The Role of Membrane Architecture in Primary Cilium Signaling

项目摘要 纤毛是普遍存在的以微管为基础的细胞器，参与感觉、发育 信号、液体流动和细胞运动的失调导致一系列人类疾病状态，包括 癌症和纤毛病。不活动的初级纤毛起信号天线的作用。它有一个明显的和 严格控制蛋白质和脂肪的组成，同时保持与质膜的毗连 手机。适当的纤毛功能取决于纤毛活动和纤毛活动的空间分隔 信令组件高度集中。虽然已经对蛋白质进行了广泛的表征 以及构成纤毛、纤毛膜和其底部的扩散屏障的脂类，我们知道的要少得多。 关于纤毛近端支持适当信号的膜区的组成和功能。 在许多细胞类型中，成熟的初级纤毛位于微米深的膜内陷中，称为 “纤毛袋”是一个重要的但未被充分描述的内吞活动和信号中心。这个 睫状囊的超微结构高度保守，不同于纤毛和胞浆。 膜。很难明确地将睫状囊定义为一个不同的间隔，并确定其 对纤毛信号和功能的充分贡献。这在一定程度上是由于成像的技术挑战 衍射有限的区域和缺乏工具来具体扰乱口袋的结构和组成。在这 提案I将通过一套相互关联的目标，利用创新技术来解决这一差距 包括基于邻近的标记和超分辨率显微镜以及功能信号分析，以： 1)确定睫状囊的分子组成；2)确定睫状囊的结构 建立和维持以支持信号，以及3)研究睫状囊在细胞外的作用 发信号。总之，我建议建立一张高分辨率的建筑和空间地图，并 哺乳动物睫状囊的时间动力学，并确定其在支持纤毛信号转导中的作用。 纤毛结构的缺陷会导致多种疾病状态下的信号异常。建议数 研究将扩大我们对睫状膜的结构如何支持适当的理解 初级纤毛的功能，并有可能为人类的分子基础提供新的见解 纤毛病和癌症等疾病。

Project Summary Cilia are ubiquitous and important microtubule-based organelles involved in sensation, developmental signaling, fluid flow and cell motility whose dysregulation leads to a range of human disease states including cancers and ciliopathies. The non-motile primary cilium functions as a signaling antenna. It has a distinct and tightly regulated protein and lipid composition while remaining contiguous with the plasma membrane of the cell. Proper cilium function depends on compartmentalization both for spatial separation of ciliary activities and high concentration of signaling components. While there has been extensive characterization of the proteins and lipids that compose the cilium, ciliary membrane, and the diffusion barrier at its base, much less is known about the composition and function of membrane regions proximal to the cilium that support proper signaling. In many cell types, the mature primary cilium sits in a microns-deep membrane invagination called the “ciliary pocket” that is an important but under-characterized hub of endocytic activity and signaling. The ultrastructure of the ciliary pocket is highly conserved and is distinct from both the ciliary and plasma membranes. It has been difficult to clearly define the ciliary pocket as a distinct compartment and determine its full contributions to ciliary signaling and function. This is in part due to the technical challenge of imaging this diffraction limited region and lack of tools to specifically perturb the pocket’s structure and composition. In this proposal I will address this gap through an interconnected set of aims, leveraging innovative technologies including proximity-based labeling and super-resolution microscopy as well as functional signaling assays to: 1) define the molecular composition of the ciliary pocket, 2) determine how ciliary pocket structure is established and maintained to support signaling, and 3) investigate extracellular roles of the ciliary pocket in signaling. In summary, I propose to establish a high-resolution map of the architecture and spatial and temporal dynamics of the mammalian ciliary pocket and define its role in supporting ciliary signal transduction. Defects in ciliary structure lead to aberrant signaling in a variety of disease states. The proposed research will expand our understanding of how the architecture of the periciliary membrane supports proper function of the primary cilium and has the potential to contribute new insights into the molecular basis of human diseases such as ciliopathies and cancer.