Analysis of clock proteins in their non-circadian roles

描述（由申请人提供）：生物钟是进化保守的行为和生理过程的协调者。人类生物钟的故障会导致严重的疾病，如睡眠障碍和癌症。昼夜计时是由分子反馈回路完成的，其中包括几个时钟基因及其蛋白质。两个基因周期（per）和永恒（tim）的作用已经在时钟反馈回路中得到了很好的确立，这在模式生物果蝇中起作用。这两个基因的产物，蛋白质PER和TIM，转运到细胞核并随后被降解；这两个事件对于时钟的功能都是必不可少的。令人惊讶的是，在卵巢中，这些蛋白质表现不同。它们的水平不循环；相反，它们在任何时候都保持稳定和细胞质。尽管有这种非正统的行为，卵巢PER和TIM在调节卵子产量方面具有重要的功能。我们假设PER和TIM在卵巢中的翻译后修饰可能不同于它们在时钟细胞中的修饰。我们建议使用生化和遗传工具在三个具体目标中检验这一假设。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，我们将研究与磷酸化相关的PER和TIM的翻译后修饰。最后，我们将探讨卵巢对光缺乏TIM降解的原因。本研究结果将为我们了解果蝇卵巢生物钟蛋白非昼夜节律表达的功能意义提供重要见解。越来越多的证据表明，那些被认为只起生物钟作用的基因还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.