Regulation of Synapse Formation and Function by Neurexin-1 Proteolysis

描述（申请人提供）：突触的形成、成熟和功能需要跨突触粘附复合物的精细协调，从而实现突触前和突触后信号机制的集中和控制。特别是，突触前神经素（Nrxns）与其突触后配体（如神经素、lrrtm等）的关联介导了突触功能的几个方面，如突触传递和功能性突触后位点的组装。越来越多的遗传证据表明，编码Nrxn-1的基因杂合缺失与特发性自闭症谱系障碍和精神分裂症的病理机制有关。关于Nrxn-1在发育和成年突触中的功能和调控，我们所知甚少，对其的理解可能会提高我们对神经发育障碍的认识，并揭示新的药理靶点。我在这里提供了初步数据，验证了使用两种新的小鼠模型来研究Nrxn-1，包括Nrxn-1- ha敲入小鼠，它可以改进内源性Nrxn-1的检测，以及Nrxn-1?/？条件敲除小鼠（cKO）小鼠用于功能研究。利用Nrxn-1- ha敲入蛋白，我进一步证明突触活性以金属蛋白酶依赖的方式影响Nrxn-1的加工，这可能代表了Nrxn-含粘附复合物的重要调节机制。为了更好地了解Nrxn-1在突触中的基本功能和调控，我建议确定负责活性依赖性Nrxn-1加工的特定金属蛋白酶和切割位点。然后，我将同时确定Nrxn-1在调节海马突触形成和生理功能方面的基本功能。最后，通过慢病毒在Nrxn-1 / cKO神经元中重新表达野生型、抗分裂型和诱导分裂型Nrxn-1的一系列拯救实验，我将评估Nrxn-1加工在其正常功能中的作用。预计这项拟议的研究将为Nrxn-1缺失如何引起人类神经发育障碍提供基本的见解，并将首次提供细胞外蛋白水解作为调节机制的功能分析

DESCRIPTION (provided by applicant): The formation, maturation, and function of synapses requires the exquisite coordination of transynaptic adhesion complexes, enabling the concentration and control of pre- and postsynaptic signaling machinery. In particular, the association of presynaptic neurexins (Nrxns) with their postsynaptic ligands (e.g. neuroligins, LRRTMs, etc.), mediates several aspects of synaptic function, such as synaptic transmission and the assembly of functional postsynaptic sites. A growing body of genetic evidence implicates heterozygous deletions of the gene encoding Nrxn-1 in the pathoetiology of idiopathic autism spectrum disorders and schizophrenia. Still very little is known regarding the function and regulation of Nrxn-1 at developing and adult synapses, an understanding of which may improve our knowledge of neurodevelopmental disorders and reveal novel pharmacological targets. I present here preliminary data that validates the use of two new mouse models for studying Nrxn-1, including a Nrxn-1-HA knockin mouse which will allow improved detection of endogenous Nrxn-1, as well as a Nrxn-1?/? conditional knockout mouse (cKO) mouse for functional studies. Using the Nrxn-1-HA knockin, I further show that synaptic activity influences the processing of Nrxn-1 in a metalloproteinase-dependent fashion, which may represent an important regulatory mechanism of Nrxn- containing adhesion complexes. In order to better understand the basic function and regulation of Nrxn-1 at synapses, I propose to identify the specific metalloproteinase and cleavage site responsible for activity- dependent Nrxn-1 processing. Then I will determine, in parallel, the basic function of Nrxn-1 in regulating the formation and physiological function of hippocampal synapses. Finally, through a series of rescue experiments achieved through lentiviral re-expression of wild-type, cleavage-resistant, and cleavage-inducible Nrxn-1� in Nrxn-1�/� cKO neurons, I will assess the role of Nrxn-1 processing in its normal function. It is anticipated that the proposed research will provide fundamental insight into how Nrxn-1 deletions give rise to human neurodevelopmental disorders and will offer the first functional analysis of extracellular proteolysis as a regulatory mechanism of neurexin-containing adhesion complexes.