LINKING ENDOPLASMIC RETICULUM STRESS TO NEURODEVELOPMENTAL DISORDERS

Autism spectrum disorders are a group of neurodevelopmental disorders with a strong genetic background. One of the most characterized autism-linked mutations is the R451C substitution in the synaptic protein Neuroligin3 (NLGN3). The mutation induces a local misfolding in the extracellular domain causing the retention of NLGN3 in the Endoplasmic Reticulum (ER)1. The presence of misfolded protein in the ER can lead to the activation of the Unfolded Protein Response (UPR), implicated in several neurological diseases and in the regulation of neurotransmission and plasticity2. Our aim is to ascertain whether the ER retention of the R451C NLGN3 mutant protein activates the UPR. We have generated a new PC12 Tet-On model system with inducible expression of NLGN3, either wild type or R451C proteins, for studying the UPR signaling in time-course experiments. PC12 clones were characterized for NLGN3 expression, by western blots and immunofluorescence. Wild type NLGN3 protein is correctly trafficked to the cell surface, with the R451C NLGN3 being retained in the ER, as shown by sensitivity to endoglycosidase H. Our results indicate that PC12 clones expressing the R451C mutant NLGN3, activate all UPR signaling pathways downstream of the ATF6, IRE1 and PERK stress sensors. Synthesis of R451C NLGN3 induces the up-regulation of UPR target genes, such as BiP and CHOP, before and after differentiating the cells to a neuronal phenotype. In order to understand the potential role of UPR in neurodevelopmental disorders, we are currently investigating its activation in the Knock In mouse model of autism, carrying the R451C mutation in the NLGN3 endogenous gene. Our data represent the first evidence on the effects of the R451C NLGN3 in activating the UPR and represent a solid link between UPR and neurodevelopmental disorders characterized by the retention of misfolded proteins in the ER.