Analysis of clock proteins in their non-circadian roles

描述（由申请人提供）：生物钟是行为和生理过程的进化保守协调者。人体生物钟的故障会导致严重的疾病，如睡眠障碍和癌症。昼夜节律的计时是通过分子反馈回路来完成的，其中涉及到几个时钟基因及其蛋白质。两个基因period（per）和timeless（tim）的作用已经在生物钟反馈回路中得到了很好的证实，该回路在模式生物黑腹果蝇（Drosophila melanogaster）中起作用。这两个基因的产物，蛋白质PER和TIM，转移到细胞核，随后降解;这两个事件是必不可少的时钟功能。令人惊讶的是，在卵巢中，这些蛋白质的行为不同。它们的水平不循环;相反，它们始终保持稳定和细胞质。尽管如此，卵巢PER和TIM在调节卵子生产中具有重要的功能。我们假设PER和TIM在卵巢中可能经历了与在时钟细胞中不同的翻译后修饰。我们建议使用生物化学和遗传学的工具来测试这一假设在三个特定的目标。首先，我们将研究卵巢中PER和TIM的相互作用及其功能意义。其次，我们将研究PER和TIM的翻译后修饰与磷酸化。最后，我们将探讨卵巢中缺乏TIM降解的原因。本研究的结果将为我们深入了解果蝇卵巢中时钟蛋白非昼夜节律表达的功能意义提供重要线索。越来越多的证据表明，被认为专门充当时钟组件的基因还具有其他重要的多效性作用。它们在果蝇和哺乳动物中都以非昼夜节律的方式起作用。因此，了解模式生物中时钟基因的非昼夜节律功能应该为人类健康提供有价值的见解。

DESCRIPTION (provided by applicant): Circadian clocks are evolutionary conserved coordinators of behavioral and physiological processes. Malfunctions of circadian clocks in humans lead to serious pathologies such as sleep disorders and cancer. Circadian timekeeping is accomplished by molecular feedback loops that involve several clock genes and their proteins. The role of two genes period (per) and timeless (tim) has been well established in the clock feedback loop, which operates in the model organism Drosophila melanogaster. The products of these two genes, proteins PER and TIM, translocate to cell nuclei and are subsequently degraded; both events are essential for clock function. Surprisingly, in the ovary, these proteins behave differently. Their levels do not cycle; instead, they remain stable and cytoplasmic at all times. Despite such unorthodox behavior, ovarian PER and TIM have important functions in the modulation of egg production. We hypothesize that PER and TIM may undergo different post-translational modifications in the ovary than they do in the clock cells. We propose to use biochemical and genetic tools to test this hypothesis in three specific aims. First, we will study interactions of PER and TIM in the ovary and their functional significance. Second, we will investigate posttranslational modifications of PER and TIM related to phosphorylation. Finally, we will explore the reasons for the lack of TIM degradation in response to light in the ovary. Results obtained in this study will give us important insights into the functional significance of non-circadian expression of clock proteins in the fly ovary. There is increasing evidence that genes that were thought to act exclusively as clock components have other important pleiotropic roles. They act in a non-circadian manner in both Drosophila and mammals. Therefore, understanding the non-circadian functions of clock genes in a model organism should provide valuable insights into human health.