Project Summary Autism spectrum disorder (ASD) is a neurodevelopmental disorder that affects 1 in 54 children and exhibits significant phenotypic and genetic heterogeneity. Mutations in UBE3B, which encodes an E3 ubiquitin ligase, have been identified in patients presenting with intellectual disability, lack of speech, and ASD. The specific mechanism through which disruption of UBE3B, and subsequent dysregulation of its substrates, leads to neurodevelopmental abnormalities is unknown. Our group has previously shown that a Ube3b constitutive knockout mouse model exhibits a complete loss of vocalization and defects in nest building, as well as reduced dendritic complexity, length, and spine density. In this proposal, I will investigate the neuronal function of UBE3B by characterizing changes in neuronal activity following loss of UBE3B and identifying its neuronal substrates. I will apply complimentary electrophysiological and biochemical approaches using a brain-specific conditional Ube3b knockout mouse model (cKOnestin). I will characterize the electrophysiological properties of cortical neurons from cKOnestin mice, by evaluating their basal properties, including intrinsic excitability and rheobase current, and both short-term and long-term synaptic plasticity. Furthermore, I will identify the neuronal substrates of UBE3B through analysis of protein levels, protein-protein interactions, and ubiquitination status. I will also assess the UBE3B-mediated modifications of substrates, including the site of ubiquitination and the ubiquitin chain topology. Successful completion of the proposed aims will provide new insights into the interplay of ubiquitin signaling and neurodevelopment, as well as advance our knowledge of the specific pathogenic mechanisms underlying neurodevelopmental disorders. The exceptional research environment at UT Southwestern Medical Center, combined with the collective expertise of the mentorship team, will provide excellent training. Goals for this fellowship training include becoming knowledgeable in the relevant literature and attaining proficiency in both experimental procedures and communicating scientific information to varied audiences. The skills obtained through these goals will build the foundation for an independent and successful lead investigator in scientific research.