The morphogenesis of actin-based structures in mucosal epithelia

粘膜上皮中肌动蛋白结构的形态发生

基本信息

批准号：
8681087
负责人：
Alvaro Sagasti
金额：
$ 22.32万
依托单位：
UNIVERSITY OF CALIFORNIA LOS ANGELES
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-04-01 至 2016-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8681087
关键词：
1-Phosphatidylinositol 4-Kinase Actin-Binding Protein Actins Affect Age Animals Apical Area Basal Cell Biological Models Brush Border Cell Shape Cell surface Cells Characteristics Cornea Cytoskeleton Defect Development Disease Epithelial Epithelial Cells Epithelium Esophagus Eye Eye diseases Film Fishes Gene Expression Gene Expression Profile Genetic Glycocalyx Glycoproteins Goals Height Human Hydration status Hydrophobic Surfaces Image Infection Intestines Label Larva Lead Length Life Lubrication Membrane Microfilaments Modeling Molecular Morphogenesis Mucins Mucous body substance Normal Cell Oral cavity PTEN gene Pain Pathology Pattern Periderm Pharyngeal structure Phosphatidylinositol 4,5-Diphosphate Phosphatidylinositols Phosphoric Monoester Hydrolases Play Process Production Property Proteins Recruitment Activity Relative (related person)Reporter Role Shapes Skin Staging Structure Study models Surface System Tertiary Protein Structure Testing Tissues Transgenic Organisms Width Xerostomia Zebrafish apical membrane base cellular microvillus conjunctiva eye dryness gene discovery genetic regulatory protein in vivo insight molecular imaging phosphatidylinositol 3,4,5-triphosphate phosphatidylinositol phosphate, PtdIns(4,5)P2 public health relevance research study time use tool transcriptome sequencing

项目摘要

DESCRIPTION (provided by applicant): Mucosal epithelia form the external interface of many sensitive tissues. The outer surface of cells in these epithelia displays a glycoprotein calyx and adsorbs mucins to create a mucus layer that protects those tissues from abrasion and maintains their hydration. In the cornea, for example, the mucus layer retains the tear film that keeps our eyes wet. Defects in the mucosal layer of the human cornea cause dry eye diseases (DED). These conditions are common, painful, and progress with age. DED is often accompanied by dry mouth, suggesting that its underlying causes affect properties common to mucosal epithelia. One such property is the presence of elaborate actin-based structures on the surface of these epithelial cells, known as microplicae and microridges. These structures have been little studied, but likely make a vital contribution to the functional properties of mucosal epithelia by increasin the surface area of the glycocalyx, thus maximizing their ability to hydrate tissues. Most studies of DED pathology have focused on tear production, but given the critical role of mucosal epithelia in maintaining the mucus layer and tear film, it is likely that defects in epithelial morphogenesis also contribute to these conditions. One of the main obstacles to studying the morphogenesis of microplicae and microridges has been the lack of an accessible model system. We have developed the larval zebrafish skin as a model for studying mucosal epithelial development. The entire surface of zebrafish larvae is wrapped in a single-layered mucosal epithelium known as the periderm. The apical surface of periderm cells is covered my microplicae and microridges that remarkably resemble structures on the surface of the human cornea. These cells are exceptionally accessible to transgenic labeling and confocal imaging, making it possible to visualize the formation of ridges in living animals. Moreover, the amenability of the zebrafish system to sophisticated genetic and transgenic manipulations will make it possible to uncover the underlying molecular mechanisms of microridge formation. In this proposal we combine descriptive, hypothesis-driven, and discovery-based approaches to dissect the process of ridge morphogenesis. Specifically, in Aim 1 we will use live imaging to describe the initial formation of microridges and their re-organization during cellular contraction In Aim 2 we will use molecular and imaging approaches to test the hypothesis that phosphoinositide microdomains orchestrate the formation of microridges. Finally, in Aim 3 we will use RNA-Seq to identify BAR domain proteins and actin regulators enriched in periderm and test whether select candidate proteins play roles in microridge development. Together these studies will provide the first insights into ridge morphogenesis and establish the zebrafish model as a system for understanding not only ridge formation in normal cells, but also the causes of their pathology in diseases, such as DED.

描述（由申请人提供）：粘膜上皮形成许多敏感组织的外部界面。这些上皮细胞的外表面显示出糖蛋白花萼和吸附粘蛋白，以产生粘液层，以保护这些组织免受磨损并保持其水分。例如，在角膜中，粘液层保留了使我们的眼睛湿的泪膜。人角膜的粘膜层中的缺陷会导致干眼疾病（DED）。这些条件是普遍，痛苦的，并且随着年龄的增长而进展。 DED通常伴有口干，表明其根本原因会影响粘膜上皮的常见特性。这样的特性之一是在这些上皮细胞的表面（称为微虫和微型树木）的表面上存在精致的基于肌动蛋白的结构。这些结构的研究很少，但可能通过增加糖脂的表面积来对粘膜上皮的功能特性做出至关重要的贡献，从而最大程度地提高了它们的水合组织能力。大多数DED病理学的研究都集中在泪液产生上，但是鉴于粘膜上皮在维持粘液层和泪膜中的关键作用，因此上皮形态发生的缺陷也可能导致这些疾病。研究微虫和微型矿石形态发生的主要障碍之一是缺乏可访问的模型系统。我们已经开发了幼虫斑马鱼皮，作为研究粘膜上皮发育的模型。斑马鱼幼虫的整个表面都包裹在一个称为Periderm的单层粘膜上皮。我的微虫和微型树木的顶部表面覆盖了，它们非常类似于人角膜表面的结构。这些细胞在转基因的标记和共聚焦成像中非常可访问，从而使活体动物中脊形成形成成为可能。此外，斑马鱼系统对复杂的遗传和转基因操纵的不合适性将使您有可能发现Micridge形成的基本分子机制。在此提案中，我们结合了描述性，假设驱动和基于发现的方法，以剖析脊形态发生过程。具体而言，在目标1中，我们将使用实时成像来描述AIM 2中细胞收缩过程中微型树木的初始形成及其在细胞收缩期间的重组形成，我们将使用分子和成像方法来检验以下假设：磷酸固醇磷酸肌乙酰胺微构域策划了微泳道的形成。最后，在AIM 3中，我们将使用RNA-Seq识别富含Periderm的棒域蛋白和肌动蛋白调节剂，并测试选择的候选蛋白是否在Microde of tevelopt中起着作用。这些研究将共同提供对脊形态发生的第一个见解，并建立斑马鱼模型，作为一种系统，不仅了解正常细胞中的脊形成，还可以理解其在疾病中的病理学原因，例如DED。