Genetic Evidence for Pak1 Autoinhibition and Its Release by Cdc42

Abstract

Pak1 protein kinase of Schizosaccharomyces pombe, a member of the p21-GTPase-activated protein kinase (PAK) family, participates in signaling pathways including sexual differentiation and morphogenesis. The regulatory domain of PAK proteins is thought to inhibit the kinase catalytic domain, as truncation of this region renders kinases more active. Here we report the detection in the two-hybrid system of the interaction between Pak1 regulatory domain and the kinase catalytic domain. Pak1 catalytic domain binds to the same highly conserved region on the regulatory domain that binds Cdc42, a GTPase protein capable of activating Pak1. Two-hybrid, mutant, and genetic analyses indicated that this intramolecular interaction rendered the kinase in a closed and inactive configuration. We show that Cdc42 can induce an open configuration of Pak1. We propose that Cdc42 interaction disrupts the intramolecular interactions of Pak1, thereby releasing the kinase from autoinhibition.

ACKNOWLEDGMENTS

We thank Ken Chang, Doug Johnson, and Aaron Neiman for providing DNA and yeast strains; Mike Riggs for DNA sequencing; Terry Vale, Hong Ma, Peter Gergen, and Marion Carlson for helpful discussion; the Cold Spring Harbor Laboratory Art Department for artwork; and Patricia Bird for secretarial assistance.

This work was supported by grants from the American Cancer Society and the National Cancer Institute (NIH) to M.W. M.W. is an American Cancer Society Professor.