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Originally published In Press as doi:10.1074/jbc.M002766200 on July 6, 2000

J. Biol. Chem., Vol. 275, Issue 41, 31876-31882, October 13, 2000
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Raf-1/MEK/MAPK Pathway Is Necessary for the G2/M Transition Induced by Nocodazole*

Cynthia HayneDagger , Guri Tzivion§, and Zhijun LuoDagger ||

From the Dagger  Diabetes and Metabolism Research Unit, Section of Endocrinology, Evans Department of Medicine and the  Department of Biochemistry, Boston University School of Medicine, Boston, Massachusetts 02118 and the § Diabetes Unit and Medical Services and the Department of Molecular Biology, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts 02114

The dynamic balance between polymerization and depolymerization of microtubules is critical for cells to enter and exit mitosis, and drugs that disrupt this balance, such as taxol, colchicine, and nocodazole, arrest the cell cycle in mitosis. Although the Raf/MEK/MAPK pathway can be activated by these drugs, its role in mitosis has not been addressed. Here, we characterize activation of Raf/MEK/MAPK by nocodazole when mitosis is induced. We find that at early time points (up to 3 h) in nocodazole induction, Raf/MEK/MAPK is activated, and inhibition of MAPK activation by a MEK inhibitor, PD98059 or U0126, reduces the number of cells entering mitosis by creating a block at G2. At later time points and in mitosis, activation of MEK/MAPK is severely inhibited, even though Raf-1 activity remains high and can be further increased by growth factor. This inhibition is reversed when cells are released from metaphase and enter G0/G1 phase. In addition, we find that binding of Raf-1 to 14-3-3 is progressively induced by nocodazole, reaching a maximum in mitosis, and that this binding is necessary to maintain mitotic Raf-1 activity. Our present study indicates that activation of the Raf/MEK/MAPK pathway is necessary for the G2/M progression.

* This work was supported by National Institutes of Health Grant GM 57959 (to Z. L.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

|| To whom correspondence should be addressed: Diabetes and Metabolism Research Unit, Section of Endocrinology, Evans Dept. of Medicine, Boston University School of Medicine, 650 Albany St., Rm. 820, Boston, MA 02118. Tel.: 617-414-1033; Fax: 617-638-7094; E-mail: zluo@med-med1.bu.edu.

Copyright © 2000 by The American Society for Biochemistry and Molecular Biology, Inc.
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