Identification of Yin-Yang regulators and a phosphorylation consensus for male germ cell-associated kinase (MAK)-related kinase

Zheng Fu, Katherine A. Larson, Raghu K. 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

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47 Scopus citations

Abstract

MAK (male germ cell-associated protein kinase) and MRK/ICK (MAK-related kinase/intestinal cell kinase) are human homologs of Ime2p in Saccharomyces cerevisiae and of Mde3 and Pit1 in Schizosaccharomyces 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 CDIC7/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 (RPLT1080S) 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.

Original languageEnglish (US)
Pages (from-to)8639-8654
Number of pages16
JournalMolecular and cellular biology
Volume26
Issue number22
DOIs
StatePublished - Nov 2006

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology

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    Fu, Z., Larson, K. A., Chitta, R. K., Parker, S. A., Turk, B. E., Lawrence, M. W., Kaldis, P., Galaktionov, K., Cohn, S. M., Shabanowitz, J., Hunt, D. F., & Sturgill, T. W. (2006). Identification of Yin-Yang regulators and a phosphorylation consensus for male germ cell-associated kinase (MAK)-related kinase. Molecular and cellular biology, 26(22), 8639-8654. https://doi.org/10.1128/MCB.00816-06