Secreted astrocyte proteins regulating rhythmic behavior

摘要 在哺乳动物和昆虫中，大脑星形胶质细胞与神经元共同调节睡眠和昼夜节律。 行为(在[1-3]中回顾)。为了识别调节节律行为的果蝇星形胶质细胞基因，我们 进行了星形胶质细胞特异性转录组范围的表达分析，以定义受 昼夜节律的方式。我们发现&gt；250星形胶质细胞的mRNAs在核糖体结合中表现出昼夜节律变化， 建议对这些成绩单进行有节奏的翻译(You等人提交)。许多这样的“骑行”基因 编码博曼宁(Bom)、图兰朵(Tot)的小分泌星形胶质细胞蛋白(细胞因子样或抗菌蛋白) 或生长阻滞肽(GBP)家族，已知是由机械或免疫应激诱导的。一个 这些基因中的许多受保守的Toll/免疫缺陷(IMD)信号通路和 我们的研究表明，这些通路的组成部分在成年星形胶质细胞中也大量循环。细胞因子样 分泌因子在病理性免疫反应中调节睡眠，但也有证据表明，它们 正常调节有节奏的行为。鉴于这些循环细胞因子和抗菌基因是应激- 诱导后，我们假设它们的功能是促进睡眠，以应对与 保持清醒。事实上，我们未发表的结果表明，星形胶质细胞缺乏BomBc2-一种Toll调节的， 一种编码细胞因子样蛋白的循环基因会导致睡眠时间和睡眠质量的下降。相关的 研究表明，小鼠Toll-4或Toll-2受体的缺陷分别改变了 对睡眠缺失的反应[4]或小鼠昼夜节律活动的巩固[5]。本申请提出了2个相关的 旨在验证有节奏地激活星形胶质细胞信号通路和分泌蛋白的假说 有助于调节昼夜节律行为或睡眠可塑性。目标1将定义成人的细胞类型 苍蝇大脑需要循环分泌的蛋白质或Toll/IMD信号成分来实现正常的节律行为 (昼夜节律和睡眠)由于这些分泌的蛋白质是由机械或免疫压力诱导的，这一目标将 也测试一下新的假设，即它们是由睡眠剥夺以Toll/IMD调节的方式诱导的，以 在睡眠动态平衡中起作用。目标2将确定分泌蛋白的体内表达模式 以及由循环mRNA编码的Toll/IMD信令组件。它还将询问它们是否循环进入 成虫大脑中特定细胞类型的丰度。这一目标还将使用CRISPR方法来确定 神经元和神经胶质生物钟对星形胶质细胞蛋白循环的调节作用(S)。 总之，这些目标将确定昼夜行为或睡眠可塑性所需的星形胶质细胞因子。 并确定调节其循环表达的细胞内信号通路。

Summary In both mammals and insects, brain astrocytes cooperate with neurons to modulate sleep and circadian behavior (reviewed in [1-3]). To identify Drosophila astrocyte genes that modulate rhythmic behaviors, we conducted an astrocyte-specific transcriptome-wide expression analysis to define mRNAs that are regulated in a circadian manner. We show that >250 astrocyte mRNAs exhibit circadian changes in ribosome association, suggesting rhythmic translation of these transcripts (You et al., submitted). A number of these “cycling” genes encode small secreted astrocyte proteins (cytokine-like or antibacterial) of the Bomanin (Bom), Turandot (Tot) or Growth Blocking Peptide (Gbp) families that are known to be induced by mechanical or immune stress. A number of these genes are regulated by the conserved Toll/Immune Deficiency (IMD) signaling pathways and our studies show that components of these pathways also cycle in abundance in adult astrocytes. Cytokine-like secreted factors modulate sleep during pathological immune responses, but there is also evidence that they normally regulate rhythmic behavior. Given that these cycling cytokine and antibacterial genes are stress- induced, we hypothesize that they function to promote sleep in response to the stress associated with wakefulness. Indeed, our unpublished results show that an astrocyte deficit for BomBc2 – a Toll-regulated, cycling gene that encodes a cytokine-like protein – results in decreased sleep amount and quality. Of related significance, it has been shown that deficits for the mouse Toll-4 or Toll-2 receptors, respectively, alter the response to sleep loss [4] or consolidation of mouse circadian activity [5]. This application proposes 2 related aims to test the hypothesis that rhythmically activated astrocyte signaling pathways and secreted proteins contribute to the regulation of circadian behavior or sleep plasticity. Aim 1 will define the cell types of the adult fly brain that require cycling secreted proteins or Toll/IMD signaling components for normal rhythmic behaviors (circadian and sleep). As these secreted proteins are induced by mechanical or immune stress, this aim will also test the novel hypothesis that they are induced by sleep deprivation, in a Toll/IMD-regulated manner, to function in sleep homeostasis. Aim 2 will determine the in vivo expression patterns for the secreted proteins and Toll/IMD signaling components encoded by cycling mRNAs. It will also ask whether they cycle in abundance in specific cell types of the adult fly brain. This aim will also employ CRISPR methods to determine the contribution(s) of neuronal and glial circadian clocks to the regulation of cycling astrocyte proteins. Altogether, these aims will identify astrocyte factors that are required for circadian behavior or sleep plasticity and define intracellular signaling pathways regulating their cycling expression.