权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Control of epithelial morphology and bioenergetics by Toll receptors during dynamic tissue remodeling

动态组织重塑过程中 Toll 受体对上皮形态和生物能的控制

基本信息

批准号：
10574572
负责人：
Adam Christopher Pare
金额：
$ 20.63万
依托单位：
UNIVERSITY OF ARKANSAS AT FAYETTEVILLE
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-04-01 至 2023-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10574572
关键词：
Adhesions Affect Antibodies Architecture Arkansas Attention Bioenergetics Biological Biomechanics Birth CRISPR/Cas technology Cell Adhesion Inhibition Cell Polarity Cell Shape Cell-Cell Adhesion Cells Cellular Morphology Clustered Regularly Interspaced Short Palindromic Repeats Complex Congenital Abnormality Consumption Cytoskeleton Data Analyses Data Science Defect Development Disease Drosophila genus Embryo Epithelial Cells Epithelium Family Fluorescence G Protein-Coupled Receptor Signaling G-Protein-Coupled Receptors GPCR Signaling Pathway Genetic Genus Hippocampus Glycolysis Goals Human Human Pathology Image Immune signaling Individual Intercalated Cell Link Measures Mentors Metabolic Metabolism Mitochondria Molecular Molecular Probes Morphogenesis Morphology NADH Natural Immunity Nature Neural Tube Defects Oxidation-Reduction Oxidative Phosphorylation Oxygen Consumption Pathway interactions Pattern Play Positioning Attribute Process Receptor Signaling Research Rho-associated kinase Role Shapes Signal Pathway Signal Transduction Structure Study models System Techniques Tertiary Protein Structure Testing Tissues Visualization Work cell behavior cofactor convergent extension data integration epithelial wound experimental study functional loss gain of function in vivo evaluation insight intercalation loss of function member multiphoton microscopy mutant novel overexpression receptor receptor expression receptor function repaired rho wound closure

项目摘要

Project Summary – Project Leader Adam Paré Epithelial cells undergo significant changes in shape and relative position during development to build proper tissue architecture. Errors in epithelial remodeling directly contribute to some of the most common types of congenital abnormalities––neural tube defects––which affect approximately 1 in 2000 births. However, the upstream signals that control epithelial morphology remain poorly understood. A paradigm for studying epithelial remodeling is cell intercalation in the Drosophila neurectoderm, and it was shown that three members of the highly conserved Toll receptor family are expressed in overlapping striped patterns to organize rapid cell rearrangements in this tissue. Toll receptors are widely expressed throughout human epithelia, and they have been extensively studied in the context of innate immune signaling. However, the control of cell morphology by Toll receptors has received very little attention. The focus of this proposal is to understand how non-uniform Toll receptor expression affects cortical tension, cell-cell adhesion, and mitochondrial dynamics to control cell shape and behavior during epithelial remodeling. We will use newly developed CRISPR/Cas9-derived genetic backgrounds and antibodies to characterize how Toll receptors control cell polarity to trigger intercalation; we will apply non-destructive techniques to characterize the bioenergetics of epithelial reorganization in intact living embryos; and we will investigate unaddressed links between Toll receptor, Rho, and G protein-coupled receptor signaling. Our first hypothesis is that neighboring cells sense differences in the expression of individual Toll receptor types to increase cortical tension and decrease cell-cell adhesion. We have developed a genetic system for expressing individual receptors in a single stripe that we will use to systematically characterize and compare the effects of each Toll receptor type on cell morphology and to identify the protein domains necessary for modulating cell shape. Our second hypothesis is that rapid cellular rearrangements during neurectoderm elongation require specific changes in mitochondrial architecture to fuel cytoskeletal and junctional reorganization. To test this, we will use the Agilent Seahorse flux analyzer to measure oxygen consumption rates and multiphoton microscopy to visualize cellular redox state in live embryos during epithelial remodeling, and then use gain- and loss-of-function techniques to determine what role mitochondrial fusion and fission play in epithelial reorganization. Our third hypothesis is that Toll receptor and GPCR signaling converge to activate Rho Kinase to trigger cell intercalation specifically in the neurectoderm. We will use gain- and loss-of-functional analyses to determine how these two signaling pathways intersect to control cortical tension, cell-cell adhesion, mitochondrial dynamics during epithelial remodeling. Successful completion of these experiments will give us an understanding of how Toll receptors function at a molecular level to control cell morphology and bioenergetics during dynamic tissue remodeling, which could shed light on the cell biological underpinning of neural tube defects and other epithelia-based diseases.

项目总结-项目负责人Adam Paré 在发育过程中，上皮细胞在形状和相对位置上发生了显著变化，以建立适当的组织结构上皮重塑中的错误直接导致一些最常见的类型，先天性畸形-神经管缺陷-大约每2000名新生儿中就有一人受到影响。但控制上皮形态的上游信号仍然知之甚少。研究上皮细胞的范例重塑是果蝇神经外胚层中的细胞嵌入，并且已经表明，高度保守Toll受体家族以重叠的条纹模式表达以组织快速细胞在这个组织中重新排列。Toll受体在整个人类上皮细胞中广泛表达，并且它们具有在先天免疫信号传导的背景下被广泛研究。然而，细胞形态的控制， Toll受体很少受到关注。本建议的重点是了解如何不统一收费受体表达影响皮质张力、细胞间粘附和线粒体动力学以控制细胞形状和行为的影响。我们将使用新开发的CRISPR/Cas9衍生的基因工程技术，背景和抗体来表征Toll受体如何控制细胞极性以触发嵌入;我们将应用非破坏性技术来表征完整生命中上皮重组的生物能量学我们将研究Toll受体，Rho和G蛋白偶联受体之间未解决的联系发信号。我们的第一个假设是，邻近的细胞感觉到个体Toll表达的差异，受体类型增加皮质张力和减少细胞-细胞粘附。我们开发了一个基因系统在一条带中表达单个受体，我们将用它来系统地表征和比较每种Toll受体类型对细胞形态的影响，并确定所需的蛋白质结构域，调节细胞形状。我们的第二个假设是神经外胚层发育过程中的快速细胞重排延伸需要线粒体结构的特定变化，以促进细胞骨架和连接重组。为了测试这一点，我们将使用Agilent Seahorse通量分析仪来测量氧消耗率和多光子显微镜观察上皮重塑期间活胚胎中的细胞氧化还原状态，然后使用获得和丧失功能的技术来确定线粒体融合和裂变在上皮重组我们的第三个假设是Toll受体和GPCR信号传导会聚以激活Rho 在神经外胚层中特异性触发细胞嵌入的激酶。我们将使用增益和损失函数分析以确定这两种信号通路如何交叉以控制皮质张力，细胞-细胞粘附，上皮重塑过程中的线粒体动力学成功完成这些实验将给我们了解Toll受体如何在分子水平上控制细胞形态和生物能量学在动态组织重塑过程中，这可能揭示神经管的细胞生物学基础缺陷和其他基于上皮的疾病。