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B-Raf and C-Raf are required for Ras-stimulated p42 MAP kinase activation in Xenopus egg extracts

Oncogene volume 25, pages 3307–3315 (2006)Cite this article

Abstract

During mitosis, a select pool of MEK1 and p42/p44 MAPK becomes activated at the kinetochores and spindle poles, without substantial activation of the bulk of the cytoplasmic p42/p44 MAPK. Recently, we set out to identify the MAP kinase kinase kinase (MAPKKK) responsible for this mitotic activation, using cyclin-treated Xenopus egg extracts as a model system, and presented evidence that Mos was the relevant MAPKKK . However, a second MAPKKK distinct from Mos was readily detectable as well. Here, we partially purify this second MAPKKK and identify it as B-Raf. No changes in the activity of B-Raf were detectable during progesterone-induced oocyte maturation, after egg fertilization, or during the early embryonic cell cycle, arguing against a role for B-Raf in the mitotic activation of MEK1 and p42 MAPK. Ras proteins can bring about activation of MEK1 and p42 MAPK in extracts, and Ras may contribute to signaling from the classical progesterone receptor during oocyte maturation and from receptor tyrosine kinases during early embryogenesis. We found that both B-Raf and C-Raf, but not Mos, are required for Ras-induced MEK1 and p42 MAPK activation. These data indicate that two upstream stimuli, active Ras and active Cdc2, utilize different MAPKKKs to activate MEK1 and p42 MAPK.

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Accession codes

Accessions

GenBank/EMBL/DDBJ

Abbreviations

ERK:

extracellular signal-regulated kinase

FPLC:

fast protein liquid chromatography

MEK:

MAPK kinase or ERK kinase

MAPK:

mitogen-activated protein kinase

MAPKKK:

MAP kinase kinase kinase

REKS:

Ras-activated ERK kinase stimulator

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Acknowledgements

We thank R Roth for suggesting the Ras experiments, N Ahn for providing MEK plasmids, J Cooper, J Posada, and R Bhatt for providing p42 MAPK plasmids, and members of the Ferrell lab for helpful discussions and comments on the manuscript. This work was supported by a Grant from the National Institutes of Health (GM46383). JY was supported by a postdoctoral fellowship from the American Heart Association. The sequence of Xenopus B-Raf has been deposited with the NCBI (accession no. AAU29410).

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  1. W Xiong

    Present address: W Xiong, The Scripps Research Institute, Mail Stop #SR202, 10550 N. Torrey Pines Road, La Jolla, CA 92037, USA

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  1. Department of Molecular Pharmacology, Stanford University, Stanford, CA, USA

    J Yue, W Xiong & J E Ferrell Jr

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Correspondence to J Yue.

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Yue, J., Xiong, W. & Ferrell, J. B-Raf and C-Raf are required for Ras-stimulated p42 MAP kinase activation in Xenopus egg extracts. Oncogene 25, 3307–3315 (2006). https://doi.org/10.1038/sj.onc.1209354

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