Mechanisms of Circadian Clock Output

描述（由申请人提供）：我们对昼夜节律起搏神经元用于组织日常运动行为的信号传导机制感兴趣。近年来，在确定细胞自主时钟机制的分子基础方面取得了巨大进展，该机制是昼夜节律计时的基础。这一定义允许识别大脑中的主要生物钟神经元。反过来，这也提供了一个机会，重新审视有关行为的神经基础的基本问题。该研究计划采用细胞，分子和遗传技术来了解信号如何从主要起搏器传递到反应神经元。以前我们表明，神经肽PDF是果蝇的主要昼夜节律递质。遗传学证据表明，它是初级时钟细胞（LN-v神经元）使用的最重要的信号。我们将通过实验来扩展这些信息，这些实验旨在询问PDF在大脑中的何处以及何时发出昼夜节律信号。此外，我们将鉴定和表征PDF的受体。PDF受体的定义将允许描述哪些神经元和哪些回路响应昼夜节律递质。我们也可以开始询问PDF受体激活后的长期结果（信号转导途径）是什么。由于昼夜节律信号发生在这样一个长的时间基础上（许多小时），我们假设PDF信号可能包括转录调控。我们已经确定了几个基因，其表达水平是pdf依赖性的，每天循环。现在，我们建议证明他们的作用，PDF的调解昼夜信号的目标。最后，我们进行了广泛的全基因组微阵列屏幕的昼夜节律基因表达的果蝇头部。我们已经确定了22个昼夜节律基因，其中15个是新的昼夜节律生物学的背景下。我们假设这些昼夜节律基因中的大多数参与昼夜节律输出机制，并提出实验来解决这种可能性

DESCRIPTION (provided by applicant): We are interested in signaling mechanisms used by circadian pacemaker neurons to organize daily locomotor behavior. There has been tremendous progress in recent years in defining the molecular basis of the cell autonomous clockwork mechanism that underlies circadian timekeeping. That definition has permitted the identification of the primary circadian clock neurons within the brain. In turn, that has presented the opportunity to re-examine fundamental issues concerning the neural basis of behavior. This research program features cellular, molecular and genetic techniques to understand how signals are passed from the primary pacemakers to responsive neurons. Previously we showed that the neuropeptide PDF is the principal circadian transmitter in Drosophila. Genetic evidence indicates it is the most important signal used by the primary clock cells (the LN-v neurons). We will extend that information by performing experiments that are designed to ask where and when PDF signals circadian time within the brain. Further, we will identify and characterize the receptor(s) for PDF. Definition of the PDF receptor will permit a description of which neural elements and which circuits respond to the circadian transmitter. We can also begin to ask what are the long-term consequences (signal transduction pathways) that are modulated as a result of PDF receptor activation. Because circadian signaling occurs on such a long time base (many hours), we hypothesize that PDF signaling may include transcriptional regulation. We have identified several genes whose expression levels are pdf-dependent and which cycle on a daily basis. We propose now to demonstrate their role as PDF targets for the mediation of circadian signaling. Finally, we have performed an extensive genome-wide microarray screen for circadian gene expression in the Drosophila head. We have identified 22 circadian genes, of which 15 are novel in the context of circadian biology. We hypothesize that most of these circadian genes participate in circadian output mechanisms, and propose experiments to address this possibility