Mechanisms of basement membrane secretion and assembly

基底膜分泌和组装机制

基本信息

批准号：
10352423
负责人：
Sally Horne-Badovinac
金额：
$ 31.92万
依托单位：
UNIVERSITY OF CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-04-01 至 2023-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10352423
关键词：
Apical Architecture Basement membrane Binding Biochemical Biological Assay Biology Bulla Cell surface Cells Cytoplasm Data Defect Destinations Disease Drosophila genus Dystroglycan ECM receptor Ensure Epithelial Epithelial Cells Extracellular Matrix Genetic Goals Golgi Apparatus Gossypium Heart Homeostasis Image Kidney Diseases Kinesin Lateral Lead Location Logic Lung Mammary gland Mechanics Membrane Protein Traffic Membrane Proteins Molecular Monomeric GTP-Binding Proteins Morphogenesis Motion Motor Neoplasm Metastasis Organ Pathway interactions Play Proteins Research Role Salivary Glands Shapes Signal Transduction Site Skin Stroke Surface System Testing Three-Dimensional Imaging Tissue constructs Tissues Travel Tubular formation Variant Vesicle Vision Work biophysical techniques cell motility density egg imaging approach mechanical force membrane assembly migration network architecture novel programs recruit tool trafficking

项目摘要

PROJECT SUMMARY / ABSTRACT Basement membranes (BM) are specialized extracellular matrices found at the basal surface of all epithelial tissues. These sheet-like protein networks provide mechanical stability to the cells, promote cell-cell and cell- matrix signaling, and act as a physical barrier to metastasis. Moreover, defects in BM assembly lead to skin blistering, vision problems, nephropathy, and stroke. Some aspects of BM assembly are shared by all epithelia. For example, when new BM proteins are made by the epithelial cells, post-Golgi vesicles filled with BM proteins (BM vesicles) must be trafficked exclusively to basal cellular regions for secretion. Other aspects of BM assembly are tissue-specific. For example, local variations in BM composition and density direct the branching of the mammary gland, salivary gland and lung. However, despite their clear importance to epithelial biology, we know remarkably little about how BMs are built. The goals of this proposal are: (1) to identify the molecular logic underlying polarized BM secretion, and (2) to determine how the basic BM assembly program can be modified to create a specialized BM architecture for organ morphogenesis. To this end, we are studying the BM that surrounds the Drosophila egg chamber (the multi-cellular precursor to the egg). The epithelial cells that form the egg chamber’s outer layer (follicle cells) secrete their own BM that is easily visualized on the egg chamber’s surface. We and others previously used this system to identify two small GTPases, Rab10 and Rab8, that act as master regulators of polarized BM secretion. However, how these Rabs direct the transport of BM vesicles from their point of origin at the Golgi to the basal- most region of the cell where the BM needs to be assembled is unknown. We have identified two kinesins that, when depleted, cause BM proteins to be mis-secreted. In Aim 1, we will test the hypothesis that Rab10 and/or Rab8 recruit the kinesins to BM vesicles for directed transport. This work will identify fundamental mechanisms that determine where and how a BM is built and identify new guiding principles for polarized membrane traffic. The BM we study also has a specialized architecture that allows it to act as a “molecular corset” to elongate the egg chamber. The follicle cells collectively migrate along their BM, which remains stationary. We showed that this motion synergizes with new BM secretion to build an oriented array of fibrils into the planar BM that form the heart of the molecular corset. This work has now led us to hypothesize that there may be two BM secretion pathways, a Rab10-dependent pathway that builds the planar BM and a Rab8-dependent pathway that builds the fibrils. Aim 2 will test this hypothesis. Our supporting data further suggest that BM proteins that exit the cell via the fibril-forming pathway must be placed under tension by the migrating cells for linear fibrils to form. Aim 3 will test this hypothesis. By studying these two tissue-specific mechanisms for BM formation, we will identify general principles that cells can use to build a specialized BM architecture for organ morphogenesis and identify a new role for mechanical forces in BM assembly.

项目摘要 /摘要地下膜（BM）是在所有上皮的基础表面发现的专门细胞外基质组织。这些类似薄片的蛋白网络为细胞提供了机械稳定性，促进细胞细胞和细胞 - 矩阵信号传导，并充当转移的物理障碍。此外，BM组装中的缺陷导致皮肤起泡，视力问题，肾病和中风。 BM组装的某些方面由所有上皮共享。例如，当上皮细胞制造新的BM蛋白时，充满BM的高尔基蔬菜蛋白质（BM蔬菜）必须专门贩运到基本的细胞区域进行分泌。其他方面 BM组装是组织特异性的。例如，BM组成和密度的局部变化直接乳腺，唾液腺和肺的分支。然而，尽管它们对上皮很重要生物学，我们对BMS的建立方式知之甚少。该提案的目标是：（1）确定分子逻辑基础偏光BM分泌，（2）确定基本BM组装方式如何可以修改以创建用于器官形态发生的专门BM架构。为此，我们正在研究围绕果蝇卵室的BM（多细胞鸡蛋的前体）。形成卵室外层（卵泡细胞）的上皮细胞分泌拥有在鸡蛋室表面上很容易可视化的BM。我们和其他人以前使用此系统来识别两个小的GTPases，即Rab10和Rab8，它们充当了极化BM分泌的主要调节剂。但是，这些Rabs如何将BM蔬菜从高尔基的原产地转移到基本 BM需要组装的细胞的大多数区域尚不清楚。我们已经确定了两个动力素，它们，耗尽时，会导致BM蛋白被错误分泌。在AIM 1中，我们将测试Rab10和/或的假设 Rab8将Kines招募到BM蔬菜进行定向运输。这项工作将确定基本机制这决定了BM的何处以及如何确定两极分化膜流量的新指南原则。我们研究的BM还具有专门的体系结构，使其可以充当“分子紧身胸衣” 拉长卵室。植物细胞沿其BM统一迁移，该BM保持静止。我们表明该运动与新的BM分泌协同作用，以在平面中建造一系列的原纤维阵列构成分子紧身胸衣心脏的BM。这项工作现在使我们假设可能有两个 BM分泌途径，一种依赖于Rab10的途径，构建平面BM和Rab8依赖性建立原纤维的途径。 AIM 2将检验这一假设。我们的支持数据进一步表明BM 通过纤维形成途径退出细胞的蛋白质必须由迁移细胞置于张力下线性原纤维形成。 AIM 3将检验这一假设。通过研究BM的这两种组织特异性机制形成，我们将确定细胞可以用来构建器官的专业BM体系结构的一般原则形态发生并确定机械力在BM组装中的新作用。