Function of SLEEPLESS in sleep regulation

描述（由申请人提供）：睡眠调节中的失眠功能概述睡眠具有基本的生物学功能，并且从苍蝇到人类都是保守的。这项研究的总体目标是阐明睡眠调节的分子遗传基础。通过对睡眠时间短的突变果蝇进行无偏的正向遗传筛选，我们发现了一个新的基因，sss，它是正常睡眠和剥夺后恢复睡眠所必需的。SSS编码在细胞表面表达的小的脑富集蛋白。以前被证明对睡眠调节很重要的Shaker（Sh）钾通道的表达在sss突变体中显著减少。要了解SSS如何调节睡眠，需要解决几个关键问题。Sh是否介导sss对睡眠的影响？SSS是通过形成复合物直接作用于Sh，还是通过信号通路间接作用于Sh？SSS是自主地还是非自主地作用于Sh？昼夜节律和稳态过程影响SSS和Sh表达吗？为了解决这些问题，我们提出了以下具体目标：（1）测试SSS是否通过作用于Sh来促进睡眠，以及它们是否在物理上相互作用。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。我们还将研究是否Sh的过度表达可以拯救sss突变体的短睡眠表型。此外，我们将测试SSS和Sh是否物理相互作用。(2)识别sss的作用轨迹。sss对睡眠的基础和反弹都有深远的影响，sss的鉴定使我们有机会探索稳态睡眠调节的细胞基质。我们将研究SSS蛋白的表达模式，并使用组织特异性驱动程序和sss转基因的集合进行拯救实验。将比较正常和剥夺条件下睡眠的细胞基质。(3)确定SSS是否自主地作用于Sh小区。为了了解SSS如何作用于Sh来调节睡眠，重要的是要确定Sh和SSS的作用位点。我们将使用组织特异性拯救和靶向RNAi敲低实验来确定Sh表达模式和组织需求。此外，我们将测试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。公共卫生相关性：缺乏高质量的睡眠是一个常见的问题，可能导致事故和生产力下降，医疗保健专业人员和公众越来越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会为我们提供关于我们如何以及为什么睡觉的重要线索，并可能最终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.