Cloning and Characterization of a Dictyostelium Myosin I Heavy Chain Kinase Activated by Cdc42 and Rac

doi:10.1074/jbc.271.43.27044

Cloning and Characterization of a Dictyostelium Myosin I Heavy Chain Kinase Activated by Cdc42 and Rac*

From the Department of Biochemistry, Queen's University, Kingston, Ontario K7L 3N6 and
the ^‡ Department of Physiology and Biophysics, Case Western Reserve University, Cleveland, Ohio 44106

^¶ To whom all correspondence should be addressed:
Dept. of Biochemistry, Queen's University, Kingston, Ontario, Canada K7L 3N6.
Tel.: 613-545-2998; Fax: 613-545-2987; E-mail: coteg{at}post.queensu.ca

Abstract

The motile activities of the small, single-headed class I myosins (myosin I) from the lower eukaryotes Acanthamoeba and Dictyostelium are activated by phosphorylation of a single serine or threonine residue in the head domain of the heavy chain. Recently, we purified a myosin I heavy chain kinase (MIHCK) from Dictyostelium based on its ability to activate the Dictyostelium myosin ID isozyme (Lee, S.-F., and Côté, G. P. (1995) J. Biol. Chem. 270, 11776-11782). The complete sequence of the Dictyostelium MIHCK has now been determined, revealing a protein of 98 kDa that is composed of an amino-terminal domain rich in proline, glutamine, and serine, a putative Cdc42/Rac binding motif, and a carboxyl-terminal kinase catalytic domain. MIHCK shares significant sequence identity with the Saccharomyces cerevisiae Ste20p kinase and the mammalian p21-activated kinase. Gel overlay assays and affinity chromatography experiments showed that MIHCK interacted with GTPγS (guanosine 5′-3-O-(thiotriphosphate))-labeled Cdc42 and Rac1 but not RhoA. In the presence of GTPγS-Rac1 MIHCK autophosphorylation increased from 1 to 9 mol of phosphate/mol, and the rate of Dictyostelium myosin ID phosphorylation was stimulated 10-fold. MIHCK may therefore provide a direct link between Cdc42/Rac signaling pathways and motile processes driven by myosin I molecules.

Footnotes

↵^§ American Cancer Society Junior Faculty Research Professor.
↵* This work was supported by the Medical Research Council of Canada and by National Institutes of Health Grant GM50009. 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.

The nucleotide sequence(s) reported in this paper has been submitted to the GenBank™/EMBL Data Bank with accession number(s) U67715[GenBank].
↵¹ The abbreviations used are:

SH3

Src homology 3

MIHCK

myosin I heavy chain kinase

PAK

p21-activated kinase

GST

glutathione S-transferase

GTPγS

guanosine 5′-3-O-(thio)triphosphate

Tes

2-{[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]amino}ethanesulfonic acid.
- Received August 5, 1996.