NLGs的翻译后加工对神经突触发育与功能的调控研究

Synapses are highly specialized intercellular junctions that allow one neuron to communicate with another neuron or a target cell. They are basic units of information processing in huge neural networks of the brain. Synaptic adhesion molecules, NRXs and NLGs, connect presynaptic and postsynaptic neurons at synapses, mediate transsynaptic signal, and shape the properties of neural networks by specifying synaptic functions, however, the molecular mechanism remains to be elucidated. In previous and unpublished preliminary data on Drosophila Nlgs and Nrx from our laboratory, we found that DNlgs protein undergoes post-translational processing and this phenomenon is associated with Drosophila activity, suggesting that Nlgs not only serve as mediator of synaptic scaffold molecule assembly involving transsynaptic complex formation and synapse development, but may also act as non-synaptic neurotransmitter signaling molecules in synapse development and modulation following specific cleavage. Based on these data, here we propose that the proteolytic fragments of Nlgs transduce signals in synapse development and behavior. In this proposal, we will use Drosophila and mice as animal models, apply a combination of genetic, molecular, behavioral and electrophysiological analysis as well as live cell imaging, to further investigate the mechanisms underlying regulation of synaptic adhesion complex in vivo, to track the fate and function of extracellular and intracellular domains of NLGs generated by proteolysis, to analyze different roles of DNlgs in neural development and synaptic functions in vivo undergoing specific post-translational processing. Taken together, we would provide a general paradigm for the regulation of trans-synaptic signaling in different neural circuits, and valuable information for an insight into pathogenic mechanisms of autism spectrum disorders and other cognitive diseases.

突触是神经元之间建立联系、行使功能的高度特化的胞间连接，是构成脑神经信息处理和加工的巨大神经网络的基本单位。突触粘附分子NRXs和NLGs，介导突触前、后分子的募集和组装，优化神经网络，参与脑中所有的信息处理和加工过程，但机制不清。我们前期在研究果蝇DNlgs和DNrx中发现DNlgs存在蛋白翻译后加工现象与行为关联，提示Nlgs不仅以支架分子介导突触分子的募集和组装进而参与突触的形成，而且可能还作为信号分子通过特异性切割参与非神经递质的跨突触信号传递。本研究将综合应用动物模型和生物技术，进一步研究突触粘附复合体动态调控机制和在体功能，追踪NLG分子经切割产生的胞外段和胞内段的命运，探测它们是否发挥特定的信号传递作用，解析不同Nlgs分子的特异性加工的在体分子机制及其在神经发育和功能中的作用，阐明不同的神经环路中跨突触信号调节的可能范式，也为儿童孤独症的发病机制研究提供有价值的信息。

突触是神经元之间建立联系、行使功能的高度特化的胞间连接，是构成脑神经信息处理和加工的巨大神经网络的基本单位。突触粘附分子NRXs和NLGs，介导突触前、后分子的募集和组装，优化神经网络，参与脑中所有的信息处理和加工过程，但机制不清。我们前期在研究果蝇DNlgs和DNrx中发现DNlgs存在蛋白翻译后加工现象与行为关联，提示Nlgs不仅以支架分子介导突触分子的募集和组装进而参与突触的形成，而且可能还作为信号分子通过特异性切割参与非神经递质的跨突触信号传递。本研究综合应用动物模型和生物技术，研究了突触粘附分子翻译后剪切的生物学意义。研究显示，果蝇Nlgs分子切割发生在细胞内不同部位，与啮齿类在细胞膜表面受活性调节的切割不同；剪切位点有构象依赖的（如DNlg2）、和氨基酸序列依赖的（如DNlg3）；剪切有组织特异性和非组织特异性形式，都与Nlg成熟与转运有关，是突触发育过程中的重要环节；啮齿类的Nlg1经过活性依赖性蛋白质切割突触膜表面片段后，胞内段进一步受到beta分泌酶的剪切产生的小肽段通过与SPAR结合，影响RAP1、LIMK及Cofilin活性，进而对树突棘形态及突触可塑性起调节作用；Nlgs调节突触发育与功能（包括突触囊泡释放调节）主要通过ACTIN途径，机制有所不同；WIRS基序有无，决定其是否通过WRC途径调节突触骨架，这种进化产生的多样性可能是维持突触结构与功能稳态的重要原因；发现与自闭症相关的Nlg点突变所致氨基酸变化阻止了Nlg蛋白的准确加工，影响突触发育与功能是发病机制之一。综上，Nlgs蛋白质翻译后加工在突触形成与功能中起重要作用。