Casein kinase Iγ subfamily: Molecular cloning, expression, and characterization of three mammalian isoforms and complementation of defects in the Saccharomyces cerevisiae YCK genes

Lanmin Zhai, Paul R. Graves, Lucy C. Robinson, Michelle Italiano, Michael R. Culbertson, Joie Rowles, Melanie H. Cobb, Anna A. DePaoli-Roach, Peter J. Roach

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Abstract

Casein kinase I, one of the first protein kinases identified biochemically, is known to exist in multiple isoforms in mammals. Using a partial cDNA fragment corresponding to an isoform termed CK1γ, three full-length rat testis cDNAs were cloned that defined three separate members of this subfamily. The isoforms, designated CK1γ1, CK1γ2, and CK1γ3, have predicted molecular masses of 43,000, 45,500, and 49,700. CK1γ3 may also exist in an alternatively spliced form. The proteins are more than 90% identical to each other within the protein kinase domain but only 51-59% identical to other casein kinase I isoforms within this region. Messages for CK1γ1 (2 kilobases (kb)), CK1γ2 (1.5 and 2.4 kb), and CK1γ3 (2.8 kb) were detected by Northern hybridization of testis RNA. Message for CK1γ3 was also observed in brain, heart, kidney, lung, liver, and muscle whereas CK1γ1 and CK1γ2 messages were restricted to testis. All three CK1γ isoforms were expressed as active enzymes in Escherichia coli and partially purified. The enzymes phosphorylated typical in vitro casein kinase I substrates such as casein, phosvitin, and a synthetic peptide, D4. Phosphorylation of the D4 peptide was activated by heparin whereas phosphorylation of the protein substrates was inhibited. The known casein kinase I inhibitor CK1-7 also inhibited the CK1γs although less effectively than the CK1α or CK1δ isoforms. All three CK1γs underwent autophosphorylation when incubated with ATP and Mg2+. The YCK1 and YCK2 genes in Saccharomyces cerevisiae encode casein kinase I homologs, defects in which lead to aberrant morphology and growth arrest. Expression of mammalian CK1γ1 or CK1γ3 restored growth and normal morphology to a yeast mutant carrying a disruption of YCK1 and a temperature-sensitive allele of YCK2, suggesting overlap of function between the yeast Yck proteins and these CK1 isoforms.

Original languageEnglish (US)
Pages (from-to)12717-12724
Number of pages8
JournalJournal of Biological Chemistry
Volume270
Issue number21
DOIs
StatePublished - May 26 1995

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ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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