Role of PER phosphorylation in circadian biology

生物钟调节表现为生理和行为节律的分子振荡。在所研究的所有系统中，时钟都是通过互锁的转录反馈环控制的，这些环调节时钟mRNAs的日常振荡。通过翻译后机制的调节和微调，这些mRNA振荡精确地转化为时钟蛋白水平的每日周期。动物时钟的一个保守特征是，周期(PER)蛋白每天经历水平和磷酸化状态的振荡，这些水平和磷酸化状态受双重时间(果蝇中的DBT)[哺乳动物中的酪蛋白激酶1e(CK1e)]的调节。在果蝇中，我们的实验室最近表明，过度磷酸化的PER蛋白最终靶向26S蛋白酶体，被F-box蛋白SLimb(哺乳动物中β-TrCP的同源物)降解。这项提案的总体目标是更好地理解翻译后监管在 Clock通过关注Per的磷酸化及其与泛素/蛋白酶体途径的交集来发挥作用。我将把体外分析、基于组织培养的系统和整个动物的方法结合起来。利用黑腹果蝇作为模式动物，为基因操作提供了一个强大的系统。

Circadian clocks regulate molecular oscillations that manifest into physiological and behavioral rhythms. In all systems investigated, clocks are controlled through interlocked transcriptional feedback loops, which regulate the daily oscillations of clock mRNAs. These mRNA oscillations are precisely translated to daily cycles of clock protein levels via regulation and fine-tuning by posttranslational mechanisms. A conserved feature of animal clocks is that PERIOD (PER) proteins undergo daily oscillations in levels and phosphorylation states that are regulated by DOUBLETIME (DBT in Drosophila) [casein kinase 1e (CK1e in mammals)]. In Drosophila, our lab recently showed that hyperphosphorylated PER proteins are eventually targeted to the 26S proteasome for degradation by the F-box protein Slimb (a homolog of beta-TrCP in mammals). The overall goal of this proposal is to better understand the role of posttranslational regulation in clock function by focusing on PER phosphorylation and its intersection with the ubiquitin/proteasome pathway. I will integrate in vitro assays, tissue culture based systems, and whole animal approaches. The use of Drosophila melanogaster as the model animal provides a powerful system for genetic manipulations.