Shared molecular consequences of epigenetic machinery disruption in neuronal progenitors

The Mendelian disorders of the epigenetic machinery (MDEMs) are an emerging cause of intellectual disability and growth abnormalities. Hippocampal neurons have been repeatedly found to be an effected cell type for these disorders. To directly probe the effect of disruption of the epigenetic machinery, we established a murine neuronal hippocampal differentiation model which allows rapid and systemic knockout (KO) of diverse epigenetic machinery (EM) factors. This system allows us to interrogate the consequences of each KO using ONT DNA methylation sequencing and RNA sequencing. Upon KO of KMT2A (a histone methyltransferase) or DNMT1 (a DNA methyltransferase), we observed a striking convergence in gene expression. In contrast, KO of DNMT1 lead to extensive changes in DNA methylation whereas KO of KMT2A had little-to-no effect on DNA methylation. In both models, KO accelerated neuronal differentiation, and the convergence in gene expression appeared linked to this differentiation state. The transcription factor GSX1 was differentially methylated, differentially expressed and ∼20% of its known down-stream targets were differentially expressed genes. Extending our methylation analysis to 46 EM genes, we found that loss of DNA methyltransferases caused the most profound DNAm alterations, whereas other genes induced more subtle effects. Clustering of promoter DNAm changes revealed two EM subgroups that collectively showed a marked enrichment of interactions with the DNAm machinery. Through allele-specific phasing, we identified a differentially methylated region (DMR) at Zic4 that was shared across the entire group of EM KOs, as well as 1074 haplotype-specific DMRs. Together, our results highlight shared mechanisms across diverse EM mutations and provide new insight into the molecular etiology of MDEMs.