Missense variants in RPH3A cause defects in excitatory synaptic function and are associated with a clinically variable neurodevelopmental disorder
Lisa Pavinato, Jennifer Stanic, Marta Barzasi, Antonia Gurgone, Giuseppe Chiantia, Valentina Cipriani, Ivano Eberini, Luca Palazzolo, Mónica Di Luca, Alex Costa, Andrea Marcantoni, Elisa Biamino, Marco Spada, Susan M. Hiatt, Whitley V. Kelley, Letizia Vestito, Sanjay M. Sisodiya, Stéphanie Efthymiou, Prem Chand, Rauan Kaiyrzhanov, Alessandro Bruselles, Simona Cardaropoli, Marco Tartaglia, Silvia De Rubeis, Joseph D. Buxbaum, Damian Smedley, Giovanni Battista Ferrero, Maurizio Giustetto, Fabrizio Gardoni, Alfredo Brusco
Abstract
Purpose RPH3A encodes a protein involved in the stabilization of GluN2A subunit of NMDA-type glutamate receptors at the cell surface, forming a complex essential for synaptic plasticity and cognition. We investigated the effect of variants in RPH3A in patients with neurodevelopmental disorders (NDDs). Methods By using trio-based exome sequencing, GeneMatcher, and screening of 100,000 Genomes Project data, we identified six heterozygous variants in RPH3A. In silico and in vitro models, including rat hippocampal neuronal cultures, have been used to characterize the effect of the variants. Results Four cases had a NDD with untreatable epileptic seizures [p.(Gln73His)dn; p.(Arg209Lys); p.(Thr450Ser)dn; p.(Gln508His)], and two cases [p.(Arg235Ser); p.(Asn618Ser)dn] showed high-functioning autism spectrum disorder (ASD). Using neuronal cultures, we demonstrated that p.(Thr450Ser) and p.(Asn618Ser) reduce the synaptic localization of GluN2A; p.(Thr450Ser) also increased the surface levels of GluN2A. Electrophysiological recordings showed increased GluN2A-dependent NMDAR currents for both variants, and alteration of postsynaptic calcium levels. Finally, expression of the Rph3AThr450Ser variant in neurons affected dendritic spine morphology. Conclusion Overall, we provide evidence that missense gain of function variants in RPH3A increase GluN2A-containing NMDARs at extrasynaptic sites, altering synaptic function and leading to a clinically variable neurodevelopmental presentation ranging from untreatable epilepsy to ASD.