Functional analysis of asd genes in zebrafish

A central challenge in the genetics of ASD is advancing from gene discovery to the elucidation of biologically relevant mechanisms. The objectives of this research are to analyze the function of multiple ASD-associated genes in the developing vertebrate brain and to identify common pathways uniting these genes at the behavioral, brain structural, circuit, and molecular levels. We conducted parallel in vivo analyses of 10 high-confidence ASD genes at the behavioral, brain structural, and circuit levels in zebrafish. Using CRISPR/Cas9, we generated frameshift mutations in the zebrafish orthologs of ASD genes. Next, we performed automated assays of arousal and sensory processing behaviors and whole-brain structural and circuit analyses in zebrafish ASD gene mutants. Whole-brain RNA-sequencing was conducted in 2 ASD gene zebrafish mutants displaying the most robust phenotypes. We found that zebrafish mutants of ASD genes display a range of convergent and divergent phenotypes at the behavioral, structural, and circuit levels. Whole-brain analyses identify the forebrain (telencephalon) as the most significant contributor to brain size differences across ASD genes and that regions involved in sensory-motor control are associated with differences in baseline brain activity. Whole-brain RNA-sequencing of 2 ASD gene mutants identifies dysregulation of highly conserved ASD gene-associated pathways. Our study identifies both unique and shared effects of ASD gene loss of function in the developing vertebrate brain. We find evidence for points of biological convergence at the brain structural, activity, and molecular levels, revealing pathways relevant to ASD biology.