Restrepo_2019_J.Neurosci_39_9065

Presynaptic alpha-neurexins are highly expressed and more frequently linked to neuropsychiatric and neurodevelopmental disorders than beta-neurexins. However, how extracellular sequences specific to alpha-neurexins enable synaptic transmission is poorly understood. We identified a mutation in an extracellular region of neurexin-3alpha (A687T), located in a region conserved among alpha-neurexins and throughout vertebrate evolution, in a patient diagnosed with profound intellectual disability and epilepsy. We systematically interrogated this mutation using a knockdown-replacement approach, and discovered that the A687T mutation enhanced presynaptic morphology and increased two critical presynaptic parameters: (1) presynaptic release probability, and (2) the size of the readily releasable pool exclusively at excitatory synapses in mixed sex primary mouse hippocampal cultures. Introduction of the mutation in vivo and subsequent analysis in ex vivo brain slices made from male and female mice revealed a significant increase in excitatory presynaptic neurotransmission that occluded presynaptic but not postsynaptic LTP. Mechanistically, neurexin-3alpha(A687T) enhanced binding to LRRTM2 without altering binding to postsynaptic neuroligin-1. Thus, neurexin-3alpha(A687T) unexpectedly produced the first neurexin presynaptic gain-of-function phenotype and revealed unanticipated novel insights into how alpha-neurexin extracellular sequences govern both transsynaptic adhesion and presynaptic neurotransmitter release.SIGNIFICANCE STATEMENT Despite decades of scientific scrutiny, how precise alpha-neurexin extracellular sequences control synapse function remains enigmatic. One largely unpursued avenue to identify the role of precise extracellular sequences is the interrogation of naturally occurring missense mutations. Here, we identified a neurexin-3alpha missense mutation in a compound heterozygous patient diagnosed with profound intellectual disability and epilepsy and systematically interrogated this mutation. Using in vitro and in vivo molecular replacement, electrophysiology, electron microscopy, and structure-function analyses, we reveal a novel role for neurexin-3alpha, unanticipated based on alpha-neurexin knock-out models, in controlling presynaptic morphology and neurotransmitter release at excitatory synapses. Our findings represent the first neurexin gain-of-function phenotype and provide new fundamentally important insight into the synaptic biology of alpha-neurexins.