Project description:Multiple human autism risk genes are predicted to converge on the β‐catenin (β‐cat)/Wnt pathway. However, direct tests to link β‐cat up‐ or down‐regulation with autism are largely lacking, and the associated pathophysiological changes are poorly defined. Here we identify excessive β‐cat as a risk factor that causes expression changes in several genes relevant to human autism. Our studies utilize mouse lines with β‐cat dysregulation in forebrain excitatory neurons, identified as cell types with convergent expression of autism‐linked genes in both human and mouse brains. We show that mice expressing excessive β‐cat display behavioral and molecular changes, including decreased social interest, increased repetitive behaviors, reduced parvalbumin and altered expression levels of additional genes identified as potential risk factors for human autism. These behavioral and molecular phenotypes are averted by reducing β‐cat in neurons predisposed by gene mutations to express elevated β‐cat. Using next-generation sequencing of the prefrontal cortex, we identify dysregulated genes that are shared between mouse lines with excessive β‐cat and autism‐like behaviors, but not mouse lines with reduced β‐cat and normal social behavior. Our findings provide critical new insights into β‐cat, Wnt pathway dysregulation in the brain causing behavioral phenotypes relevant to the disease and the molecular etiology which includes several human autism risk genes.
Project description:Using RNA-seq to sequence the transcriptomes of β-cat lox(ex3) (Ctrl) and β-cat lox(ex3);Nex-Cre (β-cat Ovp) mice, we did not find significant change in the expression level of classical Wnt target genes.
Project description:To compare total RNA levels in miR-124 and mock-transfected cells (Figure S3), 5-10 ug of total RNA from miR-124-transfected cells or mock-transfected cells or universal reference RNA (Stratagene Cat# 740000) was reverse transcribed with Superscript III (Invitrogen Cat# 18080085) in the presence of aminoallul-dUTP 5-(3-aminoallyl)-dUTP (Ambion Cat# AM8439) and natural dNTPs (GE Healthsciences Cat# US77212) with 10 ug of N9 primer (Invitrogen). Subsequently, amino-allyl-containing cDNAs from miR-124 and mock-transfected cells were covalently linked to Cy5 NHS-monoesters, and universal reference cDNA was covalently linked to Cy3 NHS-monoesters (GE HealthSciences Cat# RPN5661). Cy5- and Cy3-labeled cDNAs were mixed and diluted into 50 ul of solution containing 3x SSC, 25 mM Hepes-NaOH (pH 7.0), 20 ug human Cot-1 DNA (Invitrogen Cat# 15279011), 20 ug of poly(A) RNA (Sigma Cat# P9403), 25 ug of yeast tRNA (Invitrogen Cat# 15401029), and 0.3% SDS. The sample was incubated at 95 C for 2 min, spun at 14,000 rpm for 10 mins in a microcentrifuge, then hybridized at 65 C
Project description:To understand the function of Wnt/β-catenin signaling-activated cardiomyocytes (β-cat ON CMs), we performed RNA-Seq on β-cat ON and OFF CMs at the atrioventricular canal (AVC) in the heart, respectively. Our analyses indicated that β-cat ON CMs upregulate genes related to blood vessel development.
