Function of SLEEPLESS in sleep regulation

描述(申请人提供)：睡眠调节中的失眠功能摘要睡眠具有基本的生物学功能，从苍蝇到人类都是如此。这项拟议研究的总体目标是阐明睡眠调节的分子遗传学基础。通过对短眠突变果蝇的无偏正向遗传筛选，我们发现了一种新的基因--无眠(Sss)，它是正常睡眠和剥夺后恢复睡眠所必需的。SSS编码一种表达在细胞表面的小的、富含大脑的蛋白质。以前被证明对睡眠调节重要的振荡器(Sh)钾通道的表达在sss突变体中显著减少。要了解SSS是如何调节睡眠的，需要解决几个关键问题。Sh是否在sss对睡眠的影响中起中介作用？SSS是通过形成复合体直接作用于Sh，还是通过信号通路间接作用于Sh？SSS对Sh细胞的作用是自主的还是非细胞的？昼夜节律和动态平衡过程是否影响SSS和Sh的表达？为了解决这些问题，我们提出了以下具体目标：(1)检验SSS是否通过作用于Sh而促进睡眠，以及它们之间是否存在身体上的相互作用。为了证实sss和sh之间潜在的遗传相互作用，我们将寻找降低sss和sh活性对睡眠的协同作用。我们还将研究Sh的过度表达是否可以挽救sss突变体的短眠表型。此外，我们将测试Sss和Sh是否存在身体上的相互作用。(2)确定社会保障体系的作用轨迹。SSS的识别对基础睡眠和反弹睡眠都有深远的影响，使我们有机会探索稳态睡眠调节的细胞底物。我们将研究SSS蛋白的表达模式，并使用一系列组织特异性驱动因素和SSS转基因进行救援实验。我们将比较正常和剥夺条件下睡眠的细胞底物。(3)确定SSS是否自主作用于Sh细胞。为了了解SSS如何作用于Sh来调节睡眠，确定Sh和Sss的作用轨迹是很重要的。我们将通过组织特异性救援和靶向RNAi敲除实验来确定Sh的表达模式和组织需求。此外，我们还将测试SSS是否可以分泌，以及分泌的SSS是否可以挽救Sss突变体的睡眠表型。与公共健康相关：缺乏高质量的睡眠是一个常见的问题，可能会导致事故和生产力下降，医疗保健专业人员和公众越来越多地意识到睡眠是一个关键的健康问题。阐明睡眠调节的分子机制可能为我们提供关于睡眠方式和睡眠原因的重要线索，并最终可能导致睡眠障碍的新疗法。

DESCRIPTION (provided by applicant): Function of SLEEPLESS in Sleep Regulation Summary Sleep serves essential biological functions, and is conserved from flies to humans. The overall goal of the proposed research is to elucidate the molecular genetic basis of sleep regulation. Through an unbiased forward-genetic screen for short-sleeping mutant flies, we identified a novel gene, sleepless (sss), which is required for normal sleep as well as recovery sleep after deprivation. sss encodes a small, brain-enriched protein expressed on the cell surface. Expression of the Shaker (Sh) potassium channel, previously shown to be important for sleep regulation, is markedly reduced in sss mutants. Several key questions need to be addressed to understand how SSS regulate sleep. Does Sh mediate the effect of sss on sleep? Does SSS act directly on Sh by forming a complex or does it act indirectly via a signaling pathway? Does SSS act on Sh cell-autonomously or non-cell-autonomously? Do circadian and homeostatic processes affect SSS and Sh expression? To address these questions, we propose the following specific aims: (1) Test whether SSS promotes sleep by acting on Sh and whether they physically interact. To confirm a potential genetic interaction between sss and Sh, we will look for a synergistic effect on sleep of reducing both sss and Sh activity. We will also examine whether overexpression of Sh can rescue the short-sleeping phenotype of sss mutants. In addition, we will test whether SSS and Sh physically interact. (2) Identify the locus of action of sss. Identification of sss, which has profound effects on both baseline and rebound sleep, allows us an opportunity to explore the cellular substrates of homeostatic sleep regulation. We will examine the expression pattern of the SSS protein, and carry out rescue experiments using a collection of tissue-specific drivers and an sss transgene. Cellular substrates of sleep under normal and deprived conditions will be compared. (3) Determine if SSS acts on Sh cell-autonomously. To understand how SSS acts on Sh to regulate sleep, it is important to determine the locus of action of Sh as well as that of sss. We will determine the Sh expression pattern and tissue requirement using tissue-specific rescue and targeted RNAi knockdown experiments. In addition, we will test whether SSS can be secreted and whether secreted SSS can rescue the sleep phenotype of sss mutants. PUBLIC HEALTH RELEVANCE: Lack of good-quality sleep is a common problem that can lead to accidents and reduced productivity, and there is growing awareness among healthcare professionals and the public that sleep is a critical health issue. Elucidation of molecular mechanisms underlying sleep regulation may provide us with important clues as to how and why we sleep, and may ultimately lead to novel treatments for sleep disorders.