Open AccessIdentification of Yin-Yang Regulators and a Phosphorylation Consensus for Male Germ Cell-Associated Kinase (MAK)-Related Kinase
Author(s) -
Zheng Fu,
Katherine A. Larson,
Raghu Chitta,
Sirlester A. Parker,
Benjamin E. Turk,
Matthew W. Lawrence,
Philipp Kaldis,
Konstantin Galaktionov,
Steven M. Cohn,
Jeffrey Shabanowitz,
Donald F. Hunt,
Thomas W. Sturgill
Publication year - 2006
Publication title -
molecular and cellular biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.14
H-Index - 327
eISSN - 1067-8824
pISSN - 0270-7306
DOI - 10.1128/mcb.00816-06
Subject(s) - biology , cyclin dependent kinase 2 , cyclin dependent kinase 7 , cyclin dependent kinase 9 , map kinase kinase kinase , mitogen activated protein kinase kinase , cyclin dependent kinase , map2k7 , cyclin dependent kinase 1 , cyclin dependent kinase 4 , kinase , autophosphorylation , biochemistry , microbiology and biotechnology , protein kinase a , cell cycle , cell
MAK (male germ cell-associated protein kinase) and MRK/ICK (MAK-related kinase/intestinal cell kinase) are human homologs of Ime2p inSaccharomyces cerevisiae and of Mde3 and Pit1 inSchizosaccharomyces pombe and are similar to human cyclin-dependent kinase 2 (CDK2) and extracellular signal-regulated kinase 2 (ERK2). MAK and MRK require dual phosphorylation in a TDY motif catalyzed by an unidentified human threonine kinase and tyrosine autophosphorylation. Herein, we establish that human CDK-related kinase CCRK (cell cycle-related kinase) is an activating T157 kinase for MRK, whereas active CDK7/cyclin H/MAT1 complexes phosphorylate CDK2 but not MRK. Protein phosphatase 5 (PP5) interacts with MRK in a complex and dephosphorylates MRK at T157 in vitro and in situ. Thus, CCRK and PP5 are yin-yang regulators of T157 phosphorylation. To determine a substrate consensus, we screened a combinatorial peptide library with active MRK. MRK preferentially phosphorylates R-P-X-S/T-P sites, with the preference for arginine at position −3 (P−3) being more stringent than for prolines at P−2 and P+1. Using the consensus, we identified a putative phosphorylation site (RPLT1080 S) for MRK in human Scythe, an antiapoptotic protein that interacts with MRK. MRK phosphorylates Scythe at T1080 in vitro as determined by site-directed mutagenesis and mass spectrometry, supporting the consensus and suggesting Scythe as a physiological substrate for MRK.