2i Maintains a Naive Ground State in ESCs through Two Distinct Epigenetic Mechanisms

Mouse embryonic stem cells (ESCs) are maintained in serum with leukemia inhibitory factor (LIF) to maintain self-renewal and pluripotency. Recently, a 2i culture method was reported using a combination of MEK inhibition (MEKi) and GSK3 inhibition (GSK3i) with LIF to maintain ESCs in a naive ground s...

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Bibliographic Details
Published in:	Stem cell reports Vol. 8; no. 5; pp. 1312 - 1328
Main Authors:	Sim, Ye-Ji, Kim, Min-Seong, Nayfeh, Abeer, Yun, Ye-Jin, Kim, Su-Jin, Park, Kyung-Tae, Kim, Chang-Hoon, Kim, Kye-Seong
Format:	Journal Article
Language:	English
Published:	United States Elsevier Inc 09-05-2017 Elsevier
Subjects:	Animals Cell Line DNA (Cytosine-5-)-Methyltransferases - genetics DNA (Cytosine-5-)-Methyltransferases - metabolism DNA Methylation Epigenesis, Genetic G9a Glycogen Synthase Kinase 3 - antagonists & inhibitors Glycogen Synthase Kinase 3 - metabolism GSK3i Induced Pluripotent Stem Cells - cytology Induced Pluripotent Stem Cells - metabolism JMJD2C Jumonji Domain-Containing Histone Demethylases - metabolism MAP Kinase Kinase 1 - antagonists & inhibitors MAP Kinase Kinase 1 - metabolism MEKi methylation Mice Mouse Embryonic Stem Cells - cytology Mouse Embryonic Stem Cells - metabolism phosphorylation PRDM14 Protein Kinase Inhibitors - pharmacology Proteolysis TET1 Transcription Factors - genetics Transcription Factors - metabolism Ubiquitination MEKi methylation 2i JMJD2C TET1 PRDM14 GSK3i phosphorylation G9a
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Summary:	Mouse embryonic stem cells (ESCs) are maintained in serum with leukemia inhibitory factor (LIF) to maintain self-renewal and pluripotency. Recently, a 2i culture method was reported using a combination of MEK inhibition (MEKi) and GSK3 inhibition (GSK3i) with LIF to maintain ESCs in a naive ground state. How 2i maintains a ground state of ESCs remains elusive. Here we show that MEKi and GSK3i maintain the ESC ground state by downregulating global DNA methylation through two distinct mechanisms. MEK1 phosphorylates JMJD2C for ubiquitin-mediated protein degradation. Therefore, MEKi increased JMJD2C protein levels but decreased DNMT3 expression. JMJD2C promotes TET1 activity to increase 5-hydroxymethylcytosine (5hmC) levels. GSK3i suppressed DNMT3 expression, thereby decreasing DNA methylation without affecting 5hmC levels. Furthermore, 2i increased PRDM14 expression to inhibit DNMT3A/B protein expression by promoting G9a-mediated DNMT3A/B protein degradation. Collectively, 2i allows ESCs to maintain a naive ground state through JMJD2C-dependent TET1 activation and PRDM14/G9a-mediated DNMT3A/B protein degradation. [Display omitted] •MEKi increases JMJD2C protein levels and decreases DNMT3 expression in ESCs•JMJD2C promotes TET1 hydroxylase activity to increase global 5hmC levels•GSK3i decreases global DNA methylation without affecting 5hmC levels•2i-induced PRDM14 expression promotes G9a-mediated DNMT3A/B protein degradation In this article, Kim and colleagues show that MEKi and GSK3i maintain the ESC ground state by downregulating global DNA methylation through two distinct mechanisms. MEKi increased JMJD2C protein levels, thereby enhancing TET1 activity, whereas GSK3i decreased DNMT3 family expression. Therefore, they uncover that DNA demethylation pathways under 2i conditions are governed by the JMJD2C/TET1 and PRDM14/G9a/DNMT3 complexes.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 Co-first author
ISSN:	2213-6711 2213-6711
DOI:	10.1016/j.stemcr.2017.04.001