Genetic Dissection of Signaling and Cilia

项目概要/摘要 纤毛引起了科学家们的极大兴趣，因为科学家们认识到它们是一种基本的细胞器 需要信号。解开调节Sonic hedgehog（Shh）信号传导的特定机制， 纤毛是困难的，因为许多破坏纤毛发生的突变体也影响Hh信号传导。我们早就 我们的研究集中在一个小的睫状GTlymphoma，ARL 13 B，我们假设它整合了纤毛发生的调节， 通过不同效应子及其下游通路的Hh信号传导。作为一个GTdR，单碱基对 预测ARL 13 B的GT3结构域内的突变破坏个体效应子途径。ARL13B 纤毛非常丰富。通过工程改造仅表达不定位于ARL 13 B的ARL 13 B变体的小鼠， 为了从基因上将ARL 13 B在纤毛发生中的作用与其在信号传导中的作用分开，我们在基因上将ARL 13 B在纤毛发生中的作用与其在信号传导中的作用分开。我们专注于 哺乳动物神经发育，并通过正向遗传筛选确定小鼠突变体在几个 ARL 13 B相关蛋白此外，GTP酶的ARL家族中的其他蛋白质涉及纤毛 以及通过似乎是ARL 13 B相关机制的信号传导。在未来五年，我们建议 使用小鼠突变体来确定这些参与者在体内和特定细胞中的调控关系， 类型这些实验将揭示ARL 13 B在纤毛发生、蛋白质向纤毛/在纤毛内的运输以及 嘘信号转导在前所未有的分辨率。因此，我们的建议将产生一种分子遗传学， 工具包，该领域将准备区分纤毛的调节Hh信号。这是 重要的是我们的纤毛，纤毛发生和纤毛结构的基本理解，以及我们的基本 对Hh通路的理解。

Project Summary/Abstract Cilia sparked phenomenal interest as scientists recognized them as a fundamental cellular organelle required for signaling. Untangling the specific mechanisms that regulate Sonic hedgehog (Shh) signaling within the cilium is difficult since so many mutants that disrupt ciliogenesis also affect Hh signaling. We have long focused on a small ciliary GTPase, ARL13B, that we hypothesize integrates the regulation of ciliogenesis and Hh signaling through distinct effectors and their downstream pathways. As a GTPase, single basepair mutations within the GTPase domain of ARL13B are predicted to disrupt individual effector pathways. ARL13B is highly enriched in cilia. By engineering mouse expressing only an ARL13B variant that does not localize to cilia, we genetically uncoupled the role of ARL13B in ciliogenesis from its role in signaling. We focus on mammalian neural development and through forward genetic screens identified mouse mutants in several proteins related to ARL13B. Additionally, other proteins in the ARL family of GTPases are implicated in cilia and signaling through what appear to be ARL13B related mechanisms. In the next five years, we propose using mouse mutants to define the regulatory relationships among these players in vivo and in specific cell types. These experiments will unravel ARL13B function in ciliogenesis, traffic of proteins to/within cilia, and Shh signal transduction at unprecedented resolution. Thus, our proposal will generate a molecular genetic toolkit from which the field will be poised to distinguish the regulation of cilia from that of Hh signaling. This is important to our fundamental understanding of cilia, ciliogenesis and cilia structure, as well as our basic comprehension of the Hh pathway.