Synaptic neurexin-1 assembles into dynamically regulated active zone nanoclusters

Neurexins are well-characterized presynaptic cell adhesion molecules that engage multifarious postsynaptic ligands and organize diverse synapse properties. However, the precise synaptic localization of neurexins remains enigmatic. Using superresolution microscopy, we demonstrate that neurexin-1 forms discrete nanoclusters at excitatory synapses, revealing a novel organizational feature of synaptic architecture. Synapses generally contain a single nanocluster that comprises more than four neurexin-1 molecules and that also includes neurexin-2 and/or neurexin-3 isoforms. Moreover, we find that neurexin-1 is physiologically cleaved by ADAM10 similar to its ligand neuroligin-1, with similar to 4-6% of neurexin-1 and similar to 2-3% of neuroligin-1 present in the adult brain as soluble ectodomain proteins. Blocking ADAM10-mediated neurexin-1 cleavage dramatically increased the synaptic neurexin-1 content, thereby elevating the percentage of Homer1(+) excitatory synapses containing neurexin-1 nanoclusters from 40-50% to similar to 80%, and doubling the number of neurexin-1 molecules per nanocluster. Taken together, our results reveal an unexpected nanodomain organization of synapses in which neurexin-1 is assembled into discrete presynaptic nanoclusters that are dynamically regulated via ectodomain cleavage.