Interdisciplinary studies of the Drosophila Circadian Clock

我们一直在以果蝇为模型研究昼夜行为节律的分子控制 系统最初在果蝇中表征的基因的直系同源物，现在已经与控制节奏性 行为和生理学的脊椎动物，包括鱼，青蛙，老鼠和人类。在这里，我们提出三个 果蝇生物钟的跨学科研究。(1)我们将进行结构性合作 研究可以帮助我们确定具体的监管行动，以前认识到遗传和 在生物化学上，由PER蛋白的某些结构域执行。PER的结构是否表明 它与TIM和DBT相互作用？这些数据是否表明TIM如何抑制DBT引起的PER磷酸化？ PER N端附近的LOV结构域是否具有黄素结合结构域？(2)我们将 产生新的突变，用于通过条件蛋白剪接分析重要的时钟基因。我们的研究 GSK-3、dbt、vri和Pdp-1的超型和亚型突变表明，每一种都是 果蝇时钟然而，由于这些基因的无效突变是致命的，因此难以 决定它们对时钟的全部影响。我们正在开发一种化学遗传学， 可扩散的分子将可逆地控制每种重要蛋白质在活蝇中的存在。(3)我们将生产 新的微阵列和统计方法，以澄清节奏基因组活动的特点，并确定是否 苍蝇使用单一的分子机制来产生所有的昼夜节律。开始将时钟链接到 特定的行为和生理输出，我们将研究一组新的基因，这些基因由一种 光和生物钟的配对作用。

We have been studying the molecular control of circadian behavioral rhythms using Drosophila as a model system. Orthologs of genes initially characterized in the fly,have now been linked to the control of rhythmic behavior and physiology in vertebrates, including fish, frogs, mice and humans. Here we propose three classes of interdisciplinary investigations of the Drosophila clock. (1) We will conduct collaborative structural studies that can help us determine how specific regulatory actions, previously recognized genetically and biochemically, are performed by certain domains of the PER protein. Does the structure of PER indicate how it interacts with TIM and DBT? Do such data suggest how TIM suppresses PER's phosphorylation by DBT? Does a putative LOV domain near the N-terminus of PER possess a flavin binding domain? (2) We will generate new mutations for the analysis of vital clock genes by Conditional Protein Splicing. Our studies of hyper- and hypo-morphic mutations of GSK-3, dbt, vri, and Pdp-1 indicate that each is a key component of the Drosophila clock. However, because null mutations of these genes are lethal, it has been difficult to determine their full effects on the clock. We are developing a form of chemical genetics in which small diffusible molecules will reversibly control the presence of each vital protein in living flies. (3) We will produce new microarray and statistical approaches to clarify features of rhythmic genome activity and to determine if flies use a single molecular mechanism to generate all circadian rhythms. To begin to link the clock to specific behavioral and physiological outputs, we will investigate a novel set of genes that are regulated by a paired action of light and the circadian clock.