Autism spectrum disorder (ASD) is a complex and heterogenous neurodevelopmental disease that affects about 1% of children in the United States. ASD is characterized by deficits in verbal communication, impaired social interaction, and limited, repetitive interests. A subset of patients with ASD display early brain overgrowth. We have produced two relevant models that model important aspects of early brain overgrowth in ASD: mouse models deficient for Dvl1 and Dvl3 (Dvl1-/-3+/- mutants) that display adult social behavior abnormalities associated with transient embryonic brain enlargement during deep layer cortical formation; and human neuronal progenitor cells (NPCs) models produced from induced pluripotent stem cells (iPSCs) derived from ASD individuals with early brain overgrowth that displayed enhanced proliferation compared to non-ASD controls. Remarkably, in both models, these aberrant ASD phenotypes were caused by down-regulation of ?- catenin activity and its direct target BRN2. We hypothesize that the ?-catenin/BRN2 transcriptional cascade is a key pathway that exquisitely regulates NPC proliferation and differentiation during brain development of mouse and human, resulting in normal social behavior, while dysregulation results in abnormal social behavior and at least some aspects of ASD. We propose to address key questions posed by this hypothesis.
Aim 1 : Identify the transcriptional targets of ?-catenin and BRN2 in NPCs from iPSCs derived from human patients with and without early brain overgrowth and from Dvl1-/-3+/- mutant mice. To determine specificity of the ?-catenin/BRN2 transcriptional cascade in ASD with early brain overgrowth, we will produce NPCs from iPSCs from: controls and ASD patients with and without early brain overgrowth; patients with PTEN mutations that display macrocephaly with or without ASD; and two ASD lines with corrected Wnt pathway mutations. We will utilize RNA-seq and ChIP-seq combined with gene ontology (GO) analysis to identify downstream pathways that are directly regulated by ?-catenin and BRN2 in mouse NPCs from wild-type and Dvl1-/-3+/- embryos and in human NPCs derived from iPSCs of control and ASD patients.
Aim 2 : Characterize common downstream pathway(s) that are misregulated in human ASD and Dishevelled mouse models. We will intersect the downstream pathways that mediate the effects of the ?- catenin/BRN2 transcriptional cascade in mouse and human NPCs, and select pathways that are commonly dysregulated using novel bioinformatics tools and approaches.
Aim 3 : Validate the disruption of pathways identified in Aim 2 in in NPCs and organoids made from iPSCs from control and ASD patients as well as embryonic brains of Dvl1-/-3+/- mice. We will confirm and validate the disruption of pathways identified in Aim 2 and assess their importance in ASD pathology by perturbing them in our mouse and human-derived NPCs and brain organoid models via genome editing. Mouse mutants for genes/pathways with strong effects will be produced by genome editing to validate their significance in brain overgrowth and social behavior in vivo.
We have identified a novel, conserved ?-catenin/BRN2 transcriptional cascade that regulates embryonic neural progenitor proliferation in mouse models with adult social behavior deficits and brain abnormalities, as well as human induced pluripotent stem cell models of autism. Our main hypothesis is that the ?-catenin/BRN2 transcriptional cascade is a key pathway that exquisitely regulates neural progenitor proliferation and differentiation during brain development of mouse and human, resulting in normal social behavior, while dysregulation results in abnormal social behavior. We propose to address this hypothesis by determining the downstream pathways that are directly regulated by ?-catenin and BRN2 the using genome-wide RNA-seq and ChIP-seq. These studies will advance our understanding of the relationship of the ?-catenin/BRN2 transcriptional cascade to embryonic brain enlargement and autism. Further study of this cascade will likely provide important insights into mammalian development and behavior.