Structural studies of tight junction proteins

紧密连接蛋白的结构研究

基本信息

批准号：
8909863
负责人：
John M Flanagan
金额：
$ 4.79万
依托单位：
PENNSYLVANIA STATE UNIV HERSHEY MED CTR
依托单位国家：
美国
项目类别：
财政年份：
2013
资助国家：
美国
起止时间：
2013-01-01 至 2016-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8909863
关键词：
Alzheimer&apos s Disease Area Asthma Binding Biochemical Biological Biological Assay Biology Blood Blood-Retinal Barrier Brain Brain Neoplasms Cell physiology Cells Cellular Structures Coiled-Coil Domain Collaborations Complex Cytoplasmic Tail Data Disease Drug Delivery Systems Endothelial Cells Epithelial Epithelial Cells Extracellular Fluid Family Functional disorder Head Health Herpes zoster disease Human Hybrids Individual Infection Ions Irritable Bowel Syndrome Kidney Diseases Knowledge Lead Length Lung Malignant Neoplasms Medicine Membrane Membrane Proteins Methods Michigan Modeling Molecular N-terminal Pennsylvania Permeability Pharmaceutical Preparations Phosphorylation Physiological Physiology Property Protein Binding Proteins Protocols documentation Publishing Regulation Regulatory Element Research Resolution Retina Retinal Diseases Role Serine Site Stroke Structure Surface Techniques Testing Therapeutic Therapeutic Agents Tight Junctions Tissues Tyrosine Universities Vascular Dementia Work X-Ray Crystallography apical membrane base basolateral membrane college disorder prevention fluid flow gastrointestinal epithelium human disease improved in vivo insight novel novel strategies novel therapeutics occludin pathogen public health relevance receptor research study response solute tool treatment strategy uptake

项目摘要

DESCRIPTION (provided by applicant): Tight junctions (TJs) are essential cellular structures that form the selective paracellular barriers in endothelial and epithelial cells, including the blood-brain, and blood-retinal barriers, lung and gut epithelium. The barrier properties of TJs regulate metabolite flux, fluid flow and drug uptake into these tissues. Dysfunction in TJ barrier properties is observed in a large number of human diseases including cancer (e.g. brain tumors), stroke, retinopathies, kidney disorders, irritable bowel syndrome, Alzheimer's disease and vascular dementia, and asthma. Moreover, TJ components are used as cellular receptors by a range of pathogens. Thus, an improved understanding of the basic structural and functional biology of TJs will have wide ranging impact for human health. Specifically, this information may lead to improved drug delivery methods, novel protocols for regulating TJ permeability in disease and prevention of some infections. Currently, little is known about the molecular basis for TJ barrier properties and how they are regulated. In this proposal, we describe a combined structural, biochemical and cellular and molecular approach to illuminate the function of occludin (Occ), a transmembrane component of TJs, which has been implicated in the regulation of TJ barrier properties. These studies build upon our previously published and unpublished data implicating Occ, and its binding partner zona occludens 1 (ZO-1) protein, in regulating barrier properties conferred by TJs. The planned studies are organized into three specific aims. In Aim1, experiments are described, using a cell biological approach to determine the molecular mechanism(s) for the effects of phosphorylation of Serine 471 in Occ (S471) on TJ cellular structure function. In Aim2, we describe experiments to determine the structural details of the ZO-1/Occ protein-binding core, which includes S471, and with their functional binding complexes In Aim 3, we extend these approaches to determine the contribution of S471 phosphorylation of Occ to its interaction with ZO-1 using full length Occ and dimeric ZO-1 constructs. Together, the results of these three aims will provide a structural and biochemical basis for understanding Occ function and starting point for developing novel strategies for modulating TJ barrier properties that target the ZO-1/Occ complex.

描述（由申请人提供）：紧密连接（TJS）是必需的细胞结构，在内皮和上皮细胞中形成了选择性细胞屏障，包括血脑和血脑屏障，以及血液 - 视网膜屏障，肺和肠道屏障，肺和肠道上皮。 TJ的屏障特性调节代谢物通量，流体流量和药物吸收到这些组织中。在包括癌症（例如脑肿瘤），中风，视网膜病，肾脏疾病，肠易激综合症，阿尔茨海默氏病和血管性痴呆和哮喘在内的大量人类疾病中观察到TJ屏障特性的功能障碍。此外，TJ成分被一系列病原体用作细胞受体。因此，对TJ的基本结构和功能生物学的了解将对人类健康产生广泛的影响。具体而言，这些信息可能会导致改善药物输送方法，这是调节疾病中TJ渗透性和预防某些感染的新方案。当前，关于TJ屏障特性的分子基础及其调节方式知之甚少。在此提案中，我们描述了一种结合结构，生化和细胞和分子方法，以阐明occludin（occ）的功能（occ），TJ的跨膜成分，与TJ屏障性质的调节有关。这些研究基于我们先前发表的未发表的数据，涉及OCC及其结合伴侣Zona coccludens 1（ZO-1）蛋白，以调节TJ赋予的屏障特性。计划的研究分为三个特定目标。在AIM1中，使用细胞生物学方法来描述实验，以确定OCC（S471）中丝氨酸471磷酸化对TJ细胞结构功能的影响。在AIM2中，我们描述了确定ZO-1/OCC蛋白结合核的结构细节（包括S471）的实验，并在AIM 3中及其功能结合复合物，我们扩展了这些方法，以确定OCC的S471磷酸化的贡献，使用全长OCC和Dimeric ZO-1构造与ZO-1的相互作用。总之，这三个目标的结果将为理解OCC功能的结构和生化基础提供了开发新型策略的起点，以调节针对ZO-1/OCC复合物的TJ屏障特性。