Project Summary: The proposed R00 phase of this project will take place in the Wells Laboratory, which opened in September 2021 as part of the Department of Human Genetics in the David Geffen School of Medicine at the University of California Los Angeles. Recent reports estimate that 1 out of every 6 children in the United States meet the diagnostic criteria for neurodevelopmental disorders such as autism spectrum disorders (ASD), attention-deficit hyperactivity disorder (ADHD), and intellectual disability (ID). The prevalence of ASDs, which are characterized by persistent social impairments, language deficits, and repetitive behaviors, has increased by 120% over the past 15 years, a problem further exacerbated by the fact that the disease mechanisms underlying ASDs are largely unknown and no targeted therapeutic interventions exist. Recent progress in human genome sequencing has begun to illuminate pathways to disease through the identification of several genetic risk factors, the most common of which is the deletion of 16p11.2 locus (16p11.2del). Initial studies have nominated specific genes in the 16p11.2 locus in neuronal dysfunction. This proposal aims to elucidate the disease mechanisms underlying 16p11.2del phenotypes using in vitro induced pluripotent stem cell (iPSC)-derived human brain cells. In the first aim, we will attempt to identify the 16p11.2 genes contributing to disease-relevant molecular and phenotypic defects using a pooled CRISPR activation approach in a neural progenitor cell village composed of dozens of patient and neurotypical control lines. In the second aim, we will assess and rescue abnormal molecular and cellular responses to major signaling pathway activation in a village of patients and controls. The successful completion of these aims could lead to the identification of genetic targets for therapeutic intervention, while also dramatically changing the way the field conducts in vitro modeling of human brain disorders.