The Role of Myosin 2 in Contact Guidance

肌球蛋白 2 在接触指导中的作用

• 批准号: 9557314
• 负责人: Robert Adelstein
• 金额: $ 51.71万
• 依托单位: NATIONAL HEART, LUNG, AND BLOOD INSTITUTE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9557314
• 关键词: Acrylamides; Actins; Address; Adhesions; Adult; Affect; Architecture; Cell Adhesion; Cell Line; Cell model; Cells; Chemicals; Cytoskeleton; Deletion Mutation; Development; Elasticity; Embryo; Embryonic Development; Fibroblasts; Fibronectins; Gel; Gene Expression; Genetic Transformation; Geometry; Knock-out; Maintenance; Mechanics; Microtubules; Modeling; Myosin ATPase; Myosin S-2; Pattern; Positioning Attribute; Protein Isoforms; Proteins; Reading; Regulation; Reproducibility; Research; Role; Series; Signal Transduction; Solid; Structure; Surface; Techniques; Testing; Tissues; Tubulin; blastomere structure; cell motility; in vivo; migration; non-muscle myosin; polydimethylsiloxane

To investigate the contribution of NMs 2 to cell contact guidance we established a microprinting technique. We tested the effect of the NM 2B isoform knockout by visualizing cell spreading and dynamics of the cytoarchitecture on fibronectin coated micro-patterns on soft acrylamide gels and hard polydimethylsiloxane surfaces. This allows us to elucidate proteins that guide spreading and migration of fibroblast cells. We found that absence of NM 2B altered contact guidance of fibroblasts manifesting in a substantial loss of cellular projections and uniaxial directional polarization as well as increased contractility and elasticity of cells. Using cytoskeleton targeting compounds and a series of genetic transformations we verified a possible role for the NM 2B isoform in contact guidance and proposed a model in which the roles of NM 2A and 2B isoforms diverge in control over structure of the actin cytoskeleton and its crosstalk with microtubules. To assess reproducibility and verify the role of NMs 2 in contact guidance, an independent series of cell lines will be tested. This second series will accommodate treatment with cytoskeleton targeting compounds and a set of genetic transformation to affect gene expression in a manner similar to the one used previously for fibroblast cells. We are creating and testing a model of cell cytoskeleton architecture dynamics that incorporates actin-tubulin-myosin dependent regulation of contact guidance. This model creates a platform for the study of the roles of NMs 2 in tissue development.

为了研究 NMs 2 对细胞接触引导的贡献，我们建立了一种微印刷技术。我们通过可视化软丙烯酰胺凝胶和硬聚二甲基硅氧烷表面上纤连蛋白包被微图案上的细胞扩散和细胞结构动力学来测试 NM 2B 同种型敲除的效果。这使我们能够阐明指导成纤维细胞扩散和迁移的蛋白质。我们发现，NM 2B 的缺失改变了成纤维细胞的接触引导，表现为细胞投射和单轴定向极化的大量损失以及细胞收缩性和弹性的增加。 使用细胞骨架靶向化合物和一系列遗传转化，我们验证了 NM 2B 同工型在接触引导中的可能作用，并提出了一个模型，其中 NM 2A 和 2B 同工型在控制肌动蛋白细胞骨架的结构及其与微管。为了评估重现性并验证 NMs 2 在接触指导中的作用，将测试一系列独立的细胞系。第二个系列将采用细胞骨架靶向化合物进行治疗，并进行一系列遗传转化，以类似于之前用于成纤维细胞的方式影响基因表达。我们正在创建和测试细胞骨架结构动力学模型，该模型结合了肌动蛋白-微管蛋白-肌球蛋白依赖性接触引导调节。该模型为研究 NMs 2 在组织发育中的作用创建了一个平台。