Position of UNC-13 in the active zone regulates synaptic vesicle release probability and release kinetics.
Position of UNC-13 in the active zone regulates synaptic vesicle release probability and release kinetics.
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作者：
Zhou K;Stawicki TM;Goncharov A;Jin Y
突触前活动区蛋白UNC-13/Munc13S通过直接与突触囊泡融合装置相互作用，对突触囊泡的胞吐起重要作用。一个悬而未决的问题是，它们与活动区的联系，从而它们与钙离子进入部位的位置，如何调节SV的释放。主要的UNC-13/Munc13亚型的N-末端含有一个非钙结合的C2a结构域，它介导蛋白质的同质或异质相互作用。在这里，我们证明了秀丽线虫UNC-13的C2a结构域调节诱导释放的释放概率及其精确的活动区定位。SV释放的动力学分析支持，由含有N-末端的C2a结构域介导的UNC-13与钙离子进入位点的接近是加速神经递质释放的关键。此外，C2a结构域是自发释放所必需的。这些数据揭示了UNC-13C2a结构域的多重作用，并提示自发释放和诱发释放的快速阶段可能涉及活动区的公共SVs池。DOI：http://dx.doi.org/10.7554/eLife.01180.001神经元通过称为突触的连接相互连接。当沿着神经元传播的电信号到达突触时，它会导致称为突触小泡的泡状结构释放，其中含有称为神经递质的化学物质。当被小泡释放时，这些神经递质与第二个神经元上的受体结合，并允许信号继续通过神经系统。神经元释放突触小泡在很大程度上取决于通过称为离子通道的结构进入神经元的钙离子的数量，也取决于钙离子进入神经元的速度。囊泡以三种方式之一释放：它们可以响应钙离子的流入而迅速释放(在几毫秒内)；它们可以随着钙离子的流入而缓慢释放(在几十或数百毫秒的时间内)；或者它们可以在与钙离子流入无关的随机时间释放。众所周知，突触附近某些钙感受器的敏感性会影响囊泡的释放。曾经有人认为，钙离子进入神经元的“活动区”和囊泡所在区域之间的距离也可能影响释放速度，但这一假说背后的分子机制尚不清楚。周等人。通过进行一系列实验，现在已经为这个问题提供了新的线索，这些实验涉及操纵线虫线虫神经元中一种名为UNC-13的蛋白质--已知这种蛋白质参与囊泡的释放。首先，研究表明，UNC-13在活动区的准确位置取决于蛋白质中称为C2a结构域的结构域。其次，研究表明UNC-13蛋白与钙离子通道之间的距离强烈影响囊泡的快速释放模式。最后，周等人。结果表明，C2a结构域对囊泡的自发释放也有显著的影响，这表明一个共同的囊泡车队可能同时用于快速和自发的囊泡释放模式。周等人。还产生了模仿神经疾病癫痫发作的突变蠕虫，并表明消除C2a结构域可以缓解与疾病相关的一些症状。许多神经疾病是由突触信号传递不正确引起的，因此，除了为突触活动的基本机制提供见解外，这些结果还可能有助于开发管理神经疾病的新策略。DOI：http://dx.doi.org/10.7554/eLife.01180.002
The presynaptic active zone proteins UNC-13/Munc13s are essential for synaptic vesicle (SV) exocytosis by directly interacting with SV fusion apparatus. An open question is how their association with active zones, hence their position to Ca2+ entry sites, regulates SV release. The N-termini of major UNC-13/Munc13 isoforms contain a non-calcium binding C2A domain that mediates protein homo- or hetero-meric interactions. Here, we show that the C2A domain of Caenorhabditis elegans UNC-13 regulates release probability of evoked release and its precise active zone localization. Kinetics analysis of SV release supports that the proximity of UNC-13 to Ca2+ entry sites, mediated by the C2A-domain containing N-terminus, is critical for accelerating neurotransmitter release. Additionally, the C2A domain is specifically required for spontaneous release. These data reveal multiple roles of UNC-13 C2A domain, and suggest that spontaneous release and the fast phase of evoked release may involve a common pool of SVs at the active zone. DOI: http://dx.doi.org/10.7554/eLife.01180.001 Neurons are connected to each other by junctions called synapses. When an electrical signal travelling along a neuron arrives at a synapse, it causes the release of bubble-like structures called synaptic vesicles that contain chemicals called neurotransmitters. When released by the vesicles these neurotransmitters bind to receptors on a second neuron and allow the signal to continue on its way through the nervous system. The release of synaptic vesicles from the neuron depends largely on the number of calcium ions that enter this neuron via structures called ion channels, and also on the rate at which they enter. Vesicles are released in one of three ways: they can be released quickly (within a few milliseconds) in response to the influx of calcium ions; they can be released slowly (over a period of tens or hundreds of milliseconds) in response to the influx; or they can be released at random times that are not related to the influx. It is known that the sensitivity of certain calcium sensors near the synapse influences the release of the vesicles. It had been thought that the distance between the “active zone” where the calcium ions enter the neuron and the region where the vesicles reside might also influence rate of release, but the molecular mechanism underlying this hypothesis is poorly understood. Zhou et al. have now shed new light on this question by performing a series of experiments that involved manipulating a protein called UNC-13 – which is known to be involved in the release of vesicles – in neurons from C. elegans, a nematode worm. First it was shown that the precise position of UNC-13 in the active zone depended on a domain within the protein called the C2A domain. Next it was shown that the distance between the UNC-13 protein and the calcium ion channels strongly influences the quick mode of vesicle release. Finally, Zhou et al. showed that the C2A domain also had a significant influence on the spontaneous release of vesicles, which suggests that a common fleet of vesicles might be used for both the quick and the spontaneous modes of vesicle release. Zhou et al. also generated mutant worms that mimicked a neurological disease, epileptic seizure, and showed that eliminating the C2A domain can relieve some of the symptoms associated with the disease. Many neurological diseases are caused by signals not being transmitted properly at synapses, so in addition to providing insights into the basic mechanism underlying synaptic action, these results could also assist with the development of new strategies for managing neurological diseases. DOI: http://dx.doi.org/10.7554/eLife.01180.002