Molecular mechanisms of synapse assembly and function

描述（由申请人提供）：突触功能异常是神经系统疾病的标志，从自闭症到精神健康障碍再到神经变性。突触活性区（Synaptic active zone, AZs）是调节神经递质释放的专门部位。释放动力学在AZs之间有显著差异，并根据神经活动进行调节。尽管调节神经递质释放对神经系统功能的重要性，但我们对AZs如何组织以获得精确的释放特性以及它们如何根据活动进行重组的理解仍然有限。我们最近发现了Fife，一种无脊椎短笛同系物。Piccolo先前被认为是无脊椎动物基因组中缺失的一种AZ蛋白，它通过与AZ蛋白的多重连接来调节释放动力学。果蝇是研究短笛/短笛功能的理想模型，因为它缺乏大管和相关的基因冗余，这些基因冗余已经减缓了哺乳动物系统的进展。我们产生了无效的life等位基因，并发现了life在AZ组织中的关键作用，神经递质

DESCRIPTION (provided by applicant): Aberrant synaptic function is a hallmark of neurological diseases ranging from autism to mental health disorders to neurodegeneration. Synaptic active zones (AZs) are specialized sites for the regulated release of neurotransmitter. Release dynamics vary significantly between AZs and are modulated in response to neural activity. Despite the importance of regulated neurotransmitter release to nervous system function, our understanding of how AZs are organized to achieve precise release properties and how they are reorganized in response to activity remains limited. We recently identified Fife, an invertebrate Piccolo homolog. Piccolo, previously believed absent from invertebrate genomes, is an AZ protein hypothesized to regulate release dynamics through its multiple connections to AZ proteins. Drosophila is an ideal model for studying Fife/Piccolo function as it lacks Bassoon and the associated genetic redundancy that has slowed progress in mammalian systems. We generated null fife alleles and found a critical role for Fife in AZ organization, neurotransmitter release and behavior. Here, we test the model, consistent with our functional data and vertebrate biochemical data, that Fife promotes neurotransmitter release by organizing Ca2+ channels and synaptic vesicles (SVs) and in close proximity for reliable Ca2+- dependent exocytosis (Aim 1). A key advantage of the Drosophila model is the ability to rapidly progress to in vivo functional tests of mechanistic models based on protein interactions. We have identified components of the cAMP-signaling pathway as Fife interacting proteins in tandem mass spectrometry (MS/MS) experiments, and propose experiments to functionally test the in vivo significance of these interactions in synaptic plasticity (Aim 2A and B). Finally, we propose a novel, highly efficient genetic screen that takes advantage of neuronally derived female sterility in fife mutants to identify functional interactors that may be inaccessible to proteomic identification (Aim 2C).