A multidisciplinary approach to study protocadherin-19: from neuronal function to the “cellular interference” pathogenic mechanism

Mutations in the X-chromosome gene PCDH19 cause a female-limited form of infant-onset epilepsy (Epileptic encephalopathy, early infantile, EIEE9) that is associated with intellectual disability and autistic features. PCDH19 encodes protocadherin-19, whose function in the brain remains unknown. EIEE9 is characterized by a peculiar inheritance pattern as it affects females, with the exception of somatic mosaic males. To explain gender differences, a cellular interference model has been proposed but never proven: random chromosome X-inactivation in females leads to tissue mosaicism in which cells expressing wild-type PCDH19 and cells not expressing or expressing a mutant PCDH19 coexist. This tissue mosaicism is proposed to scramble cell-to-cell communication (cellular interference). Recently, we generated a conditional PCDH19 KO mouse. Preliminary in vivo recordings of neuronal activity show that PCDH19 mosaic expression in vivo triggers spontaneous epileptiform activity. In vitro, electrophysiological recordings demonstrated that iPSC-derived neurons depleted of PCDH19 are characterized by an hyperexcitable phenotype. Overall, these data suggest a role of PCDH19 in synaptic connectivity and support its involvement in neuronal circuit formation. We will characterize Pcdh19 conditional mouse brain from a morphological and functional point of view in order to investigate EIEE9 aetiology with a focus on tissue mosaicism; we will characterize the phenotype of patients with PCDH19 mutations in terms of brain excitability, brain connectivity and morphology and we will generate and characterize disease-relevant cellular models for EIEE9 exploiting patient-derived iPSCs and SHED (Human Exfoliated Deciduous teeth). Finally, we will develop new strategies based on CRISPR/Cas9 technology able to rescue the “cellular interference” in vitro and in vivo.