DNMT3B supports meso-endoderm differentiation from mouse embryonic stem cells

Abstract

The correct establishment of DNA methylation patterns during mouse early development is essential for cell fate specification. However, the molecular targets as well as the mechanisms that determine the specificity of the de novo methylation machinery during differentiation are not completely elucidated. Here we show that the DNMT3B-dependent DNA methylation of key developmental regulatory regions at epiblast-like cells (EpiLCs) provides an epigenetic priming that ensures flawless commitment at later stages. Using in vitro stem cell differentiation and loss of function experiments combined with high-throughput genome-wide bisulfite-, bulk-, and single cell RNA-sequencing we dissected the specific role of DNMT3B in cell fate. We identify DNMT3B-dependent regulatory elements on the genome which, in Dnmt3b knockout (3BKO), impair the differentiation into meso-endodermal (ME) progenitors and redirect EpiLCs towards the neuro-ectodermal lineages. Moreover, ectopic expression of DNMT3B in 3BKO re-establishes the DNA methylation of the master regulator Sox2 super-enhancer, downmodulates its expression, and restores the expression of ME markers. Taken together, our data reveal that DNMT3B-dependent methylation at the epiblast stage is essential for the priming of the meso-endodermal lineages and provide functional characterization of the de novo DNMTs during EpiLCs lineage determination.

TDG orchestrates ATF4-dependent gene transcription during retinoic acid-induced cell fate acquisition

“…Differentiation skewing towards cardiac mesoderm has also been observed in vivo in TET mutants (Li et al, 2016), suggesting a common requirement of various components of the DNA demethylation machinery for neural differentiation. Consistent with the idea that DNA demethylation associates with neural differentiation, the knock-out of the DNA methyltransferase gene Dnmt3b in mESCs inversely skews differentiation toward neurectoderm (Lauria et al, 2023). Although a number of differentially methylated regions (DMRs) observed in EpiSC-like cells derived from Dnmt3b-KO ESCs (Lauria et al, 2023) were found to overlap with the TSSs of genes repressed in TDG-null ECCs, they also overlapped in a similar proportion with TSSs of activated genes (data not shown).…”

WENDY: Covariance Dynamics Based Gene Regulatory Network Inference

“…2 for the GRNs at different time points. We can see that initially DNMT3L (catalytically inactive regulatory factor of DNA methyltransferases) [62] and DNMT3B (establishes DNA methylation patterns in embryos) [36] are major targets of regulation. As time increases, many regulatory effects show a weakening in the latter three time points, relative to the initial state of the GRNs.…”

ZFP281 coordinates DNMT3 and TET1 for transcriptional and epigenetic control in pluripotent state transitions

“…However, the two proteins are known to have unique functions in ESCs and early development. 27,33,48,49 Future studies are needed to distinguish the functions and targets of DNMT3A versus DNMT3B (and possibly the isoforms associated with each gene) in different PSCs and during pluripotent state transitions. In addition, DNMT1 was reported to possess de novo DNA methylation activity in mouse oocytes, 50 13 and indicated with dashed inhibition lines, would lead to downregulation of ZFP281-regulated genes including Dnmt3a/3b (and thus reduced 5mC levels), which in turn relieves TET1 from the competition, leading to heightened TET1 chromatin binding (and thus increased 5hmC levels).…”

“…However, the two proteins are known to have unique functions in ESCs and early development. 27, 33, 48, 49 Future studies are needed to distinguish the functions and targets of DNMT3A versus DNMT3B (and possibly the isoforms associated with each gene) in different PSCs and during pluripotent state transitions. In addition, DNMT1 was reported to possess de novo DNA methylation activity in mouse oocytes, 50 but we can only speculate that DNMT1 may compensate for the loss of de novo DNA methylation activities in establishing Dnmt3a/3b- DKO cEpiSCs.…”