Characterization of Pak2p, a Pleckstrin Homology Domain-containing, p21-activated Protein Kinase from Fission Yeast

Mary Ann Sells,Justin T Barratt,Juliane Caviston,Sabine Ottilie,Ekkehard Leberer,Jonathan Chernoff

doi:10.1074/jbc.273.29.18490

Mary Ann Sells, Justin T Barratt + Show 4 more

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https://doi.org/10.1074/jbc.273.29.18490

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Abstract

p21-activated kinases (PAKs) bind to and are activated by Rho family GTPases such as Cdc42 and Rac. Since these GTPases play key roles in regulating cell polarity, stress responses, and cell cycle progression, the ability of PAK to affect these processes has been examined. We previously showed that fission yeast pak1+ encodes an essential protein that affects mating and cell polarity. Here, we characterize a second pak gene (pak2+) from Schizosaccharomyces pombe. Like the Saccharomyces cerevisiae proteins Cla4p and Skm1p, fission yeast Pak2p contains an N-terminal pleckstrin homology domain in addition to a p21-binding domain and a protein kinase domain that are common to other members of the PAK family. Unlike pak1+, pak2(+) is not essential for vegetative growth or for mating in S. pombe. Overexpression of the wild-type pak2+ allele suppresses the lethal growth defect associated with deletion of pak1+, and this suppression requires both the pleckstrin homology- and the p21-binding domains of Pak2p, as well as kinase activity. A substantial fraction of Pak2p is associated with membranous components, an association mediated both by the pleckstrin homology- and by the p21-binding domains. These results show that S. pombe encodes at least two pak genes with distinct functions and suggest that the membrane localization of Pak2p, directed by its interactions with membrane lipids and Cdc42p, is critical to its biological activity.

Highlights

Signal transduction by small GTPases of the Rho family has become the focus of intensive investigation in recent years
The predicted Pak2p protein contains a pleckstrin homology (PH) domain at the N terminus, followed by a p21binding domain, and a serine/threonine protein kinase domain that occupies the C-terminal half of the protein (Fig. 1B)
Like Pak1p, Pak2p binds to activated Cdc42p and affects cell morphology when overexpressed

Summary

Introduction

Signal transduction by small GTPases of the Rho family has become the focus of intensive investigation in recent years. Rho GTPases (in mammals, Cdc, Rac, and Rho) regulate a variety of signaling pathways in higher eukaryotes, including those that affect the organization of cortical actin, stress responses, gene transcription, and cell cycle progression [1]. Several candidate effectors for Rho GTPases have been identified recently, including a variety of protein kinases, GTPase activating proteins, and other proteins that lack obvious enzymatic function. Dominant interfering forms of PAK block stress-activated protein kinase activation by Cdc and Rac, suggesting that PAKs may be effectors for these GTPases in stress signaling pathways. We recently characterized a PAK-like protein in S. pombe This kinase, known as Pak1p or Shk1p, is essential for viability and mating and may play important roles in the generation and/or maintenance of cell polarity [14, 15]. We report the biochemical and functional characterization of Pak2p

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