Höfundar:
Juan Ouyang, Hans Tómas Björnsso, Kaan Okay, Katrín Möller
Introduction: Mutations in epigenetic machinery (EM) components often lead to intellectual disability (ID) and growth retardation. Mendelian disorders of Epigenetic Machinery (MDEMs) are caused by mutations in these EM factors. Recently, our group and others have uncovered specific DNA methylation (DNAm) signature in one of these disorders (Kabuki syndrome). This project aims to uncover DNAm signature for 60 different EM genes in neuronal progenitors (NPCs) and identify if they share overlapping DNAm signatures.
Methods: In order to prioritize the important locations of DNAm abnormalities, we knock out EM genes with CRISPR/Cas9 in primary NPCs and detect the DNA epi-signatures via Nanopore sequencing. We will then overlap the differentially methylated CpG regions (DMRs) and validate the shared ones via CRISPR-interference to identify functional CpGs.
Results: We have identified thousands of DMRs after EM gene knockouts, including Ep300 and Kmt2a. Specifically, hyperDMRs are enriched in gene bodies and promoter regions, whereas hypoDMRs are more located in transcription binding sites and gene bodies. However, not all of the disease-causing EM genes show significant DMRs, suggesting different mechanisms behind the observed phenotypes of those MDEMs. Further work will highlight which of the CpGs have functional consequences.
Conclusions: Our findings suggest that many disease-causing EM genes significantly impact DNAm. Interestingly, genes with large DMRs belong to a group that is predicted to be highly intolerant towards mutations and associates strongly with neurodevelopmental disorders. This study can therefore identify functionally important CpG sites in MDEMs and provide important understanding of the mechanism of these disorders.