Cytoskeletal control of microridge morphogenesis on mucosal epithelial cells of the zebrafish skin

斑马鱼皮肤粘膜上皮细胞微脊形态发生的细胞骨架控制

• 批准号: 9534703
• 负责人: Alvaro Sagasti
• 金额: $ 29.26万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-01 至 2021-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9534703
• 关键词: Actin-Binding Protein; Actins; Address; Adopted; Affect; Animals; Apical; Architecture; Bacterial Artificial Chromosomes; Cell Shape; Cell Surface Extensions; Cell membrane; Cell surface; Cells; Cellular Structures; Complement; Complex; Cornea; Cytokinesis; Cytoskeletal Proteins; Cytoskeleton; Data; Defect; Development; Disease; Elements; Epithelial; Epithelial Cells; Epithelium; Esophagus; Etiology; Expression Profiling; Eye diseases; F-Actin; Fingerprint; Fishes; Genetic Recombination; Glycoproteins; Goals; Grant; Hydration status; Image; Individual; Infection; Intermediate Filaments; Keratin; Larva; Lead; Length; Link; Lubrication; Membrane; Microfilaments; Modeling; Molecular; Molecular Conformation; Morphogenesis; Mucins; Mucous Membrane; Mucous body substance; Myosin ATPase; Myosin Type II; Oral cavity; Pattern; Periderm; Pharmacology; Pharyngeal structure; Play; Process; Property; Protein Family; Protein Isoforms; Proteins; Regulation; Reporter; Rho-associated kinase; Role; Shapes; Signal Transduction; Skin; Structure; Surface; Testing; Time; Tissues; United States National Institutes of Health; Xerostomia; Zebrafish; base; cellular microvillus; conjunctiva; drebrins; experimental study; eye dryness; ezrin; inhibitor/antagonist; insight; knock-down; non-muscle myosin; overexpression; protein function; rho GTP-Binding Proteins; transcriptome sequencing

PROJECT SUMMARY To adopt forms optimized for their functions, individual cells sometimes project remarkably elaborate membrane protrusions, and even arrange them in complex patterns on their surfaces. To create and support membrane structures, the underlying cortical cytoskeleton must be arranged in specific conformations. The particular complement of cytoskeletal associated proteins, and the local regulation of their activity, thus determines the shape and pattern of cell membrane protrusions. To understand how cytoskeletal proteins together create unique cellular structures, we will study the formation of stunning actin-based structures on the surface of zebrafish skin cells called microridges. Microridges (or similar structures called microplicae) are found on most mucosal epithelial cells, which not only form the outer layer of fish skin, but also many of our own tissues, including the cornea, mouth, and parts of our gut. The microridge-covered surfaces of these cells display a glycoprotein calyx and adsorb mucins, suggesting that the unique structure of microridges is optimized for mucus retention. Mucus protects vulnerable epithelial tissues from abrasion and drying out, so understanding how microridges form could provide insight into the etiology of diseases affecting mucosal tissues, such as dry eye and dry mouth conditions. This proposal builds on successful descriptive and discovery-based studies supported by an NIH R21 grant that led to the identification of several new proteins in microridges. The experiments proposed here investigate mechanisms by which these specific proteins contribute to microridge morphogenesis, and, from a broader perspective, how they function as an ensemble to create the unique shapes and properties of microridges. In Aim 1 we will test the hypothesis that two proteins, Ezrin and Drebrin-like, initiate the first step of microridge morphogenesis, the formation of microvilli-like microridge precursors. Aim 2 investigates the interactions between F-actin and intermediate filaments (IFs) in microridges by testing if F-actin patterning determines IF patterning, and by testing the hypothesis that two candidate proteins, Envoplakin and Periplakin, link these cytoskeletal elements together. Finally, in Aim 3, live imaging, pharmacology, and molecular approaches will be used to characterize how myosin-based contraction and Rho GTPase signaling contribute to microridge morphogenesis. Collectively these studies will provide mechanistic insights into microridge morphogenesis, illuminate how cytoskeletal proteins together create elaborate cellular structures, and potentially point to how defects in epithelial morphogenesis contribute to diseases afflicting mucosal epithelia.

项目概要 为了采用针对其功能优化的形式，单个细胞有时会表现得非常复杂 膜突起，甚至在其表面排列成复杂的图案。创建和支持 膜结构中，底层的皮质细胞骨架必须以特定的构象排列。这 细胞骨架相关蛋白的特定补体及其活性的局部调节，因此 决定细胞膜突起的形状和图案。 为了了解细胞骨架蛋白如何共同创造独特的细胞结构，我们将研究其形成 斑马鱼皮肤细胞表面令人惊叹的基于肌动蛋白的结构，称为微脊。微脊（或 大多数粘膜上皮细胞上都发现了类似的结构（称为微皱襞），这些细胞不仅形成外部 鱼皮层，还有我们自己的许多组织，包括角膜、口腔和肠道的一部分。这 这些细胞的微脊覆盖的表面显示出糖蛋白花萼并吸附粘蛋白，这表明 独特的微脊结构针对粘液保留进行了优化。粘液保护脆弱的上皮组织 由于磨损和干燥，因此了解微脊是如何形成的可以帮助深入了解微脊的病因学 影响粘膜组织的疾病，例如干眼症和口干症。 该提案建立在 NIH R21 拨款支持的成功的描述性和基于发现的研究的基础上 这导致微脊中几种新蛋白质的鉴定。这里提出的实验调查了 这些特定蛋白质促进微脊形态发生的机制，并且从更广泛的角度 从角度来看，它们如何作为一个整体发挥作用，创造微脊的独特形状和特性。在 目标 1 我们将测试以下假设：两种蛋白质（Ezrin 和 Drebrin 样）启动微脊的第一步 形态发生，微绒毛样微脊前体的形成。目标 2 研究相互作用 通过测试 F-肌动蛋白图案是否决定 IF，在微脊中区分 F-肌动蛋白和中间丝 (IF) 模式，并通过测试两个候选蛋白 Envoplakin 和 Periplakin 将这些蛋白联系起来的假设 细胞骨架元件在一起。最后，在目标 3 中，实时成像、药理学和分子方法将 用于表征基于肌球蛋白的收缩和 Rho GTPase 信号传导如何促进微脊 形态发生。总的来说，这些研究将为微脊形态发生提供机制见解， 阐明细胞骨架蛋白如何共同创造复杂的细胞结构，并可能指出如何 上皮形态发生的缺陷导致粘膜上皮疾病。