Analysis of the R451C Neuroligin3 Knock-In mouse, a model of a monogenic form of autism

Autism Spectrum Disorders (ASDs) are neurodevelopmental syndromes, in which several environmental risk factors act on a vulnerable genetic background. Among genes whose mutations have been associated with ASDs, the R451C substitution in the synaptic protein Neuroligin3 (NLGN3) has been highly characterized. It is known from in vitro studies, that the mutation affects folding of the extracellular domain of the protein, causing its retention in the Endoplasmic Reticulum (ER) and the activation of the Unfolded Protein Response (UPR). It has been shown both in vitro and in vivo, that only ~10% of the mutant protein reach the synapse, causing loss of NLGN3 on the cell surface and leading to alterations in synaptic neurotransmission. In this work, we have evaluated whether UPR was activated in vivo, in the brain of the knock-in mouse model carrying the R451C mutation in the endogenous NLGN3. We showed a selective increase of UPR markers levels in the cerebellum of the R451C mice, along with an increase in the frequency of the miniature excitatory currents in the Purkinje cells, that resulted to be UPR-dependent. At the same time, in order to find a strategy to rescue NLGN3 folding and expression on the cell surface, we have generated and characterized a new cell-based model system that allowed studying NLGN3 protein trafficking. By using this system, we have screened an FDA-approved library of compounds for improving impaired protein folding. Among the compounds that have been tested, several members of the glucocorticoid family showed efficacy in increasing mutant protein trafficking and restoring membrane localization. Collectively, our data indicated that the ER-retention of R451C NLGN3 in vivo, caused UPR activation and alterations of synaptic function in the cerebellum of a mouse model of a monogenic form of autism. Furthermore, we identified compounds improving NLGN3 folding and rescuing impaired trafficking.