frequency (frq) gene is an essential regulatory component in the clock. We have found that frq gene product, FRO, is progressively phosphorylated and the phosphorylation of FRO is important for its degradation and the clock function.
In Specific Aim 1, we will use both genetic and biochemical approaches to identify the kinase(s) that phosphorylates FRO. The gene(s) encoding the kinase(s) will be cloned and its clock function analyzed. We know that the degradation of FRO is an important regulatory aspect of the clock, therefore, the understanding the mechanism by which FRO is degraded will be our focus in Specific Aim 2. We will test several specific hypotheses of the mechanism of FRO degradation, and we will examine the role of FRO degradation in the clock. Temperature is a major clock affecting environmental factor and it affects several fundamental clock properties for all circadian clocks.
In Specific Aim 3, we will study the molecular mechanisms of temperature regulations in the Neurospora clock. Specifically, we will study how temperature regulates the expression of FRO and the molecular mechanism of temperature compensation of the clock. Together, these studies will allow us to describe the framework of the Neurospora clock in genetic and molecular terms.
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