Role of PER phosphorylation in circadian biology

生物钟调节分子振荡，表现为生理和行为节奏。在所有研究的系统中，时钟都是通过互锁的转录反馈回路来控制的，这些反馈回路调节时钟mRNA的日常振荡。这些mRNA振荡通过翻译后机制的调节和微调被精确地翻译为时钟蛋白水平的每日周期。动物生物钟的一个保守特征是PERIOD（PER）蛋白的水平和磷酸化状态每天发生振荡，受DOUBLETIME（DBT在果蝇中）[酪蛋白激酶1 e（CK 1 e在哺乳动物中）]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26 S蛋白酶体，由F-box蛋白Slimb（哺乳动物中β-TrCP的同源物）降解。本提案的总体目标是更好地理解翻译后调节在以下方面的作用： 通过关注PER磷酸化及其与泛素/蛋白酶体途径的交叉来研究生物钟功能。我将整合体外分析，组织培养为基础的系统，和整个动物的方法。果蝇作为模式动物的使用为遗传操作提供了一个强有力的系统。

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.