Mechanisms of Intracellular Membrane Fusion

细胞内膜融合的机制

• 批准号: 8032992
• 负责人: JAMES ROTHMAN
• 金额: $ 16.55万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-03-23 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8032992
• 关键词: Adhesions; Adhesives; Affect; Amino Acid Sequence; Area; Behavior; Biochemistry; Biological; Biological Process; Biophysics; Cell membrane; Cells; Cellular Membrane; Chimeric Proteins; Coiled-Coil Domain; Collaborations; Complex; Confocal Microscopy; Development; Diabetes Mellitus; Disease; Electrophysiology (science); Endocrine; Environment; Event; Exhibits; Exocytosis; Exposure to; Funding; Grant; Homo; Image; Indium; Individual; Intracellular Membranes; Kinetics; Lateral; Link; Lipids; Measurement; Measures; Membrane; Membrane Fusion; Micromanipulation; Molecular; Molecular and Cellular Biology; Mutation; Nature; Outcome; Paris, France; Peptide Sequence Determination; Phenotype; Physicians; Physiological; Physiology; Pore Proteins; Process; Protein Isoforms; Proteins; Research; Research Personnel; Retinal Cone; Running; SNAP receptor; Sampling; Shapes; Structure; Surface; Surface Tension; Surgeon; System; Technology; Time; Transmembrane Domain; Universities; Vesicle; Viral Fusion Proteins; Work; college; flexibility; insight; interdisciplinary approach; lipid structure; mast cell; mutant; optical imaging; palmitoylation; programs; protein folding; proteoliposomes; reconstitution; syntaxin

DESCRIPTION (provided by applicant): Membrane fusion is central to many areas of endocrine and exocrine physiology, and imbalances in these processes give rise to important diseases, such as diabetes. As a result of the work supported by this grant during the current cycle of funding, the core principle of cellular membrane fusion is now well established, consisting of the assembly of cognate SNARE proteins initially residing in apposing membranes to yield a stable, bridging complex that triggers the bilayers to merge. How does the assembly of SNARE proteins between membranes drive membrane fusion? A mechanistic understanding of the fusion event, in which as many as four separate proteins fold together to fuse apposing bilayers, will require the combined power of a variety of biophysical approaches. Such an undertaking has become possible only recently, thanks to continuing advances in both SNARE fusion biochemistry and biophysical membrane technologies. We will use different complementary technologies (cellular and molecular biology, electrophysiology, surface force/adhesion, and optical imaging), to follow SNARE dynamics and function in real time. SNARE proteins will be reconstituted in both synthetic and biological membranes, thus allowing measurements in fully reconstituted membrane environments as well as in less flexible but more physiological cellular membranes. Within these systems, we will determine the energetics of SNARE-assembly, the consequences of membrane composition and membrane tension on fusion, and the dynamics of the fusion pore itself. Insights gained in the different approaches can be linked by comparing the effects of critical mutations and other perturbations. By following this program, whose value has been proven with viral fusion proteins, we expect new and important information concerning cellular membrane fusion to emerge during the next five years.

描述（由申请人提供）：膜融合是内分泌和外分泌生理学许多领域的核心，这些过程中的不平衡引起重要疾病，如糖尿病。由于在当前的资助周期中获得了该资助支持的工作，细胞膜融合的核心原则现已确立，包括最初位于并列膜中的同源SNARE蛋白的组装，以产生稳定的桥接复合物，触发双层合并。SNARE蛋白在膜间的组装如何驱动膜融合？一个机械的理解融合事件，其中多达四个单独的蛋白质折叠在一起融合并列双层，将需要各种生物物理方法的综合力量。由于SNARE融合生物化学和生物物理膜技术的不断进步，这种任务直到最近才成为可能。我们将使用不同的补充技术（细胞和分子生物学，电生理学，表面力/粘附和光学成像），以遵循SNARE动态和功能在真实的时间。SNARE蛋白质将在合成膜和生物膜中重建，从而允许在完全重建的膜环境中以及在柔性较低但更具生理性的细胞膜中进行测量。在这些系统中，我们将确定陷阱组装的能量学，膜组成和膜张力对融合的影响，以及融合孔本身的动力学。通过比较关键突变和其他扰动的影响，可以将不同方法中获得的见解联系起来。通过遵循这一计划，其价值已被证明与病毒融合蛋白，我们预计新的和重要的信息，有关细胞膜融合出现在未来五年。