Hong Zhang, James D. Berger
Department of Zoology, University of British Columbia, 6270 University Boulevard, Vancouver, B. C. V6T 1Z4 Canada
Two mitotic cyclin homologues, CYC1 and CYC2 have been identified and cloned from ciliate Paramecium tetraurelia. CYC1 contains 324 amino acids with a predicted molecular mass of 38 kDa whereas CYC2 is 337 amino acids long with a predicted molecular mass of 40 kDa. They display about 42~51% sequence identity to other eukaryotic mitotic cyclins within the cyclin box region. Even though they are small when compared to other mitotic cyclins, the essential elements such as cyclin box and destruction box can be identified within the sequences. Genomic Southern analysis indicated that CYC1 gene has two isoforms, and they share 92.3% and 85.9% identity at the amino acid level and at the nucleotide level, respectively. Both CYC1 and CYC2 proteins show characteristic accumulation and destruction during the vegetative cell cycle, with CYC1 peaking at the point of commitment to division (PCD), and CYC2 reaching the maximal level during cytokinesis. Immunoprecipitation experiments indicate that both CYC1 and CYC2 associate with, but distinct p34cdc2 homologues. CYC1 can be precipitated by p13suc1 while CYC2 is in a complex with CDC2PTB, a p34cdc2 kinase associated with cytokinesis. Both cyclin immunoprecipitates exhibit histone H1 kinase activity that oscillate with the cell cycle in parallel with their respective amount of cyclins present. Histone H1 kinase activity associated with CYC1 reaches a peak at PCD while CYC2 shows maximal activity during cytokinesis. We propose that CYC1 may be involved in the PCD in association with CDK that binds to p13suc1, and the CYC2/CDC2PTB complexes regulate cytokinesis. These are the first cyclins that have ever been documented in ciliates. Similar sequences have also been identified in Tetrahymena, Sterkiella, Copodia and Blepharisma, suggesting the conservation of cyclin within ciliates. Our study support the idea that CDK/cyclin motif is conserved in ciliates during evolution even though the cell cycle control of this group appears different from other eukaryotes.