Mechanisms of Membrane Fusion

膜融合机制

• 批准号: 9278209
• 负责人: WILLIAM Tobey WICKNER
• 金额: $ 74.7万
• 依托单位: DARTMOUTH COLLEGE
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-07-01 至 2021-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9278209
• 关键词: Bacteria; Biological Assay; Biological Models; Cells; Chemicals; Complex; Controlled Study; Coxiella burnetii; Ebola virus; Frankfurt-Marburg Syndrome Virus; Genetic; Genetic study; Hormones; Human; In Vitro; Infection; Light; Lipid Bilayers; Lipids; Medical; Membrane; Membrane Fusion; Membrane Proteins; Molecular Chaperones; Organelles; Physiology; Proteins; Regulation; Role; SNAP receptor; Somatotropin; Stress; System; Testing; Vacuole; Variant; Yeasts; cell growth; neurotransmission; pathogen; pathogenic bacteria; proteoliposomes; public health relevance; rab GTP-Binding Proteins; reconstitution; snRNP Structural Core Protein

 DESCRIPTION (provided by applicant): Membrane fusion underlies hormone secretion, neurotransmission, and all exocytic and endocytic traffic. Its mechanism is conserved from yeast to humans. A current paradigm suggests that membrane proteins termed SNAREs inexorably drive fusion when anchored in apposed membranes as a trans-SNARE complex. While SNAREs are required, genetic studies from yeast to humans show that Rab GTPases, their effectors, SM proteins, and SNARE chaperones are also essential. Our studies of yeast vacuole fusion are providing a new paradigm, illuminating the integrated mechanisms of these other essential proteins and showing that their actions extend beyond regulation of trans-SNARE complex levels. Vacuole fusion studies have progressed from initial genetics through our extensive study of in vitro organelle fusion to proteoliposome fusion, which we have reconstituted with all purified and defined proteins and lipids: SNAREs, SNARE disassembly chaperones Sec18p/Sec17p, the Rab GTPase Ypt7p, a hexameric Rab effector complex HOPS, and lipids which include acidic and bilayer-averse headgroups and specific fatty acyl chains. With a rigorous fusion assay of protected lumenal content mixing, our studies show that fusion is driven by several cooperating factors: bilayer stress from trans-SNARE complex assembly, membrane destabilization by nonbilayer lipids, and bilayer bending through the action of a multisubunit tether. Fusion is blocked by the omission of SNAREs, of nonbilayer-prone lipids, or of tethering factors, even though SNAREs still pair in trans in the latter 2 conditions. Our chemically-defined reconstitution of fusion allows independent variation of SNARE concentration, nonbilayer lipid concentration, and the HOPS and Rab tethering proteins, all while assaying both physical associations and fusion function. Testing and extending this model system is changing our view of fusion. In light of the fundamental role of fusion throughout human physiology, and the central role of human HOPS for cellular infection by Marburg and Ebola viruses and by bacteria such as the pathogen Coxiella burnetii, these studies will be of medical significance as well.

 描述（由适用提供）：膜融合是激素分泌，神经传递以及所有外囊肿和内吞流量的基础。它的机制是从酵母到人类的。当前的范式表明，当将膜蛋白称为网状物质时，当锚定在应用机制中作为跨鼻涕复合物时，无可呈驱动的融合。虽然需要咆哮，但从酵母到人类的遗传研究表明，Rab GTPases，其作用，SM蛋白和圈圈伴侣也是必不可少的。我们对酵母菌融合的研究正在提供新的范式，从而阐明了这些其他必要蛋白质的综合机制，并表明它们的作用超出了跨效复合物水平的调节。 Vacuole fusion studies have progressed from initial genetics through our extensive study of in vitro organic fusion to proteoliposome fusion, which we have reconstituted with all purified and defined proteins and lipids: SNAREs, SNARE disassembly chainones Sec18p/Sec17p, the Rab GTPase Ypt7p, a hexameric Rab effector complex HOPS, and lipids which include acidic and避开双层的头组和特定的脂肪酰基链。通过对受保护的腔内含量混合的严格融合测定，我们的研究表明，融合是由几个合作因素驱动的：跨源头复合物组装中的双层应力，非叶叶脂质的膜不稳定，而双层均通过二体基团Tether的作用而弯曲。融合会因省略窃贼，非质易纤维脂质或绑扎因子而阻塞，即使在后2条条件下，小偷仍然在trans中配对。 我们的化学定义融合重构允许对圈圈浓度，非叶质脂质浓度以及啤酒花和Rab绑扎蛋白的独立变化，同时既主张物理关联和融合功能。测试和扩展此模型系统正在改变我们对融合的看法。鉴于融合在整个人类生理学中的基本作用，以及人类啤酒花在Marburg和Ebola病毒以及通过细菌（例如病原体Coxiella burnetii）感染细胞感染的核心作用，这些研究也具有医学意义。