How do VRI and PDP1 regulate circadian rhythms?

描述（由申请人提供）：一个晚上的睡眠紊乱就足以让我们意识到正常的睡眠/觉醒节奏对身心健康的重要性。这些节律是由体内的昼夜节律（约24小时）时钟控制的，我们从果蝇的研究中了解了生物钟的大部分功能。事实上，第一个被发现的人类睡眠障碍基因是生物钟基因的同源物，最早是在果蝇身上被发现和克隆的。作为一种简单的行为，昼夜节律为理解大脑如何控制行为的基本机制提供了机会。特别是，果蝇时钟是了解转录因子如何动态调节神经元功能的一个很好的系统，这是神经科学中一个广泛而重要的话题。在此，我们建议研究时钟转录因子如何控制时钟神经元的活动。我们重点研究了Vrille （VRI）和PDP1，它们构成了果蝇的第二个时钟反馈回路。VRI和PDP1在维持反馈回路本身的稳健性方面发挥作用，并且是通过调节在黎明达到峰值的基因将生物钟与节律输出联系起来的最下游因素。

DESCRIPTION (provided by applicant): One night of disturbed sleep is all that it takes for us to realize how important normal sleep / wake rhythms are for mental and physical health. These rhythms are controlled by an internal circadian (~24hr) clock, and we have learnt most about how the clock functions from studies in Drosophila. Indeed, the first human sleep disorder gene identified is a homologue of the period clock gene, first identified and cloned in Drosophila. As a simple behavior, circadian rhythms provide the opportunity to understand fuindamental mechanisms of how brains control behavior. In particular, the Drosophila clock is an excellent system to understand how transcription factors dynamically regulate neuronal function, a topic of broad general importance in Neuroscience. Here, we propose to study how clock transcription factors control clock neuron activity. We focus on Vrille (VRI) and PDP1, which comprise the second Drosophila clock feedback loop. VRI and PDP1 have roles in maintaining the robustness of the feedback loops themselves, and are the most downstream factors that link the clock to rhythmic outputs by regulating genes that peak at dawn. In Aim 1, we propose a detailed analysis of the VRI/PDP1 clock loop. We have identified that additional regulators exist in this loop. One regulator is Cryptochrome (CRY), previously characterized as a circadian photoreceptor, but recently established by my lab to also function as a transcriptional represser in most clock neurons. Here we propose to study cry regulation by PDP1 in different clock neurons. A second regulator of vri and Pdp1 expression may be Stich1, a homologue of the mammalian Dec clock proteins, which are potent transcriptional repressors, but have not been formally fitted into the clockworks. Finally the identification of a cry null allele will allow us to test in which clock neurons in the brain CRY is a repressor. In Aim 2, we seek to extend exciting findings that should help link the clock to output pathways. We have developed a behavioral genomics approach in which gene expression from purified pacemaker neurons can be assayed across the whole genome. We have currently analyzed different times of day, but we will extend this analysis to different clock mutants with known effects on the outputs of pacemaker neurons. Thus we can correlate patterns of gene expression with known behavioral outputs. In our initial studies, we identified a group of genes which may underlie rhythmic neuronal activity and neurotransmission from pacemaker neurons. We propose to study mutants in these genes for defects in circadian behaviors. Many previously unstudied genes are also tightly clock-regulated, and we will narrow done which to study further through additional GeneChip experiments in vri and Pdp1 mutants.