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The Molecular Biophysics and Tissue Biomechanics of Somite Morphogenesis

体节形态发生的分子生物物理学和组织生物力学

基本信息

批准号：
9896870
负责人：
SCOTT A HOLLEY
金额：
$ 38.36万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-04-13 至 2022-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9896870
关键词：
Adhesions Apical Automobile Driving Binding Proteins Biological Biological Assay Biology Biomechanics Biophysics Cadherins Cell Adhesion Cell Adhesion Molecules Cell Nucleus Cell Separation Cell Surface Proteins Cell model Cell-Cell Adhesion Cells Computer Models Computer Simulation Data Development Diffusion Disease Embryo Ensure Ephrins Epithelial Epithelial Cells Epithelium Exercise Extracellular Matrix Feedback Fibronectin Receptors Fibronectins Fluorescence Fluorescence Resonance Energy Transfer Genes Homeostasis ITGA5 gene Image Integrin alpha5beta1 Integrins Ligands Measures Mechanics Mediating Membrane Mesenchyme Mesoderm Modeling Molecular Molecular Conformation Morphogenesis Motion Movement Pathway interactions Protein Conformation Proteins Receptor Protein-Tyrosine Kinases Regulation Resources Role Side Signal Transduction Somites Spectrum Analysis Systems Analysis Techniques Testing Tissues Transgenic Organisms Vertebral column Zebrafish complement system epithelial to mesenchymal transition experimental study in silico in vivo kinematics laboratory experiment molecular dynamics molecular scale mutant protein protein interaction segregation spatiotemporal spine bone structure

项目摘要

Reciprocal regulation between the ECM and associated epithelial cells is integral to development, homeostasis and disease. Somites are segmental precursors of the vertebral column and musculature that form via a mesenchymal to epithelial transition. Somite morphogenesis is dependent upon a Fibronectin ECM, the Fibronectin receptor Integrin 51, the cell adhesion protein Cadherin 2 and bidirectional signaling via the receptor tyrosine kinase EphA4 and its membrane bound ligand Ephrin-B2a. These genes/pathways mediate cell-ECM adhesion, cell-cell adhesion and contact mediated cell repulsion, and our hypothesis is that the physical organizing activity of the somite boundary emerges via specific spatiotemporal intertwining of differential cell adhesion and ECM constrained cell repulsion. In Aim 1, fluorescence correlation spectroscopy (FCS) and fluorescence crosscorrelation spectroscopy (FCCS) will be used quantify protein diffusion and protein binding constants in vivo. These experiments will determine whether the segregation of these cell surface proteins occurs via diffusion and capture or active mobilization. Additionally, the roles of integrin 5, cadherin 2 and ephrin-b2a in driving these changes in subcellular localization will be elucidated by performing FCCS in live mutant embryos. In Aim 2, a systems analysis of cell motion will be used to quantify tissue biomechanics during somite morphogenesis in wild-type and mutant embryos. In Aim 3, we quantify the relative levels of Integrin activation via cytoplasmic signals versus via positive feedback through the ECM. Positive and negative feedback between biological mechanisms creates network effects that are hard to predict a priori and difficult to fully explore experimentally. in silico modeling will be used to systematically examine the relationships between cell adhesion, cell-ECM adhesion and cell contact mediated repulsion in somite morphogenesis in order to help interpret and prioritize more resource intensive wet-lab experiments.

ECM和相关上皮细胞之间的相互调节是发育、体内平衡和疾病。体节是脊椎骨的节段性前体通过间充质向上皮转化形成的柱状和肌肉组织。体节形态发生依赖于纤连蛋白ECM，纤连蛋白受体整合素细胞粘附蛋白Cadherin 2和通过受体的双向信号传导酪氨酸激酶EphA 4及其膜结合配体Ephrin-B2 a。这些基因/通路介导细胞-ECM粘附、细胞-细胞粘附和接触介导的细胞排斥，假设体节边界的物理组织活动是通过差异细胞粘附和ECM约束细胞的特定时空交织排斥在目标1中，荧光相关光谱（FCS）和荧光交叉相关光谱（FCCS）将用于定量蛋白质扩散和蛋白质体内结合常数。这些实验将确定是否分离的这些细胞表面蛋白通过扩散和捕获或主动移动而发生。此外，整合素β 5、钙粘蛋白2和肝配蛋白-b2 a在驱动这些变化中的作用，亚细胞定位将通过在活的突变胚胎中进行FCCS来阐明。在目的2，细胞运动的系统分析将用于量化组织生物力学，在野生型和突变体胚胎体节形态发生。在目标3中，我们量化了通过细胞质信号与通过正反馈的整合素激活水平 ECM。生物机制之间的正反馈和负反馈产生网络效应很难事先预测，也很难通过实验充分探索。计算机建模将用于系统地研究细胞粘附、细胞-ECM 粘附和细胞接触介导的排斥在体节形态发生，解释和优先考虑更多的资源密集型湿实验室实验。