Mechanisms of Synaptic Specificity in C. elegans

描述(申请人提供)：化学突触是一种特殊的细胞连接结构，对神经元之间的交流是必不可少的。在发育过程中，在特定的亚细胞隔间的特定神经元之间形成突触。突触靶点选择、轴突运输和突触前组装是突触形成的不可或缺的步骤，但人们对此知之甚少。在这里，我建议扩大我们的研究，以了解突触形成的两个基本方面：极化轴突运输和活动区蛋白聚集是如何受到调节的。突触通常形成在远端轴突和树突上，这给细胞体和突触之间的细胞内物质的有效交换带来了一个挑战性的问题。微管和MT相关的马达介导细胞内的运输。一般认为，运输的方向取决于两个因素：MTS的极性和所涉及的电机类型。基于我们已发表和未发表的数据，我们已经确定了两条对突触前成分运输至关重要的细胞周期蛋白依赖的激酶通路。在没有这两条通路的情况下，绝大多数突触小泡蛋白和活动区标志物无法定位到轴突，而是由于动蛋白马达的错误调节而在树突中被发现。突触形成中另一个鲜为人知的问题是，突触小泡的池大小是如何确定的。许多突触显示出突触小泡团的固定大小，这表明分子机制调节突触前终末局部突触小泡前体的组装。当适当数量的囊泡被招募时，可能会有一个负反馈系统来关闭组装途径。我们推测，如果这种反馈机制是有缺陷的，人们应该会看到突变的突触和异常的囊泡池。事实上，在正向遗传筛选中，我们分离出了一个突变体，在该突变体中，近端的突触异常大，而远端的突触含有很少的物质。我们还发现了一个类似Arf的GTPase，负责调节突触前特化的位置和大小。为了从机制上深入了解这些基因的功能，我特别计划了解：1)细胞周期蛋白依赖的激酶通路如何通过调节分子马达来控制轴突运输和突触定位，以及2)类似GTPase的ARF，ARL8如何调节突触囊泡池的大小和突触前组装。鉴于许多神经疾病都与突触连接的改变有关，而且细胞周期蛋白依赖性蛋白激酶与神经退行性疾病有关，希望这个项目将有助于理解大脑在生理和病理条件下的发育。

DESCRIPTION (provided by applicant): Chemical synapses are specialized cellular junction structures that are essential for communication between neurons. During development, synapses form between specific neurons at defined subcellular compartments. Synaptic target selection, axonal transport and presynaptic assembly are integral steps of synapse formation that are poorly understood. Here, I propose to expand our research to understand two essential aspects of synapse formation: polarized axonal trafficking and how aggregation of active zone proteins is regulated. Synapses are usually formed on distal axon and dendrites, creating a challenging problem for effective exchange of intracellular material between cell bodies and synapses. Microtubules and MT associated motors mediate intracellular trafficking. It is generally believed that the direction of transport depends on two factors: the polarity of MTs and the type of motor involved. Based on our published and unpublished data, we have identified two cyclin-dependent kinase pathways that are essential for the trafficking of presynaptic components. In the absence of both pathways, the vast majority of synaptic vesicle proteins and active zone markers fail to localize to axon and instead are found in dendrites due to misregulation of kinesin motors. Another poorly understood question in synapse formation is how the pool size of synaptic vesicles is determined. Many synapses display stereotyped size of synaptic vesicle clusters, suggesting that molecular mechanisms regulate the assembly of synaptic vesicle precursors locally at the presynaptic terminals. When the appropriate number of vesicles is recruited, there might be a negative feedback system to shut down the assembly pathway. We reasoned that if this feedback mechanism is defective, one should expect to see mutant synapses with abnormal vesicle pool. Indeed in a forward genetic screen, we isolated a mutant in which the proximal synapses are abnormally large while the distal synapses contain little material. We have also identified an Arf like GTPase to be responsible for the regulation the location and size of presynaptic specializations. To gain mechanistic insights on the functions of these genes, I specifically plan to understand: 1) how the cyclin-dependent kinases pathways control axonal transport and synapse localization through the regulation of molecular motors, and 2) how an ARF like GTPase, ARL8 regulate the synaptic vesicle pool size and presynaptic assembly. Given that many neural disorders are associated with alterations in synaptic connectivity and that cyclin dependent kinases have been implicated in neurodegenerative diseases, it is hopeful that this project will help to understand brain development under both physiological and pathological conditions.