Elucidating the Cell Type-Specific Gene Regulatory Landscape of the Developing Brain in the Investigation of Autism Spectrum Disorder-Related Genes

Abstract

Autism Spectrum Disorder (ASD) is a complex neurodevelopmental disorder distinguished by a spectrum of symptoms and severity levels, predominantly impacting social engagement and motor behavior. The study of ASD is significantly complicated by its exceptional heterogeneity, involving over 1000 genes and diverse developmental processes. However, the emergence of single-cell RNA sequencing (scRNA-seq) and the structure of Gene Regulatory Networks (GRNs) offer a promising pathway to navigate this heterogeneity. By combining various ASD-related transcripts under one regulatory mechanism, we can gain a more comprehensive understanding of the disorder. Capitalizing on these tools, this study aims to unite diverse ASD-related transcripts under a common regulatory framework, thereby offering a more holistic understanding of the disorder. Specifically, we sought to identify those cell type-specific GRNs in which ASD-related Transcription Factors (TFs) control a disproportionately greater number of unique target gene interactions compared to other cell types throughout neurodevelopment. We call this subset of unique ASD related TF-gene interactions the ‘ASD regulon’. The research identified an enrichment in ASD regulon activity in certain populations of cells within the mouse brain, including Layer 4 and 6 neurons, interneurons, projection neurons, Cajal-Retzius cells, and several glial cell types. We further sought to delineate the effect of time on the activity of ASD regulons by modeling ASD regulon activity as a function of time. The progression of neurodevelopment was found to have a significant effect on the increase in ASD regulon activity in the developing forebrain of mice from prenatal to neonatal period, while a decrease in ASD regulon activity was observed throughout adolescent-adult whole mouse brain development. The study also identified Ctcf, a highly conserved, ubiquitously expressed protein, as a key driver of ASD regulon activity in the enriched cell types. Ctcf was found to orchestrate the expression of ~150-300 genes across the enriched cell types and was solely culpable for the designation of a cell type as ASD regulon enriched. However, these results were not reproducible in humans, highlighting the translational difficulties in investigating ASD in a murine system. The study acknowledges limitations such as the potential overlooking of crucial players in ASD etiology due to the focus on TF driven enrichment in ASD activity. In conclusion, the study developed a GRN reconstruction pipeline that serves as a tool for the investigation of cell type-specific changes in GRNs across the dynamic gene expression landscape of brain development and further identified cell types and time periods which warrant further investigation.