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首页> 外文期刊>The Journal of Neuroscience: The Official Journal of the Society for Neuroscience >Titration of Syntaxin1 in mammalian synapses reveals multiple roles in vesicle docking, priming, and release probability
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Titration of Syntaxin1 in mammalian synapses reveals multiple roles in vesicle docking, priming, and release probability

机译：哺乳动物突触中Syntaxin1的滴定揭示了在囊泡对接，启动和释放概率中的多种作用

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摘要

Synaptic vesicles undergo sequential steps in preparation for neurotransmitter release. Individual SNARE proteins and the SNARE complex itself have been implicated in these processes. However, discrete effects of SNARE proteins on synaptic function have been difficult to assess using complete loss-of-function approaches. We therefore used a genetic titration technique in cultured mouse hippocampal neurons to evaluate the contribution of the neuronal SNARE protein Syntaxin1 (Stx1) in vesicle docking, priming, and release probability. We generated graded reductions of total Stx1 levels by combining two approaches, namely, endogenous hypomorphic expression of the isoform Stx1B and RNAi-mediated knockdown. Proximity of synaptic vesicles to the active zone was not strongly affected. However, overall release efficiency of affected neurons was severely impaired, as demonstrated by a smaller readily releasable pool size, slower refilling rate of primed vesicles, and lower release probability. Interestingly, dose-response fitting of Stx1 levels against readily releasable pool size and vesicular release probability showed similar Kd (dissociation constant) values at 18% and 19% of wild-type Stx1, with cooperativity estimates of 3.4 and 2.5, respectively. This strongly suggests that priming and vesicle fusion share the same molecular stoichiometry, and are governed by highly related mechanisms.
机译：突触小泡经历了连续的步骤以准备释放神经递质。单个的SNARE蛋白和SNARE复合物本身已牵涉到这些过程中。但是，使用完整的功能丧失方法难以评估SNARE蛋白对突触功能的离散影响。因此，我们在培养的小鼠海马神经元中使用了一种基因滴定技术来评估神经元SNARE蛋白Syntaxin1（Stx1）在囊泡对接，启动和释放概率中的作用。通过结合两种方法，即同工型Stx1B的内源性亚态表达和RNAi介导的敲低，我们产生了Stx1总水平的分级降低。突触小泡对活动区的接近程度没有受到很大影响。但是，受影响的神经元的总体释放效率受到严重损害，这表现为较小的易于释放的池大小，较慢的初生囊泡补充速度以及较低的释放概​​率。有趣的是，Stx1水平对易于释放的库大小和囊泡释放概率的剂量-反应拟合在野生型Stx1的18％和19％处显示相似的Kd（解离常数）值，协同性估计分别为3.4和2.5。这强烈表明，引发和囊泡融合具有相同的分子化学计量，并且受高度相关的机制支配。

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