Genomic analysis of autism spectrum disorder and functional characterization of rare variants

Autism spectrum disorder (ASD) is a multifactorial neurodevelopmental disorder, characterized by a complex genetic background. Genetic variants involved in ASD etiology can either be common or rare, can range in size from large chromosomal aberrations to point mutations and can be inherited or arise de novo in ASD individuals. The first aim of this Ph.D. project was the investigation of ASD genetic landscape through the genomic analysis rare coding variation in a family-based ASD cohort. The study sample included 116 families, for a total of 435 individuals, 144 of which with ASD, and whole exome and whole genome sequencing were performed in 29 and 105 families respectively. With this approach we were able to produce a molecular diagnosis for 10% of cases and to study the rare inherited and de novo variation present in our cohort. These analyses led, among other results, to the identification of 37 rare de novo potentially damaging sequence variants, affecting both known ASD risk genes and new promising candidates. Another purpose of our investigation was to functionally characterize specific de novo variants in genes of interest. One of the 37 de novo missense potentially damaging variants affected RAB11B, encoding for a Rab GTPase involved in vesicular transport and endocytic recycling, biological processes with a central role in neurodevelopment. In the second aim of this Ph.D. project, the RAB11B missense variant from our cohort was functionally characterized in vitro, alongside two additional missense variants in the same gene reported in literature but not functionally described. The experimental design involved immunofluorescence analyses investigating the localization and the cellular effects of the mutated recombinant RAB11B proteins. The results of the second aim gave us significant insight into the impact of different missense variants in the function of the Rab protein and its potential role in ASD susceptibility.