The Molecular Basis for Integrin-Mediated Bidirectional Signaling

白细胞是保护身体免受外来入侵者的重要免疫成分。 粘附、迁移、外渗和细胞间通讯都是通过膜介导的。 β2整合素是在细胞膜上发现的整合膜蛋白， 白细胞表面由于β2整合素的复杂和多功能作用， 与自身免疫、心脏和肺部病变以及传染病有关， 几种癌症。虽然整联蛋白是主要的治疗靶点，但药物开发一直是 阻碍由于意外的副作用，产生了巨大的差距，在我们的理解， 驱动特异性和整合素激活的机制。我会破译出 β2整合素活化、配体识别和双向信号传导 approach.我将利用我在冷冻电镜方法开发方面的专业知识， 分离的β2整联蛋白和配体结合的复合物的构象快照，以揭示 与信号转导相关的动态结构重排，并确定关键残基 介导配体特异性。使用细胞表面表达的整合素，我将评估功能性 突变这些残基对配体和构象特异性结合的影响 抗体和粘附、吞噬作用和细胞运动。深入了解整合素 在接近天然的环境中变构，我将开发一个膜模拟系统，使用下一个- 第二代苯乙烯马来酸共聚物。膜脂质影响整合素活化， 配体结合并且是形成稳定复合物的关键。这些聚合物将提供一个流- 提取和纯化包埋有预形成的整联蛋白-配体的纳米盘的线性方法 复杂的自然环境。我将用这个系统来研究整合素与 talin是一种与细胞质上的保守基序结合的中心整合素激活蛋白， 整合素区域。这将揭示分子的细节整合素是如何被激活中继信号 变构穿过质膜，并定义了双向调节的分子基础。 信号，并提供了一个框架，用于设计生物化学，生物物理，和机械， 灵敏的实验，研究更大的复合物，并获得全面的洞察整合素 功能最终，这项工作将提供一个结构蓝图的合理设计， 治疗自身免疫性疾病，这是实验室的长期目标。

Leukocytes are essential immune components protecting the body against foreign invaders. Adhesion, migration, extravasation, and cell-cell communication are mediated though the bidirectional signaling of β2 integrins, which are integral membrane proteins found on the leukocyte surface. Due to their complex and multifunctional roles, dysregulation of β2 integrins is linked to autoimmune, cardiac and pulmonary pathologies as well as infectious diseases and several cancers. Although integrins are prime therapeutic targets, drug development has been hindered due to unanticipated side effects that arise from large gaps in our understanding of the mechanisms that drive specificity and integrin activation. I will decipher the molecular basis for β2 integrin activation, ligand recognition, and bidirectional signaling using an integrative approach. I'll build on my expertise in cryoEM method development to capture high- resolution conformational snapshots of isolated β2 integrin and ligand-bound complexes to reveal the dynamic structural rearrangements associated with signal transduction and identify key residues mediating ligand specificity. Using cell-surface expressed integrins, I'll assess the functional consequences of mutating these residues on binding of ligands and conformation- specific antibodies and on adhesion, phagocytosis, and cell motility. To gain broad insight into integrin allostery in a near-native context, I'll develop a membrane mimetic system using next- generation styrene maleic acid copolymers. Membrane lipids influence integrin activation and ligand binding and are key to forming stable complexes. These polymers will afford a stream- lined method to extract and purify nanodiscs embedded with pre- formed integrin-ligand complexes in their native environment. I will use this system to study integrin in complex with talin, the central integrin- activator protein that binds a conserved motif on the cytoplasmic region on integrin. This will reveal in molecular detail how integrins are activated to relay signals allosterically across the plasma membrane and define a molecular basis for bidirectional signaling as well as provide a framework for designing biochemical, biophysical, and mechano- sensitive experiments to study larger complexes and gain comprehensive insight into integrin function. Ultimately, this work will provide a structural blueprint for the rational design of therapeutics for autoimmune diseases, which is a long-term goal of the lab.