Investigation of the Cellular and Molecular Mechanisms of Thrombocyte Integrin Signaling

血小板整合素信号传导的细胞和分子机制研究

• 批准号: 10616738
• 负责人: Zhao Wang
• 金额: $ 39.66万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-05-05 至 2026-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10616738
• 关键词: 3-Dimensional; Adaptor Signaling Protein; Affinity; Agonist; Architecture; Binding; Biochemical; Biological Assay; Biology; Blood Coagulation Disorders; Blood Platelets; Cell Surface Receptors; Cell surface; Cells; Cessation of life; Cholesterol; Classification; Clinical; Complex; Cryo-electron tomography; Cryoelectron Microscopy; Cytoskeleton; Defect; Detergents; Development; Disease; Dose; Drug Design; Drug Targeting; Drug usage; EGF gene; Equilibrium; Event; Family; Feasibility Studies; Fibrinogen; Future; Goals; Hematological Disease; Hemorrhage; Hour; Human; Hybrids; Imaging Techniques; Immunotherapy; In Situ; In Vitro; Inflammation; Integral Membrane Protein; Integrins; Investigation; Ions; Kinetics; Laboratories; Length; Licensing; Ligands; Link; Macrophage-1 Antigen; Malignant Neoplasms; Maps; Membrane; Modeling; Molecular; Molecular Conformation; Molecular Probes; Monoclonal Antibodies; Myocardial Infarction; Pathology; Pharmaceutical Preparations; Physiological; Platelet Activation; Platelet aggregation; Process; Property; Protein Structure Initiative; Proteins; Regulation; Reporting; Research; Resolution; Roentgen Rays; Role; Shapes; Signal Transduction; Specimen; Stroke; Structure; Surface; Techniques; Therapeutic; Thermodynamics; Thick; Thigh structure; Thrombasthenia; Thrombocytopenia; Thrombosis; Time; Vesicle; Visualization; antagonist; biophysical techniques; clinical effect; clinically relevant; effective therapy; experimental study; improved; in situ imaging; inhibitor; insight; microvesicles; nanodisk; nanometer resolution; novel; particle; platelet function; pre-clinical; receptor; reconstitution; skills; small molecule; structural biology; success; targeted treatment; therapeutic target; thrombotic; tomography; von Willebrand Factor

Abstract Platelet integrin αIIbβ3 is central to platelet activation, which occurs through highly complex molecular interactions and structural alteration. Ultimately, αIIbβ3 matures its affinity for fibrinogen and von Willebrand factor. Dysregulation in αIIbβ3 affinity maturation events causes Glanzmann's thrombasthenia or thrombocytopenia. Therefore, fine-tuning these interactions through the αIIbβ3 receptor is a proven and effective strategy for both therapeutic thrombotic pathology targeting and immunotherapy. Currently, there are no αIIbβ3 inhibitors for long-term clinical use, while current inhibitors for short-term use may cause serious thrombocytopenia. The overall goal of our proposed research is to understand the molecular basis of the αIIbβ3 shapeshifting and the mechanochemistry of αIIbβ3 activation and inhibition by small molecules, including clinical- drugs. Our guiding hypothesis is that multiple forms of αIIbβ3 exist in a dynamic conformational equilibrium that is temporally and spatially regulated by cell signaling, association with the cytoskeleton, and interactions with exocellular ligands. We will determine, for the first time, the single-particle cryoEM structure of intact αIIbβ3, characterize structural transitions at the atomic level as they relate to physiological ligands (e.g., fibrinogen), and determine the effects of clinically relevant antagonists on αIIbβ3 conformational equilibrium both in situ and in vitro. All specific aims will elaborate integrin αIIbβ3 events at the molecular level pertaining to these objectives. Aim 1 experiments will characterize shapeshifting structural changes of αIIbβ3 induced by ligands or clinical- relevant drugs. We will investigate the kinetics of the conformational dynamics of the purified full-length αIIbβ3 using a single-particle cryoEM approach and wide-ranging biochemical techniques for the protein in solution. Aim 2 experiments will study architectural changes in intact αIIbβ3 by probing its conformational states on the cell surfaces using conformation-reporting mAbs and by directly solving in situ structures using cutting-edge advanced cryoET imaging techniques. Collectively, our proposed studies will provide unique molecular insights into structural and bidirectional signaling regulation of αIIbβ3, which will advance our understanding of integrin biology and may identify new antagonists for modulating αIIbβ3 function.

摘要