Nanoscale cellular and molecular fingerprints of synaptic diversity

功能性突触的多样性对于传递和编码大脑中的感觉信息至关重要。然而，这种多样性的分子和生物物理基础仍未被探索。我们假设，活性区支架蛋白的数量和位置的纳米级变化，以及控制其融合的分子机制的突触特异性表达，有助于功能性突触多样性。利用小鼠体感觉皮层和小脑，我们将确定负责感觉知觉的抑制性和兴奋性突触（强突触和弱突触）多样性的分子和生物物理机制。在果蝇模型系统中已经发现了许多影响突触行为的候选分子。在这里，我们建议使用突触分子的超分辨率成像来识别它们的结合伙伴，并表征它们的纳米级结构（分子指纹），最终影响钙通道和突触囊泡之间的距离。为了建立结构函数关系，我们将使用光学和生物物理相结合的方法来估计单钮扣的耦合距离。最后，通过基因改变（Crisper/Cas9基因编辑）对关键活性区支架蛋白进行因果实验，确认其在偶联距离设定中的具体作用。我们期望阐明界定整个大脑功能多样性的一般分子规则。由于突触前蛋白参与了几种脑部疾病，以及我们研究的分子性质和跨脑广度，这项研究将为未来的治疗干预提供基础。

Functional synaptic diversity is critical for routing and encoding sensory information in the brain. Yet, the molecular and biophysical underpinnings of such diversity remain unexplored. We hypothesize that nanoscale variations in the number and location of active zone scaffold proteins, as well as synapse-specific expression of the molecular machinery controlling their fusion, contribute to functional synaptic diversity. Using the mouse somatosensory cortex and cerebellum we will identify the molecular and biophysical mechanisms underlying the diversity of inhibitory and excitatory synapses (strong and weak), which are responsible for sensory perception. Many candidate molecules influencing synaptic behavior have been identified in the drosophila model system. Here we propose to use super-resolution imaging of synaptic molecules to identify their binding partners and characterize their nanoscale architecture (molecular fingerprint), which ultimately influences the distance between calcium channels and synaptic vesicles. In order to establish a structure function relationship, we will estimate coupling distances using combining optical and biophysical approaches at single boutons. Finally, causal experiments will be performed by genetic alteration (Crisper/Cas9 gene editing) of key active zone scaffold proteins to confirm their specific role in setting coupling distance. We expect to elucidate general molecular rules defining functional diversity throughout the brain. Because of the involvement of presynaptic proteins in several brain disorders and the molecular nature and cross-brain breadth of our study, this research will provide a foundation for therapeutic future interventions.