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Two Distinct Classes of Mitotic Cyclin Homologues, Cyc1 and Cyc2, Are Involved In Cell Cycle Regulation In the Ciliate Paramecium Tetraurelia

ABSTRACT The eukaryotic cell cycle is regulated by the sequential activation of different CDK/cyclin complexes. Two distinct classes of mitotic cyclin homologues, CYC1 and CYC2, have been identified and cloned for the first time in the ciliate Paramecium. Cyc1 is 324 amino acids long with a predicted molecular mass of 38 kDa, whereas Cyc2 is 336 amino acids long with a predicted molecular mass of 40 kDa. They display 42‐51% sequence identity to other eukaryotic mitotic cyclins within the ‘cyclin box’ region. the conserved ‘cyclin box’ and ‘destruction box’ elements can be identified within each of the sequences. Genomic Southern blot analysis indicated that the CYC1 gene has two isoforms, with 92.3% and 85.9% identity at the amino acid level and at the necleotide level, respectively. Both Cyc1 and Cyc2 proteins showed characteristic patterns of 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 late in the cell cycle. Immunoprecipitation experiments with antibodies specific to Cyc1 and Cyc2 indicated that Cyc1 and Cyc2 associate with distinct CDK homologues. Both immunoprecipitates exhibited histone H1 kinase activity that oscillated in the cell cycle in parallel with the respective amount of cyclins present. Histone H1 kinase activity associated with Cyc1 reached a peak at PCD while Cyc2 showed maximal activity when about 75% cells have completed cytokinesis. We propose that Cyc1 may be involved in commitment to division, in association with the CDK that binds to p13suc1, Cdk3, and that the Cyc2/Cdk2 complex may regulate cytokinesis. PCR‐amplification revealed similar sequences in Tetrahymena, Sterkiella, Colpoda and Blepharisma. suggesting the conservation of the cyclin genes within ciliates. Although cell cycle regulation in ciliates differs in some respects from that of other eukaryotes, the cyclin motifs have clearly been conserved during evolution.