Spatio-temporal dynamics of GEF-GTPase networks

描述（由申请人提供）：Rho家族GTPases是普遍存在的分子开关，控制着非常多样化的细胞过程。它们被鸟嘌呤核苷酸交换因子（gef）激活，其数量大约是GTPase本身的5倍，并整合许多控制GTPase功能的细胞输入。全球环境基金和全球环境基金形成了复杂的网络，这些网络是临时和本地为特定目的而构成的。生物化学、遗传、分子和结构分析已经揭示了大量关于这些至关重要的途径，但最重要的功能特性，它们的时空调节，只能在完整细胞的背景下完全理解。该PPG汇集了具有不同专业知识的团队成员，以开发创新技术，实现GEF/GTPase网络在体内的研究，从成像数据中提取网络结构和信号动力学的计算工具，以及对严重依赖GEF/GTPase动力学的细胞行为的深入了解。（项目1- Hahn）将提供基于不同GEF结构类别设计的GEF生物传感器。在与GEF结构专家Sondek的合作下，不同的生物传感器设计将通过特定的上游输入和内源性GEF的激活来报告GEF的激活。（项目2- Danuser）将开发同时成像和/或光电操纵任何一对GEF和GTPase的活性的能力，用于GEF/GTPase时空协调的高分辨率研究。新的计算工具将结合来自不同实验的数据来模拟大型网络，并从成像数据中提取网络结构和信号动力学。这些方法将在复杂的GEF-GTPase反馈相互作用的研究中进行测试。（项目3-大厅）：这个以生物学为重点的项目将把我们的工作扩展到多细胞系统。我们将关注GEF在细胞-细胞连接和隐层板足中的激活，并确定GEF调节集体迁移。（项目4- Burridge）将解决GEF/GTPase网络在机械转导中的作用，探索在细胞-基质和细胞-细胞粘附过程中以及细胞核中RhoA信号的机械调节的新发现。

DESCRIPTION (provided by applicant): Rho family GTPases is ubiquitous molecular switches that control extraordinarily diverse cellular processes. They are activated by guanine nucleotide exchange factors (GEFs) that are roughly 5-fold more numerous than the GTPases themselves and integrate the many cellular inputs controlling GTPase function. GEFs and GTPases form complex networks that are constituted transiently and locally for specific purposes. Biochemical, genetic, molecular, and structural analyses have unraveled a great deal about these critically important pathways, but the most important functional property, their spatio-temporal regulation, can only be fully understood in the context of intact cells. This PPG brings together team members with diverse expertise to develop innovative technologies enabling the study of GEF/GTPase networks in vivo, computational tools to extract network architecture and signaling kinetics from imaging data, and in-depth knowledge of cell behaviors critically dependent on GEF/GTPase dynamics: (Project 1- Hahn) will deliver GEF biosensors based on designs addressing different GEF structural classes. In a collaborative effort with Sondek, expert in GEF structure, different biosensor designs will report GEF activation by specific upstream inputs, and activation of endogenous GEFs. (Project 2- Danuser) will develop the ability to simultaneously image and/or photomanipulate the activity of any pair of GEFs and GTPases, for high resolution studies of GEF/GTPase spatio-temporal coordination. New computational tools will combine data from different experiments to model large networks, and to extract network architecture and signaling kinetics from imaging data. These methods will be tested in studies of complex GEF-GTPase feedback interactions. (Project 3- Hall): This biologically focused project will extend our work to multicellular systems. We will focus on GEF activation in cell-cell junctions and cryptic lamellipodia, and identify GEFs regulating collective migration. (Project 4- Burridge) will address the role of GEF/GTPase netvvorks in mechanotransduction, exploring novel findings regarding the mechanical regulation of RhoA signaling at cell-matrix and cell-cell adhesions during initiation of protrusions, and in the nucleus.