Evolution and diversity of synaptic transmission: Roles for voltage-gated calcium channels and PDZ-domain mediated scaffolding

突触传递是神经系统功能的核心。在我的研究中，我试图了解突触结构和功能在远亲动物之间的差异，以及突触是如何进化的。事实上，获得对突触及其进化的深入的系统发育理解将有助于拓宽我们对这种复杂而动态的细胞-细胞信号装置的理解，基础科学可以帮助指导未来治疗涉及突触传递缺陷的神经系统疾病的努力。* 有趣的是，最近的基因组测序研究表明，早期分化的缺乏神经系统的“原始”动物，如小型海洋无脊椎动物粘毛虫，拥有突触传递所需的大部分基因。同样有趣的是，已知的最早分化的动物，栉水母，如Mnemiopsis leidyi，具有神经系统和突触传递，似乎是独立进化的。我在分子生物学，电生理学和基因组学/转录组学方面的广泛背景使我和我的工作人员能够利用这些动物来深入了解突触多样性和进化。在这5年的NSERC资助中，我和我的团队将通过完成以下目标来探索这些主题：*1）目标1：电压门控钙（Cav）通道的比较结构-功能和生理学研究，Cav通道在突触传递中起着至关重要的作用，并与许多人类疾病有关。我们将比较人类和其他无脊椎动物的Cav通道与Trichoplax和Mnemiopsis的高度分歧的直系同源物/同源物，以获得一般的见解保守和定义的生物物理特征的每个Cav通道类型，并确定是否Cav通道在Trichoplax和Mnemiopsis中发挥的作用类似于已知的突触功能的Cav通道在其他研究良好的生物体。* 2）目的2：高通量蛋白质组学以评估作为突触支架基础的关键蛋白质-蛋白质相互作用（包括Cav通道与突触前神经递质分泌的关联）是否在Trichoplax中不存在，这与其缺乏突触一致，并定义独立进化的且知之甚少的拟记忆体突触的分子结构。* 3）目标3：Trichoplax中突触的缺失，以及Mnemiopsis独立进化的突触，应该通过关键突触蛋白质-蛋白质相互作用的缺失/差异来反映。我们将使用从目标1和2中收集的信息，为果蝇的体内拯救实验提供信息，使用来自Trichoplax和Mnemiopsis的引入蛋白质相互作用结构域的野生型与“体外进化”的突触基因，以测试有关可能促进突触进化的关键分子创新的假设。

Synaptic transmission lies at the very core of nervous system function. In my research, I seek to understand how synaptic structure and function differs between distantly-related animals, and how the synapse evolved. Indeed, gaining a deep, phylogenetic understanding of the synapse and its evolution will help broaden our understanding of this complex and dynamic cell-cell signaling apparatus, basic science that can help guide future efforts to treat diseases of the nervous system involving deficiencies in synaptic transmission. ***Interestingly, recent genome sequencing studies have revealed that early-diverging “primitive” animals that lack nervous systems, such as the small marine invertebrate Trichoplax adhaerens, harbour a majority of genes required for synaptic transmission. Also interesting is that the most early-diverging animals known, ctenophores such as Mnemiopsis leidyi, bear nervous systems and synaptic transmission that appears to have evolved independently. My broad background in molecular biology, electrophysiology and genomics/transcriptomics uniquely positions me and my personnel to exploit these animals for gaining insights into synaptic diversity and evolution. Over this 5 year NSERC grant, my team and I will explore these subjects by completing the following objectives: ***1) Objective 1: Comparative structure-function and physiology studies of voltage-gated calcium (Cav) channels, which play crucial roles in synaptic transmission and are implicated in numerous human diseases. We will compare human and other invertebrate Cav channels with highly divergent orthologues/homologues from Trichoplax and Mnemiopsis, to gain general insights into conserved and defining biophysical features of each Cav channel type, and to determine whether the roles that Cav channels play in Trichoplax and Mnemiopsis resemble known synaptic functions of Cav channels in other well-studied organisms. ***2) Objective 2: High-throughput proteomics to evaluate if key protein-protein interactions that underlie synaptic scaffolding, including the association of Cav channels with pre-synaptic neurotransmitter secretion, are absent in Trichoplax, in accordance with its lack of synapses, and to define the molecular architecture of the independently evolved and poorly understood synapses of Mnemiopsis. ***3) Objective 3: The absence of synapses in Trichoplax, and the independently evolved ones of Mnemiopsis, should be reflected by absences/differences in key synaptic protein-protein interactions. We will use information gathered from objectives 1 and 2 to inform in vivo rescue experiments in the fruit fly Drosophila, using wildtype vs. “in vitro evolved” synaptic genes from Trichoplax and Mnemiopsis with introduced protein interaction domains, to test hypotheses about key molecular innovations that might have facilitated synapse evolution.**