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ERK1 and ERK2 regulate embryonic stem cell self-renewal through phosphorylation of Klf4

Nature Structural & Molecular Biology volume 19, pages 283–290 (2012)Cite this article

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Abstract

Understanding and controlling the mechanism by which stem cells balance self-renewal versus differentiation is of great importance for stem cell therapeutics. Klf4 promotes the self-renewal of embryonic stem cells, but the precise mechanism regulating this role of Klf4 is unclear. We found that ERK1 or ERK2 binds the activation domain of Klf4 and directly phosphorylates Klf4 at Ser123. This phosphorylation suppresses Klf4 activity, inducing embryonic stem cell differentiation. Conversely, inhibition of Klf4 phosphorylation enhances Klf4 activity and suppresses embryonic stem cell differentiation. Notably, phosphorylation of Klf4 by ERKs causes recruitment and binding of the F-box proteins βTrCP1 or βTrCP2 (components of an ubiquitin E3 ligase) to the Klf4 N-terminal domain, which results in Klf4 ubiquitination and degradation. Overall, our data provide a molecular basis for the role of ERK1 and ERK2 in regulating Klf4-mediated mouse embryonic stem cell self-renewal.

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Figure 1: Klf4 is a substrate of ERK1 or ERK2.
Figure 2: ERK1 phosphorylates Klf4 Ser123.
Figure 3: Inhibiting ERK signaling induces self-renewal of embryonic stem cells.
Figure 4: Klf4 Ser123 phosphorylation has a role in embryonic stem cell self-renewal.
Figure 5: Klf4 Ser123 phosphorylation induces βTrCP binding.
Figure 6: Effect of the βTrCP2 phosphodegron in Klf4 on binding with βTrCP2.

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Acknowledgements

We thank M. Pagano (New York University School of Medicine) for the kind gift of plasmids, including βTrCP1/2 and several F boxes. We thank H. Niwa (RIKEN Center for Developmental Biology (CDB)) for the kind gift of plasmids, including the Lefty1 core promoter. We wish to thank T.M. Poorman for secretarial assistance. This work was supported by The Hormel Foundation.

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Author notes

  1. Yong-Yeon Cho

    Present address: Current Address: College of Pharmacy, The Catholic University of Korea, Bucheon-si, Korea.,

  2. Myoung Ok Kim and Sung-Hyun Kim: These authors contributed equally to this work.

Authors and Affiliations

  1. The Hormel Institute, University of Minnesota, Austin, Minnesota, USA

    Myoung Ok Kim, Sung-Hyun Kim, Yong-Yeon Cho, Janos Nadas, Chul-Ho Jeong, Ke Yao, Dong Joon Kim, Dong-Hoon Yu, Young-Sam Keum, Kun-Yeong Lee, Zunnan Huang, Ann M Bode & Zigang Dong

  2. Center for Laboratory Animal Resources, KyungPook National University, Daegu, Republic of Korea

    Sung-Hyun Kim

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Contributions

M.O.K. and S.-H.K. designed experiments, participated in writing the manuscript, cultured embryonic stem cells and conducted immunoprecipitation experiments and screening. Y.-Y.C. designed experiments and participated in writing the manuscript. J.N. and Z.H. carried out the computational biology and modeling. C.-H.J. participated in experimental design and cultured embryonic stem cells. K.Y. cultured embryonic stem cells and did the immunofluorescence and kinase assays. D.J.K. did the kinase assays. D.-H.Y. cultured embryonic stem cells. Y.-S.K. participated in designing the experiments and cultured the lentivirus for knockdown of Klf4. K.-Y.L. cultured embryonic stem cells. A.M.B. and Z.D. supervised the design of the experiments and the manuscript editing and writing.

Corresponding author

Correspondence to Zigang Dong.

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Kim, M., Kim, SH., Cho, YY. et al. ERK1 and ERK2 regulate embryonic stem cell self-renewal through phosphorylation of Klf4. Nat Struct Mol Biol 19, 283–290 (2012). https://doi.org/10.1038/nsmb.2217

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