Genetic and Molecular Dissection of the Neurospora Clock

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Virtually all eukaryotic organisms appropriately examined have been shown to possess the capacity for endogenous temporal control and organization known as a circadian rhythm. The cellular machinery responsible for generating rhythms is collectively known as the biological clock. A healthy circadian clock underlies both physical and mental health. Because of the ubiquity of its influence on human mental and physiological processes - from circadian changes in basic human physiology to the clear involvement of rhythms in work/rest cycles and sleep - understanding the clock is basic to prevention and treatment of many physical and mental illnesses, from metabolic disorders to sleep/wake dysfunction and cancer. Our research uses genetic and molecular studies of the model eukaryote Neurospora, as well as mammalian cells in culture, to further our understanding of the organization of the circadian oscillator, a one- step transcription-translation feedback loop whose regulatory architecture is conserved from fungi to mammals. Planned research lies within three foci. Focus #1 builds upon our understanding of the interplay between structure and function in core clock components. We will determine how phosphorylations and interactions among clock components lead to repression within the feedback loop; address a controversy as to whether negative element turnover has a role in the mammalian oscillator; probe how clock-controlled phosphorylation guides essential interactions and activities of clock components leading to the canonical circadian property of temperature compensation, and how modulation of RNA metabolism and gene expression contribute to nutritional compensation. Focus #2 pioneers new territory and exploits recently developed techniques, expanding the use of cell biological tools to complement genetics in defining the spatio-temporal dynamics of clock components within the cell. We will show how, as well as where in the cell the clock operates. Focus #3 will build upon our strong grounding in the genetics and genomics of light-regulation, using computational and informatic tools to define the hierarchical network of transcription factors that govern the response of Neurospora to light and time. The aim is to provide the first concrete model for global circadian control of a eukaryotic genome. Our long term goals are to describe, in the language of genetics and biochemistry, the feedback cycle comprising the circadian clock, how this cycle is synchronized with the environment, and how time information generated by the feedback cycle is used to regulate the behavior of cells and organisms. These projects are complementary and mutually enriching in that they rely on genetic and molecular techniques to dissect, and ultimately to understand, the organization of cells as a function of time.