Interaction between active Pak1 and Raf-1 is necessary for phosphorylation and activation of Raf-1

Mengwei Zang, Cynthia Hayne, Zhijun Luo

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101 Scopus citations


Activation of Raf-1 is a complex process in which phosphorylation of Ser338-Tyr841 is a critical step. Previous studies have shown that Pak1/2 is implicated in both Ras-dependent and -independent activation of Raf-1 by phosphorylating Raf Ser338. The present study explores the structural basis of Raf-1 phosphorylation by Pak1. We found that Pak directly associates with Raf-1 under both physiological and overexpressed conditions. The association is greatly stimulated by 4β-12-O-tetradecanoylphorbol-13-acetate and nocodazole and by expression of the active mutants of Rac and Ras. The active forms of Pak generated by mutation of Thr423 to Glu or truncation of the amino-terminal moiety exhibit a greater binding to Raf than the wild type, whereas the kinase-dead mutant Pak barely binds Raf. The extent of binding to Raf-1 is correlated with the ability of Pak to phosphorylate Raf and induce mitogen-activated protein kinase activation. Furthermore, the Raf-1 binding site is defined to the carboxyl terminus of the Pak catalytic domain. In addition, our results suggest that the amino-terminal regulatory region of Raf inhibits the interaction. Taken together, the results indicate that the interaction depends on the active conformations of Pak and Raf. They also argue that Pakl is a physiological candidate for phosphorylation of Raf Ser338 during the course of Raf activation.

Original languageEnglish (US)
Pages (from-to)4395-4405
Number of pages11
JournalJournal of Biological Chemistry
Issue number6
StatePublished - Feb 8 2002
Externally publishedYes

ASJC Scopus subject areas

  • Molecular Biology
  • Biochemistry
  • Cell Biology


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