In a second line of research, we demonstrated a multi-step model of synapse assembly mediated by the adhesion complex neurexin/neuroligin. Our team first developed a novel method to probe the kinetic rates of ligand/receptor interactions, based on counting single neurexin1β-conjugated Quantum Dots on neurons expressing neuroligin-1, revealing long lasting bonds between neurexin and neuroligin (Saint-Michel et al., Biophys J. 2009).
Second, the use of neurexin-coated microspheres, allowed us to show that AMPARs, mediating the fast component of excitatory transmission at glutamatergic synapses, could be recruited at neurexin/neuroligin contacts independently of synaptic activity, challenging the classical view that NMDARs had to be present first at nascent synapses and activated by pre-synaptic inputs (Heine et al., PNAS 2008). This study also resolved the fact that the activity-dependent recruitment of AMPARs was dependent on their subunit composition.
Third, using single nano-particle tracking and a series of molecular constructs (mutated neuroligins, shRNA to neuroligin-1 and PSD-95, cultures and slices from neuroligin-1 KO mice), we showed that AMPARs were recruited at PSD-95 scaffolds through surface diffusion and trapping (Mondin et al., J. Neurosci. 2011). Such rapid stabilization of a post-synaptic scaffold by neurexin/neuroligin adhesion could represent a critical molecular step in synapse initiation at nascent axon-dendrite contacts, and point to a new role of AMPARs in synaptogenesis. Overall, our results show distinctive roles of N-cadherin and neurexin/neuroligin adhesions at the synapse, the former related to morphological plasticity in connection to actin motility, the later being associated to the recruitment of scaffolding and functional elements (PSD-95 and AMPA receptors).
Fourth, using neurexin-1 clusters, single neuroligin molecule tracking, and biochemistry, we revealed for the first time a signaling mechanism associated with neurexin/neuroligin adhesion. Specifically, we demonstrated that neurexin-1β binding to neuroligin-1 triggers tyrosine phosphorylation of neuroligin-1 and induces the differential recruitment of PSD-95 versus gephyrin, thereby potentially controlling the balance between excitatory and inhibitory synapses (Giannone et al., Cell Reports 2013).