Isotani, A. | Search bioRxiv

bioRxiv2021

Inhibition of N-myristoyltransferase Promotes Naive Pluripotency in Mouse and Human Pluripotent Stem Cells

Naive and primed states are distinct states of pluripotency during early embryonic development that can be captured and converted to each other in vitro. To elucidate the regulatory mechanism of pluripotency, we performed a recessive genetic screen of homozygous mutant mouse embryonic stem cells (mESCs) and found that suppression of N-myristoyltransferase (Nmt) promotes naive pluripotency. Disruption of Nmt1 in mESCs conferred resistance to differentiation. Suppression of Nmt in mouse epiblast stem cells (mEpiSCs) promoted the conversion from the primed to the naive state. This effect was independent of Src, which is a major substrate of Nmt and is known to promote differentiation of mESCs. Suppression of Nmt in naive-state human induced pluripotent stem cells (hiPSCs) increased the expression of the naive-state marker. These results indicate that Nmt is a novel target for the regulation of naive pluripotency conserved between mice and humans.

developmental biology↗

bioRxiv2020

Ets2 frame-shift mutant models express in-frame mRNA by exon skipping that complements Ets2 function in the skin

The Ets2 transcription factor has been implicated in various biological processes. An Ets2 mutant model, which lacks the DNA-binding domain (ETS domain), was previously reported to exhibit embryonic lethality caused by a trophoblast abnormality. This phenotype could be rescued by tetraploid complementation, resulting in pups with wavy hair. Here, we generated new Ets2 mutant models with deletions in exon 8 and with frame-shift mutations using the CRISPR/Cas9 method. Homozygous mutants could not be obtained by natural mating as previously reported. After rescuing with tetraploid complementation, homozygous mutant mice were generated, but these mice did not exhibit wavy hair phenotype. Our newly generated mice exhibited exon 8 skipping, which led to in-frame mutant mRNA expression in the skin and thymus but not in E7.5 embryos. As this in-frame mutation contained the ETS domain, the exon 8-skipped Ets2 mRNA was likely translated into protein in the skin that complemented the Ets2 function. Thus, these Ets2 mutant models, depending on the cell types, exhibited novel phenotypes due to exon skipping and are expected to be useful in several fields of research. Summary statementNew Ets2 mutant models showed embryonic lethal phenotype by a placental abnormality but did not exhibit a wavy hair phenotype as a previous model.

developmental biology↗