Molecular Cloning of Casein Kinase II a Subunit from Dictyostelium discoideum and Its Expression in the Life Cycle

Abstract

A Dictyostelium discoideum cDNA encoding an ot-type subunit of casein kinase II was isolated, and its cDNA was used to study developmental expression of casein kinase II during the Dictyostelium life cycle. The 1.3-kb cDNA insert contained an open reading frame of 337 amino acids (M_r 39,900). The deduced amino acid sequence has high homology with those of casein kinase II a subunits from other species. Genomic Southern blot analysis suggested that there is a single gene encoding casein kinase II at subunit in D. discoideum. Northern (RNA) blot analysis showed that the casein kinase II o-subunit gene is expressed constitutively as a 1.9-kb mRNA throughout vegetative growth and multicellular development. Casein kinase purified from normal vegetative cells contained a major protein band of ~36 kDa, which was recognized by antisera raised against rat testis casein kinase II. Comparison of the in vitro transcription/translation product of the α-subunit cDNA clone and the purified 36-kDa protein by partial proteolysis indicated that the isolated cDNA clone encodes the Dictyostelium casein kinase II α subunit. No protein corresponding to a β subunit was detected in purified casein kinase. Immunoblot analysis using anti-rat casein kinase II sera showed that the α subunit of casein kinase II is expressed constitutively like its mRNA during the life cycle of D. discoideum. Casein kinase II activity measured by using a specific peptide substrate paralleled the level of at subunit detected by immunoblotting during the life cycle, with a maximum variation of -2-fold. We were unable to obtain disruptants of the casein kinase II α gene, suggesting that there is a single casein kinase II ct gene, which is essential for vegetative growth of D. discoideum.