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Ronin is essential for embryogenesis and the pluripotency of mouse embryonic stem cells.

ABSTRACT: Pluripotency is a unique biological state that allows cells to differentiate into any tissue type. Here we describe a candidate pluripotency factor, Ronin, that possesses a THAP domain, which is associated with sequence-specific DNA binding and epigenetic silencing of gene expression. Ronin is expressed primarily during the earliest stages of murine embryonic development, and its deficiency in mice produces periimplantational lethality and defects in the inner cell mass. Conditional knockout of Ronin prevents the growth of ES cells while forced expression of Ronin allows ES cells to proliferate without differentiation under conditions that normally do not promote self-renewal. Ectopic expression also partly compensates for the effects of Oct4 knockdown. We demonstrate that Ronin binds directly to HCF-1, a key transcriptional regulator. Our findings identify Ronin as an essential factor underlying embryogenesis and ES cell pluripotency. Its association with HCF-1 suggests an epigenetic mechanism of gene repression in pluripotent cells.

SUBMITTER: Dejosez M

PROVIDER: S-EPMC2495776 | biostudies-other | 2008 Jun

REPOSITORIES: biostudies-other

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Ronin is essential for embryogenesis and the pluripotency of mouse embryonic stem cells.

Dejosez Marion M Krumenacker Joshua S JS Zitur Laura Jo LJ Passeri Marco M Chu Li-Fang LF Songyang Zhou Z Thomson James A JA Zwaka Thomas P TP

Cell 20080601 7

Pluripotency is a unique biological state that allows cells to differentiate into any tissue type. Here we describe a candidate pluripotency factor, Ronin, that possesses a THAP domain, which is associated with sequence-specific DNA binding and epigenetic silencing of gene expression. Ronin is expressed primarily during the earliest stages of murine embryonic development, and its deficiency in mice produces periimplantational lethality and defects in the inner cell mass. Conditional knockout of ...[more]

PMID: 18585351

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Project description:Pluripotent cells are subject to much interest as a source of differentiated cellular material for research models, regenerative medical therapies and novel applications such as lab-cultured meat. Greater understanding of the pluripotent state and control over its differentiation is therefore desirable. The role of biomechanical properties in directing cell fate and cell behavior has been increasingly well described in recent years. However, many of the mechanisms which control cell morphology and mechanical properties in somatic cells are absent from pluripotent cells. We leveraged naturally occurring variation in biomechanical properties and expression of pluripotency genes in murine ESCs to investigate the relationship between these parameters. We observed considerable variation in a Rex1-GFP expression reporter line and found that this variation showed no apparent correlation to cell spreading morphology as determined by circularity, Feret ratio, phase contrast brightness or cell spread area, either on a parameter-by-parameter basis, or when evaluated using a combined metric derived by principal component analysis from the four individual criteria. We further confirmed that cell volume does not co-vary with Rex1-GFP expression. Interestingly, we did find that a subpopulation of cells that were readily detached by gentle agitation collectively exhibited higher expression of Nanog, and reduced LmnA expression, suggesting that elevated pluripotency gene expression may correlate with reduced adhesion to the substrate. Furthermore, atomic force microscopy and quantitative fluorescent imaging revealed a connection between cell stiffness and Rex1-GFP reporter expression. Cells expressing high levels of Rex1-GFP are consistently of a relatively low stiffness, while cells with low levels of Rex1-GFP tend toward higher stiffness values. These observations indicate some interaction between pluripotency gene expression and biomechanical properties, but also support a strong role for other interactions between the cell culture regime and cellular biomechanical properties, occurring independently of the core transcriptional network that supports pluripotency.

Project description:BackgroundPluripotent embryonic stem cells grown under standard conditions (ESC) have a markedly glycolytic profile, which is shared with many different types of cancer cells. Thus, some therapeutic strategies suggest that pharmacologically shifting cancer cells towards an oxidative phenotype, using glycolysis inhibitors, may reduce cancer aggressiveness. Given the metabolic parallels between cancer and stemness would chemotherapeutical agents have an effect on pluripotency, and could a strategy involving these agents be envisioned to modulate stem cell fate in an accessible manner? In this manuscript we attempted to determine the effects of 3-bromopyruvate (3BrP) in pluripotency. Although it has other intracellular targets, this compound is a potent inhibitor of glycolysis enzymes thought to be important to maintain a glycolytic profile. The goal was also to determine if we could contribute towards a pharmacologically accessible metabolic strategy to influence cell differentiation.Methodology/principal findingsMouse embryonic stem cells (mESC) grown under standard pluripotency conditions (in the presence of Leukemia Inducing Factor- LIF) were treated with 3BrP. As a positive control for differentiation other mESCs were grown without LIF. Overall our results demonstrate that 3BrP negatively affects pluripotency, forcing cells to become less glycolytic and with more active mitochondria. These changes in metabolism are correlated with increased differentiation, even under pluripotency conditions (i.e. in the presence of LIF). However, 3BrP also significantly impaired cell function, and may have other roles besides affecting the metabolic profile of mESCs.Conclusions/findingsTreatment of mESCs with 3BrP triggered a metabolic switch and loss of pluripotency, even in the presence of LIF. Interestingly, the positive control for differentiation allowed for a distinction between 3BrP effects and changes associated with spontaneous differentiation/loss of pluripotency in the absence of LIF. Additionally, there was a slight differentiation bias towards mesoderm in the presence of 3BrP. However, the side effects on cellular function suggest that the use of this drug is probably not adequate to efficiently push cells towards specific differentiation fates.

Dataset Information

Ronin is essential for embryogenesis and the pluripotency of mouse embryonic stem cells.

Publications

Ronin is essential for embryogenesis and the pluripotency of mouse embryonic stem cells.

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