The Role of Membrane Architecture in Primary Cilium Signaling

膜结构在初级纤毛信号传导中的作用

• 批准号: 10592274
• 负责人: Alexandra Fitzgerald Long
• 金额: $ 7.43万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-05 至 2024-04-04
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10592274
• 关键词: Actins; Architecture; Biological Assay; Biotin; Cell membrane; Cells; Centrosome; Chemicals; Cilia; Complex; Cytoskeleton; Defect; Development; Diffusion; Disease; Endocytosis; Epithelium; Esthesia; Exclusion; Functional disorder; Genetic; Image; Immunofluorescence Immunologic; Label; Lead; Ligands; Lighting; Lipids; Malignant Neoplasms; Maps; Mass Spectrum Analysis; Membrane; Microscopy; Microtubules; Modeling; Molecular; Molecular Profiling; Organelles; Play; Proteins; Proteome; Regulation; Research; Resolution; Role; SHH gene; Shapes; Signal Pathway; Signal Transduction; Structure; Techniques; Testing; Tissues; Variant; Work; base; cell motility; cell type; ciliopathy; experience; extracellular; fluid flow; human disease; improved; in vivo; inhibitor; innovative technologies; insight; neglect; protein structure; smoothened signaling pathway; superresolution microscopy; tool; trafficking

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.

项目摘要 纤毛是一种普遍存在的重要的微管细胞器，参与感觉、发育和免疫调节。 信号传导、流体流动和细胞运动，其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着信号天线的作用。它有一个独特的， 严格调节蛋白质和脂质组成，同时保持与质膜的连续性。 cell.适当的纤毛功能取决于纤毛活动的空间分离和 高浓度的信号成分。虽然已经对蛋白质进行了广泛的表征， 以及构成纤毛、睫状体膜及其底部扩散屏障的脂质， 关于纤毛附近支持适当信号传导的膜区域的组成和功能。 在许多类型的细胞中，成熟的初级纤毛位于一个微米深的膜内陷，称为 在一些实施方案中，“纤毛袋”是重要的但未充分表征的内吞活性和信号传导的枢纽。的 睫状囊的超微结构是高度保守的，并且与睫状囊和血浆都不同 膜。很难清楚地将睫状囊定义为一个独特的隔室，并确定其 对纤毛信号传导和功能的全部贡献。这在一定程度上是由于成像的技术挑战， 衍射限制区域和缺乏工具来专门扰动口袋的结构和组成。在这 我将通过一系列相互关联的目标，利用创新技术， 包括基于邻近的标记和超分辨率显微镜以及功能性信号传导分析，以： 1)定义睫状囊的分子组成，2）确定睫状囊结构如何 建立并维持以支持信号传导，和3）研究睫状体袋在细胞外的作用， 发信号。总之，我建议建立一个高分辨率的建筑和空间地图， 哺乳动物纤毛袋的时间动力学，并确定其在支持纤毛信号转导的作用。 睫状结构的缺陷导致各种疾病状态中的异常信号传导。拟议 研究将扩大我们对睫状体周膜结构如何支持适当的 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。 疾病如纤毛病和癌症。