Modeling SNARE-Mediated Membrane Fusion

SNARE 介导的膜融合建模

• 批准号: 10445738
• 负责人: Ben O'Shaughnessy
• 金额: $ 33.89万
• 依托单位: COLUMBIA UNIV NEW YORK MORNINGSIDE
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-01 至 2026-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10445738
• 关键词: Action Potentials; Beta Cell; Biochemical; Catalogs; Cell fusion; Cell membrane; Cells; Chromaffin Cells; Closure by clamp; Communities; Complex; Computer Models; Core Protein; Coupled; Data; Dense Core Vesicle; Docking; Dynamin; Entropy; Event; Evoked Potentials; Evolution; Exocytosis; Family; Functional disorder; Funding; Goals; Grain; Growth; Hormones; Hot Spot; Impairment; Informal Social Control; Kinetics; Laboratories; Machine Learning; Measurement; Mechanics; Mediating; Membrane; Membrane Fusion; Membrane Lipids; Modeling; Molecular; Mutate; Mutation; Neuraxis; Neurodegenerative Disorders; Neurodevelopmental Disorder; Neuroendocrine Cell; Neurons; Neurotransmitters; Pathway interactions; Physiological; Probability; Process; Property; Proteins; Regulation of Exocytosis; Reproducibility; Research; Respiratory Diaphragm; Rest; Running; S-nitro-N-acetylpenicillamine; SNAP receptor; Scheme; Secretory Vesicles; Shapes; Signal Transduction; Site; Stimulus; Structure; Study models; Synapses; Synaptic Cleft; Synaptic Vesicles; Testing; Thinness; Type 2 diabetic; Vesicle; experimental study; foot; hydrophilicity; in silico; insulin secretion; mathematical model; membrane model; millisecond; molecular dynamics; multi-scale modeling; nanodisk; neurotransmission; neurotransmitter release; novel; predictive modeling; reconstitution; response; sensor; simulation; synaptotagmin; synaptotagmin I; synaptotagmin II; syntaxin; target SNARE proteins; vesicular SNARE proteins; vesicular release

PROJECT SUMMARY Many basic processes rely on secretion of bioactive molecules by exocytosis, when membrane-enclosed vesicles containing neurotransmitters (NTs), hormones or other molecules fuse with the plasma membrane (PM) and release their contents through fusion pores. Neurotransmission relies on NT release at neuronal synapses, when a multicomponent machinery senses Ca!"influx triggered by an action potential and fuses small synaptic vesicles with the plasma membrane on sub-millisecond timescales, releasing NTs into the synaptic cleft to elicit a post-synaptic response. Other regulated exocytosis is slower, such as hormone release from neuroendocrine cells when a stimulus provokes large dense core vesicles to release contents after seconds or longer. In all cases, the membrane fusion step is accomplished by the SNARE proteins, when vesicle-associated VAMP (the v-SNARE) and two PM-associated t-SNAREs syntaxin and SNAP 25 zipper into a ternary complex, pulling the membranes together and fusing them. However, the mechanism of membrane fusion remains unclear. Other components in the machinery block (“clamp”) SNARE-mediated fusion, until the Ca!"signal releases the clamp. Synaptotagmin (Syt) is the Ca!"sensor for synchronous release, but the molecular identity of the clamp and the Ca!"-triggered unclamping mechanism are not established. Mutations in SNARE proteins and other NT release machinery components are associated with neurodevelopmental and neurodegenerative disorders, and impaired fusion pore dilation is associated with reduced insulin secretion by β-cells of type-2 diabetics. The proposed research aims to use mathematical modeling to establish the mechanisms of regulated membrane fusion and the mechanisms that regulate the vesicle and its pore for controlled contents release following fusion. From the previous funding period, we have working molecular dynamics (MD) simulations of SNARE-mediated membrane fusion and of the NT release machinery incorporating the core SNARE and Syt components. The simulations used sufficiently coarse-grained (CG) representations to achieve the computationally demanding millisecond physiological timescales of fusion and release. Aim 1 is to advance the SNARE-mediated fusion simulations with more realistic SNAREs, and to use machine learning to catalogue the pathways to membrane fusion as a function of the size of the fusing vesicles and other key variables. Mutated SNAREs will be simulated and compared to experiments by collaborators. Aim 2 is to use continuum mathematical modeling to establish the structure, energetics and evolution of the fused vesicle-PM complex and its fusion pore, and the mechanisms of SNARE- and Syt-mediated pore dilation. Aim 3 is to advance the MD simulations of the NT release machinery by introducing molecularly explicit representation of the membranes, and by incorporating additional components as their interactions become experimentally characterized, toward a long-term goal of “reconstituting” the machinery in silico. Simulations will test hypothesized Ca!"-triggered unclamping schemes, and will be run with mutations in the SNARE and Syt components that will be implemented experimentally by our collaborators.

项目摘要 许多基本过程依赖于胞吞作用的生物活性分子的分泌，当膜粘附时 含有神经递质（NTS），激素或其他分子与质膜（PM）融合的囊泡 并通过融合孔释放其内容。 当多组分机械感应CA时！ 带有质膜的速率在子毫秒时期，将NTS释放到突​​触裂缝中以引起 突触后的重新统治。 当astimulus引起大型密集的核囊泡时，细胞在几秒钟或更长时间后释放含量。 在所有情况下，当囊泡相关的鞋面时，膜融合步骤都是由圈蛋白完成的 （V-SNARE）和两个与PM相关的T-SNARES语法素，然后将25拉链捕入三元络合物 膜融合在一起，但是，膜融合的机理尚不清楚。 机械块中的组件（“夹具”）圈养介导的融合，直到CA！”信号释放出来。 Synaptotagmin（SYT）是CA！ CA !!”未建立触发的解开机制。 机械组件与神经发育和神经退行性疾病，且d和d 受损的融合孔扩张与2型糖尿病患者的β细胞减少有关。 拟议的研究旨在使用数学建模来建立常规膜的机制 融合及其在融合后的孔孔及其孔孔前的孔孔前的孔孔的原理。 从上一个资金期开始，我们进行了工作分子动力学（MD）模拟 膜融合和NT释放机械融合了核心网罗和SYT组件 模拟使用了足够粗糙的（CG）代表来实现计算要求 融合和释放的毫秒生理时间尺度。 模拟使用更逼真的弹头，并使用机器学习来对通往膜的途径进行分类 融合是大惊小怪的囊泡和oter键变量的函数。 与合作者的实验相比，AIM 2是使用连续数学建模 融合囊泡-PM复合物的结构，能量和演变，是融合孔，机理 SNARE和SYT介导的孔隙扩张。 通过引入分子明确表示膜，并结合其他组件 随着它们的相互作用的特征，朝着“重新构成”的长期目标 模拟中的机械将测试假设的CA！ 我们的合作者将实施的工资和SYT组件中的突变。