Spatiotemporal analysis of autism gene enrichment implicates cortex, thalamus, and hypothalamus.

Autism spectrum disorder (ASD) is a highly heritable neurodevelopmental disorder. Sequencing analyses have identified 185 ASD-associated genes, which implicate neurons, but the specific brain regions through which these neurons influence neurodevelopment remain unclear. Here, we integrate over one million single-cell RNA sequencing profiles from 20 regions of the developing human brain (4-23 post-conceptual weeks) using a new framework, STARMAPS (Sparse Task-specific Analysis for Revealing Molecular Associations in Particular Single-cell datasets). STARMAPS accounts for coordinated regional and developmental perturbations in gene expression, enabling robust cross-region comparison. We replicate prior findings that ASD-associated genes are enriched in excitatory neurons during mid-fetal development, and we extend these results to reveal distinct spatial signatures. Across 26 excitatory neuron subtypes, six clusters showed significant enrichment for ASD-associated genes. These clusters localize to both cortical and subcortical regions, including the motor, temporal, and visual cortex, as well as the thalamus and hypothalamus. Our findings support a major role for excitatory neurons across distributed brain circuits, implicating previously underappreciated subcortical structures in ASD etiology. By providing a statistically rigorous framework for spatiotemporal integration of single-cell data, STARMAPS enables refined mapping of molecular vulnerability across the developing human brain.