SETDB1 Loss Dysregulates Translation-Related Pathways and Autism-Associated Gene Networks in Mouse Embryonic Stem Cells

Autism spectrum disorder (ASD) has a strong genetic basis, yet the role of epigenetic regulators in its pathogenesis remains poorly understood. SETDB1, an H3K9 methyltransferase critical for heterochromatin silencing, has been implicated in neurodevelopment, but its impact on ASDassociated gene expression has not been systematically characterized. Here, we generated conditional Setdb1 knockout mouse embryonic stem cells (mESCs) and found that SETDB1 loss causes cell death and loss of pluripotency. RNAseq analysis revealed widespread transcriptional changes, with 1,886 upregulated and 2,170 downregulated genes (|log₂FC| ≥ 1, padj < 0.05). Notably, 266 highconfidence ASD risk genes were significantly enriched among these differentially expressed genes (hypergeometric test, p < 0.001), and these genes were primarily associated with synapse organization, chromatin remodeling, ion transport, and nervous system development. Gene set enrichment analysis uncovered a striking global downregulation of translation and ribosome biogenesis pathways (NES < –1.8, FDR < 0.05), suggesting that impaired protein synthesis contributes to the observed cell death. Proteinprotein interaction network analysis identified a highly interconnected hub gene set (e.g., Actb, Mapt, Syngap1), most of which were upregulated in KO cells, whereas Syp was downregulated. Collectively, our results identify SETDB1 as an essential safeguard of mESC survival and a key regulator of ASDlinked gene networks, providing a valuable transcriptomic resource for understanding the epigenetic control of pathways relevant to neurodevelopmental disorders.