WWP2 promotes degradation of transcription factor OCT4 in human embryonic stem cells

Key Laboratory of Stem Cell Biology, Institute of Health Sciences, [corrected] Shanghai Jiao Tong University School of Medicine, Shanghai Institutes for Biological Sciences, [corrected] 225 South Chongqing Road, Shanghai 200025, China.
Cell Research (Impact Factor: 12.41). 04/2009; 19(5):561-73. DOI: 10.1038/cr.2009.31
Source: PubMed


POU transcription factor OCT4 not only plays an essential role in maintaining the pluripotent and self-renewing state of embryonic stem (ES) cells but also acts as a cell fate determinant through a gene dosage effect. However, the molecular mechanisms that control the intracellular OCT4 protein level remain elusive. Here, we report that human WWP2, an E3 ubiquitin (Ub)-protein ligase, interacts with OCT4 specifically through its WW domain and enhances Ub modification of OCT4 both in vitro and in vivo. We first demonstrated that endogenous OCT4 in human ES cells can be post-translationally modified by Ub. Furthermore, we found that WWP2 promoted degradation of OCT4 through the 26S proteasome in a dosage-dependent manner, and the active site cysteine residue of WWP2 was required for both its enzymatic activity and proteolytic effect on OCT4. Remarkably, our data show that the endogenous OCT4 protein level was significantly elevated when WWP2 expression was downregulated by specific RNA interference (RNAi), suggesting that WWP2 is an important regulator for maintaining a proper OCT4 protein level in human ES cells. Moreover, northern blot analysis showed that the WWP2 transcript was widely present in diverse human tissues/organs and highly expressed in undifferentiated human ES cells. However, its expression level was quickly decreased after human ES cells differentiated, indicating that WWP2 expression might be developmentally regulated. Our findings demonstrate that WWP2 is an important regulator of the OCT4 protein level in human ES cells.

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Available from: Chunliang Li, Mar 06, 2014
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    • "OCT4 is subject to post translational modifications including phosphorylation [21]–[23], poly-ubiquitination [24], [25] and sumoylation [26]–[28]. For example, AKT1 phosphorylates OCT4 at threonine 235 (T235) in embryonic carcinoma cells [22]. "
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    ABSTRACT: Stem cell research can lead to the development of treatments for a wide range of ailments including diabetes, heart disease, aging, neurodegenerative diseases, spinal cord injury, and cancer. OCT4 is a master regulator of self-renewal of undifferentiated embryonic stem cells. OCT4 also plays a crucial role in reprogramming of somatic cells into induced pluripotent stem (iPS) cells. Given known vivo reproductive toxicity of cobalt and nickel metals, we examined the effect of these metals on expression of several stem cell factors in embryonic Tera-1 cells, as well as stem cells. Cobalt and nickel induced a concentration-dependent increase of OCT4 and HIF-1α, but not NANOG or KLF4. OCT4 induced by cobalt and nickel was due primarily to protein stabilization because MG132 stabilized OCT4 in cells treated with either metals and because neither nickel nor cobalt significantly modulated its steady-state mRNA level. OCT4 stabilization by cobalt and nickel was mediated largely through reactive oxygen species (ROS) as co-treatment with ascorbic acid abolished OCT4 increase. Moreover, nickel and cobalt treatment increased sumoylation and mono-ubiquitination of OCT4 and K123 was crucial for mediating these modifications. Combined, our observations suggest that nickel and cobalt may exert their reproductive toxicity through perturbing OCT4 activity in the stem cell compartment.
    Full-text · Article · Jan 2014 · PLoS ONE
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    • "Different post-translational modifications of Oct4 have been identified. For example, E3 ubiquitin ligase Wwp2 can mediate ubquitination of Oct4 to enhance its instability in ES cells [31]. Potential protein kinase A (PKA) phosphorylates serine 229 of Oct4 [32]. "
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    ABSTRACT: Cdk1 plays an important role in undifferentiated ES cells, but the underlying mechanism remains unclear. This study explores how Cdk1 collaborates with Oct4 to inhibit differentiation in mouse ES cells. We show a direct interaction between Cdk1 and Oct4, whereas other Cdk members, including Cdk2 and Cdk4, fail to associate with Oct4. By immunocytochemistry we show that Cdk1 and Oct4 co-localize in ES cells. The biological function of the Cdk1-Oct4 complex was also addressed. We found that Cdk1 enhances the binding of Oct4 on the trophectoderm marker Cdx2 and promotes Cdx2 repression. This regulation is independent of cyclins and of the kinase activity of Cdk1. Our study explains how Cdk1 and Oct4 interplay to inhibit ES cell differentiation into trophectoderm and thereby maintain stemness. STRUCTURED SUMMARY OF PROTEIN INTERACTIONS: Cdk1physically interactswithOct4byanti tag coimmunoprecipitation(View interaction) Oct4bindstoSOX-2bypull down(View interaction) Cdk1physically interactswithOct4andcyclin-B1byanti bait coimmunoprecipitation(View interaction) Oct4bindstoCdk1bypull down(View interaction) Cdk1andOct4colocalizebyfluorescence microscopy(View interaction) Oct4physically interactswithSox2byanti bait coimmunoprecipitation(View interaction).
    Full-text · Article · Oct 2012 · FEBS letters
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    • "Wwp2 catalyzes Oct4 ubiquitination in a dosage-dependent manner and also regulates its own ligase activity by auto-ubiquitination (Liao and Jin, 2010). Subsequent study shows that the human counterpart of Wwp2, WWP2, ubiquitinates OCT4, and promotes its degradation through the 26S proteasome in human ESCs (Xu et al., 2009). c-Myc has also been reported to be regulated by ubiquitin–proteasome system (UPS). "
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    ABSTRACT: The maintenance of pluripotency relies on an intricate transcriptional network hinged on a key set of transcription factors. Pluripotent stem cells have been shown to be sensitive to modulations of the cellular abundance and transcriptional activity of these key pluripotency factors. Recent evidence highlights the important role of post-translational modifications, including ubiquitination, sumoylation, phosphorylation, methylation, and acetylation, in regulating the levels and activity of pluripotency factors to achieve a balance between pluripotency and differentiation.
    Full-text · Article · Jun 2012 · Journal of Molecular Cell Biology
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