CAREER: Structural and Biochemical Studies of the Tiam1 GEF, a Paradigm for Understanding Modular Protein Signal Transduction

Rho家族的小鸟苷三磷酸(GTPase)蛋白在许多基本的生理过程中都是必不可少的，包括细胞分化、分裂、趋化、轴突发育和极性。GTP酶起到二进制“开/关”开关的作用，控制细胞内的通讯。鸟嘌呤核苷酸交换因子(GEF)密切控制GTP酶的活性状态，协调其功能。然而，全球环境基金蛋白如何被调控以及它们如何引导GTPase信号转导的基本机制仍然知之甚少。该项目的重点是Tiam1，这是一种大型、模块化的全环基金，它专门激活调节细胞极性、黏附和迁移的GTPase(Rac1)。在正常情况下，Tiam1环境基金通过蛋白质-蛋白质相互作用和/或通过磷酸化来抑制其自身的活性。一旦激活，Tiam1就能够与多个蛋白质靶点相互作用，并影响许多细胞过程。在这项研究中，将使用结构、生物物理和生化方法相结合的方法来确定Tiam1自身抑制的分子基础、它的激活和它的信号特异性。从这个项目中学到的基本概念将加深我们对信号转导的理解，并为模块信号蛋白的协调调控提供新的见解。此外，这项研究将有助于我们了解包括蛋白质变构和动力学在内的不同领域，并促进具有所需调控特性和功能的人造支架蛋白质的合理设计。广泛影响：这个职业项目的主要教育目标是让主要是本科生的教职员工和学生接触到前沿的生物物理学研究。该协会将每年举办一次“生物物理学研讨会”，与爱荷华州的PUI教员建立研究和教育合作关系。这些工作坊将向PUI的教师和他们的学生介绍现代实验生物物理学，并为他们提供必要的基础设施，以扩大他们的研究能力。此外，该协会还将设计一套“荣誉系列教程”，向高中生和本科生介绍基于网络的蛋白质和基因分析工具，以提升他们的暑期研究体验。本系列的受众将来自约100名参加校园暑期研究项目的高中生和本科生，包括SROP/McNair学者计划、爱荷华州生物科学优势计划、爱荷华州第一民族暑期计划和针对高中生的中学生培训计划。总的来说，这些活动将吸引教职员工和学生，包括科学上代表性不足的少数群体成员，参与先进的生物物理研究。该项目由分子和细胞生物科学部细胞系统集群和NSF EPSCoR办公室联合支持

The Rho-family of small guanosine triphosphatase (GTPase) proteins are essential for many basic physiological processes, including cell differentiation, division, chemotaxis, axonal development, and polarity. GTPases act as binary "on/off" switches and control communication within a cell. Guanine nucleotide exchange factors (GEFs) tightly control the active state of GTPases and coordinate their function. However, the fundamental mechanisms for how GEF proteins are regulated and how they direct GTPase signaling remain poorly understood. This project is focused on Tiam1, a large, modular GEF that specifically activates a GTPase (Rac1) that regulates cell polarity, adhesion, and migration. Under normal conditions Tiam1 GEF inhibits its own activity by protein-protein interactions and/or by phosphorylation. Upon activation, Tiam1 is capable of interacting with multiple protein targets and influencing many cell processes. In this study a combination of structural, biophysical, and biochemical approaches will be used to define the molecular basis of Tiam1 auto-inhibition, its activation, and its signaling specificity. The basic concepts learned from this project will enhance our understanding of signal transduction and provide new insights into the coordinated regulation of modular signaling proteins. In addition, this research will contribute to our understanding of diverse fields including protein allostery and dynamics, and promote the rational design of artificial scaffold proteins with desired regulatory features and functions.Broader Impacts: The major educational goal of this CAREER project is to expose faculty and students from primarily undergraduate institutions (PUI) to cutting-edge biophysics research. The PI will conduct a yearly "Biophysics Workshop" to establish research and education collaborations with PUI faculty throughout Iowa. These workshops will introduce PUI faculty and their students to modern experimental biophysics and provide access to the infrastructure necessary for them to expand their research capabilities. In addition, the PI will design an "Honors Tutorial Series" to introduce advanced high school and undergraduate students to web-based protein and gene analysis tools to enhance their summer research experience. The audience for this series will be drawn from ~100 high school and undergraduate students that participate in summer research programs across campus, including the SROP/McNair Scholars Program, Iowa Biosciences Advantage Program, Iowa First Nations Summer Program, and Secondary Student Training Program for high school students. Collectively, these activities will engage faculty and students, including members of minorities under-represented in science, in advanced biophysical research.This project is jointly supported by the Cellular Systems Cluster of the Molecular and Cellular Biosciences Division and the NSF EPSCoR Office