Genetics of New Synaptic Components and Their Functions

描述(申请人提供)：这项建议集中于突触形态的两种类型的变化：突触后膜的扩张和突触前突触的形成。解剖学上的特化是突触的标志。在突触后侧，解剖特化包括树突棘和膜折叠。在突触前一侧，突触突起增大，形成圆形突触前突触或突触前肿胀，这几乎是突触的普遍特征。这项建议的目的1测试了一个特定的假设，即活动如何调节苍蝇神经肌肉接头和哺乳动物树突棘突触后的解剖变化。特别是，它试图阐明ral的作用是 依赖活动的解剖可塑性的中介物。我们发现了一条突触可塑性的新途径，在这条途径中，小的GTP酶Ral通过激活胞外复合体，在将谷氨酸受体中的钙离子内流转导到加强膜到突触后区域的运输中发挥中心作用。因此，苍蝇神经肌肉接头(NMJ)突触后特化的膜面积以一种活动依赖的方式扩大。这项建议提出了什么模式的突触活动是必要的，以招募外囊，以及如何使Ral定位于突触后膜。并进一步探讨了Are在哺乳动物中枢神经系统树突棘形成中的意义。Aim 2使用果蝇作为一个模型系统，在这个系统中揭示了允许突触连接形成的机制。突触前终末从生长锥到突触突触的成熟是突触发生的关键后期阶段，但人们对此知之甚少。既不知道形状变化背后的细胞骨架元素，也不知道触发它的信号分子。在之前对果蝇的突变筛选中，我们发现2-3的突变在最初的突触形成后阻止了突触的形成，并阻止了在果蝇NMJ处突触的形成。我们建议通过结合生化和遗传学的方法来更详细地研究Bouton的形成过程，以发现在突触解剖的这一鲜为人知的方面的形成中的其他角色。因此，在目标2A中，我们将使用生化方法来确定2-3的结合伙伴，并确定它们是否是形成Bouton所必需的。在目标2B中，我们将通过对果蝇基因组在胚胎NMJ的突变筛选，发现更多在Bouton形成中的角色。

DESCRIPTION (provided by applicant): This proposal focuses on two types of change in synapse morphology: expansion of postsynaptic membranes and formation of presynaptic boutons. Anatomical specializations are a hallmark of synapses. On the postsynaptic side, anatomical specializations include dendritic spines and membrane folds. On the presynaptic side, an enlargement of the neurite to form a rounded presynaptic bouton or en passant swelling is a nearly universal feature of synapses. Aim 1 of this proposal tests a specific hypothesis for how activity can regulate anatomical changes at the postsynapse in the fly neuromuscular junction and in mammalian dendritic spines. In particular, it seeks to elucidate the role of Ral as a mediator of activity-dependent anatomical plasticity. We have uncovered a novel pathway for synaptic plasticity in which the small GTPase Ral, by activating the exocyst complex, serves a central role in transducing Ca2+ influx from glutamate receptors into enhanced transport of membrane to the postsynaptic region. In consequence, the membrane area of the postsynaptic specialization at the fly neuromuscular junction (NMJ) expands in an activity-dependent manner. This proposal asks what patterns of synaptic activity are necessary to recruit the exocyst and how Ral comes to be localized to postsynaptic membranes. It goes on to investigate the significance of Ral for the formation of dendritic spines in the mammalian CNS. Aim 2 uses Drosophila as a model system in which to uncover the machinery that allows synaptic boutons to form. The maturation of presynaptic terminals from growth cones to synaptic boutons is a critical late step in synaptogenesis but poorly understood. Neither the cytoskeletal elements that underlie the shape change nor signaling molecules that trigger it are known. In a previous mutant screen in Drosophila we discovered that mutations in 2-3 arrest synaptogenesis after initial synapse formation and prevent the formation of synaptic boutons at the fly NMJ. We propose to pursue the process of bouton formation in greater detail by a combination of biochemical and genetic approaches to uncover additional players in the formation of this poorly understood aspect of synaptic anatomy. Therefore, in Aim 2A we will use biochemical methods to identify binding partners for 2-3 and determine if they are required for bouton formation. In Aim 2B we will uncover additional players in bouton formation through a mutant screen of the Drosophila genome at the embryonic NMJ.