Loss of DNA methyltransferase activity in primed human ES cells triggers increased cell-cell variability and transcriptional repression

Abstract

Maintenance of pluripotency and specification towards a new cell fate are both dependent on precise interactions between extrinsic signals and transcriptional and epigenetic regulators. Directed methylation of cytosines by the <i>de novo</i> methyltransferases DNMT3A and DNMT3B plays an important role in facilitating proper differentiation, whereas DNMT1 is essential for maintaining global methylation levels in all cell types. Here, we generated single-cell mRNA expression data from wild-type, DNMT3A, DNMT3A/3B and DNMT1 knockout human embryonic stem cells and observed a widespread increase in cellular and transcriptional variability, even with limited changes in global methylation levels in the <i>de novo</i> knockouts. Furthermore, we found unexpected transcriptional repression upon either loss of the <i>de novo</i> methyltransferase DNMT3A or the double knockout of DNMT3A/3B that is further propagated upon differentiation to mesoderm and ectoderm. Taken together, our single-cell RNA-sequencing data provide a high-resolution view into the consequences of depleting the three catalytically active DNMTs in human pluripotent stem cells.