Alpha-neurexins and many of the proteins they recruit are implicated in neuropsychiatric diseases including schizophrenia, autism spectrum disorder, and mental retardation - diseases that are in desperate need of better medications. Alpha-neurexins are synaptic organizers involved in neuronal communication, and are diversified though small stretches of amino acids called splice inserts. It is not known how alpha-neurexins utilize their molecular features to bind a diverse array of partners in the synaptic cleft includin neuroligins, leucine rich repeat transmembrane proteins (LRRTMs), neurexophilins, GABAA-receptors, and likely as yet unidentified proteins as well. It is important to understand the molecular mechanisms that enable alpha-neurexins to bind their diverse partners, because when these interactions are disrupted, fundamental biological processes are altered that are thought to contribute to the pathology of many severe mental disorders. The long term goal of our laboratory is to understand on a molecular level how proteins in the synaptic cleft integrate into highly organized protein interaction networks that form and maintain functional synapses. The objective of this particular application is to reveal how the family of alpha-neurexins works as synaptic organizers to assemble diverse proteins into distinct protein complexes at different synapses. The central hypothesis is that alpha-neurexins exploit their unique molecular features to generate a portfolio of distinct and plastic binding sites. Firstly, we hypothesize that alpha-neurexins use their characteristic nine domain extracellular region to create a molecular scaffold that spatially organizes proteins in the synaptic cleft. Secondly, we hypothesize that alpha-neurexins generate distinct binding surfaces using splice insert-dependent and splice insert-independent molecular frameworks. This hypothesis is supported by strong preliminary data presented by the applicant entailing structural studies, proteomic data, biophysical data using a new method to study molecular interactions, as well as biochemical and cell-based techniques. The hypothesis will be tested by pursuing three specific aims: 1) delineate the molecular features that enable protein partners to bind in a splice insert-dependent, alpha-neurexin-dependent manner; 2) delineate the molecular features that enable protein partners to bind in a splice insert-independent, alpha-neurexin- dependent manner; and finally 3) establish the binding mode of a new partner that we have identified specific for alpha-neurexins. The rationale for this proposal is that the results will reveal how alpha-neurexins organize different molecular assemblies in the synaptic cleft which take part in biological processes involved in severe neuropsychiatric diseases. The proposal is innovative because it provides a starting point to design strategies to manipulate alpha-neurexin interactions in the synaptic cleft using small molecule compounds or biologics. This information is very significant because it could reveal completely new drug targets to reverse pathological effects of neuropsychiatric disorders and create completely new strategies to treat these devastating disorders. PUBLIC HEALTH RELEVANCE: Alpha-neurexins, a large family of synaptic organizers, have recently been implicated in neuropsychiatric diseases, including schizophrenia and autism spectrum disorder. There is growing consensus that alpha- neurexins, but also many of their protein partners, contribute to biological pathways that are disrupted in many mental disorders. The proposed research will reveal the molecular frameworks used by alpha- neurexins to recruit many different proteins into different multi-protein complexes in the synaptic cleft with different functions. This research is relevant to public health and the mission of NIMH, because detailed molecular knowledge of alpha-neurexin:protein partner interactions could provide new strategies to manipulate neurexin interactions within the synaptic cleft, in order to ameliorate behavioral deficits associated with neuropsychiatric disorders.