Cloning and characterization of Chinese hamster homologue of yeast DBF4 (ChDBF4).
The Dbf4 protein is the regulatory subunit of Cdc7 serine/threonine kinase, which is essential for entry into S phase. We report here the cloning and initial characterization of the Chinese hamster homologue of yeast DBF4. The deduced ChDbf4 protein contains 676 amino acids with a predicted molecular mass of 75.8 kDa, and shares extensive identity overall with those of human (68%) and mouse (73%). The ChDBF4 mRNA level was barely detectable in the cells arrested in the quiescent stage (G(0)) by isoleucine starvation. When cells in G(0) were released into the cell cycle, the ChDBF4 mRNA level did not significantly change until the cells reached the G(1)/S boundary, when the level rapidly increased and reached approximately 70% of the maximum level that was observed in mid to late S phase. Interestingly, gamma-irradiation rapidly and transiently downregulated the level of ChDBF4 mRNA in asynchronous cell populations. Since Dbf4-Cdc7 kinase is involved in the regulation of replication initiation, which can be transiently downregulated by irradiation (Larner et al., 1994. Mol. Cell. Biol. 14, 1901, our data raise the possibility that the downregulation of DBF4 (and, thus, the Cdc7 kinase activity) by irradiation may play a role in the cell-cycle checkpoint that functions at the G(1)/S transition and in S phase (Lee et al., 1997. Proc. Natl. Acad. Sci. USA 94, 526).
Northeastern Ontario Regional Cancer Centre, 41 Ramsey Lake Road, Sudbury, Ontario, Canada.
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D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D016466:CHO Cells; D018797:Cell Cycle Proteins; D003001:Cloning, Molecular; D006224:Cricetinae; D018076:DNA, Complementary / Q000737:chemistry / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D016193:G1 Phase; D005786:Gene Expression Regulation / Q000528:radiation effects; D049452:Green Fluorescent Proteins; D008027:Light; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D011506:Proteins / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism / Q000528:radiation effects; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016192:Resting Phase, Cell Cycle; D016196:S Phase; D029701:Saccharomyces cerevisiae Proteins; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D018797:Cell Cycle Proteins; C424441:DBF4 protein, Cricetulus griseus; D018076:DNA, Complementary; C088096:Dbf4 protein, S cerevisiae; D005656:Fungal Proteins; D008164:Luminescent Proteins; D011506:Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D049452:Green Fluorescent Proteins