Regulation of cilia by ceramide

纤毛是细胞膜的突起，具有感觉(初级纤毛)或运动(运动纤毛)功能。在……里面 星形胶质细胞和室管膜细胞、初级纤毛和运动纤毛调节细胞分裂和迁移，并推动 脑脊液(CSF)。睫状体功能障碍导致星形细胞过度生长(星形胶质细胞增生症)或 室管膜细胞功能障碍和脑积水。纤毛在细胞信号和运动中起着至关重要的作用。 纤毛数量、长度和货物蛋白的纤毛或鞭毛内运输(IFT)是动态的 受监管的。理解这一规则的一个关键障碍是缺乏关于动态激活的知识 纤毛发生和纤毛功能的影响因素。虽然纤毛是膜结构，但到目前为止，研究主要集中在 关于蛋白质在纤毛调节中的作用，对脂类在这一过程中的作用知之甚少。我们的 研究目标是确定膜脂和蛋白质如何在纤毛的调节中相互作用以及如何。 脂类代谢的调节可以用来支持星形胶质细胞和室管膜细胞的纤毛功能。 我们的中心假设是鞘磷脂神经酰胺调节纤毛长度和IFT，这对 纤毛在星形胶质细胞和室管膜细胞中的功能。我们的目标是1)测试纤毛是否受 神经酰胺相关的蛋白质复合体；2)通过使用一种新的下拉技术来定义这些复合体 富含神经酰胺和纤毛的膜小泡和双功能神经酰胺的共价交联 类似于其相互作用蛋白以确定神经酰胺结合区域；3)测试纤毛中受体的诱导 4)检测星形胶质细胞的激活和室管膜细胞的功能受 神经酰胺的体外和体内实验。我们的预期结果包括：1)确定促进 纤毛发生和支持纤毛功能，以及如何调节纤毛神经酰胺的生成；2)定义 神经酰胺与非典型蛋白激酶C、GSK-3β和 HDAC6；3)定义囊泡运输途径中的sMase激活及其对神经酰胺流向血管的作用 纤毛；4)识别与神经酰胺相关蛋白质和蛋白结构域；5)确定运输 纤毛中信号蛋白的激活，特别是sonic hedgehog途径的激活，受 神经酰胺；以及6)确定神经酰胺调节星形胶质细胞激活和室管膜细胞的机制。 驱动脑脊液流动。这个项目的影响是在BASIC中定义一个基本的和新颖的机制 神经科学和膜生物学，这对我们理解细胞的调节具有广泛的意义。 纤毛通过脂-蛋白相互作用以及这种调节对星形胶质细胞和 脑室管膜细胞在脑发育和衰老过程中的作用。目标1将检验神经酰胺稳定的假说 星形胶质细胞和室管膜细胞中的纤毛。目的2将验证神经酰胺调节IFT和 纤毛中的受体激活。目的3将验证纤毛神经酰胺调节星形胶质细胞和 室管膜细胞功能。

Cilia are protrusions of the cell membrane with sensory (primary cilia) or motor (motile cilia) function. In astrocytes and ependymal cells, primary and motile cilia regulate cell division and migration, and propel cerebrospinal fluid (CSF), respectively. Ciliary dysfunction leads to astrocytic overgrowth (astrogliosis) or ependymal cell malfunction and hydrocephalus. It is vital for the function of cilia in cell signaling and motility that cilium number, length, and intraciliary or intraflagellar transport (IFT) of cargo proteins are dynamically regulated. A critical barrier in understanding this regulation is the lack of knowledge on dynamically activated factors in ciliogenesis and cilium function. Although cilia are membrane structures, research so far has focused on the role of proteins in the regulation of cilia, and little is known about the role of lipids in this process. Our research goals are to determine how membrane lipids and proteins interact in the regulation of cilia and how modulation of lipid metabolism can be utilized to support the function of cilia in astrocytes and ependymal cells. Our central hypothesis is that the sphingolipid ceramide regulates cilium length and IFT, which is critical for the function of cilia in astrocytes and ependymal cells. Our objectives are to 1) test that cilia are regulated by ceramide-associated protein complexes; 2) define these complexes by using a novel technique to pull down ceramide enriched- and cilium-derived membrane vesicles and covalently crosslink a bifunctional ceramide analog to its interacting proteins to identify ceramide binding domains; 3) test that induction of receptors in cilia is regulated by ceramide; and 4) test that astroglial activation and ependymal cell function is regulated by ceramide in vitro and in vivo. Our expected outcomes include 1) determining ceramide species that promote ciliogenesis and support cilium function, and how the generation of ciliogenic ceramide is regulated; 2) defining a mechanism of cilium extension and IFT regulation by interaction of ceramide with atypical PKC, GSK-3β, and HDAC6; 3) defining SMase activation in vesicle transport pathways and their function for ceramide flux to the cilium; 4) identifying proteins and protein domains that associate with ceramide; 5) determining that transport and activation of signaling proteins in cilia, in particular of the sonic hedgehog pathway, are regulated by ceramide; and 6) defining a mechanism by which ceramide regulates astrocyte activation and ependymal cell- driven CSF flow. The impact of this project is on defining a fundamental and novel mechanism in basic neuroscience and membrane biology, which has broad implications for our understanding of the regulation of cilia by lipid-protein interaction and the importance of this regulation for the function of astrocytes and ependymal cells during brain development and aging. Aim 1 will test the hypothesis that ceramide stabilizes cilia in astrocytes and ependymal cells. Aim 2 will test the hypothesis that ceramide regulates IFT and receptor activation in cilia. Aim 3 will test the hypothesis that ciliogenic ceramide regulates astrocyte and ependymal cell function.