A multicopy suppressor gene of the Saccharomyces cerevisiae G1 cell cycle mutant gene dbf4 encodes a protein kinase and is identified as CDC5.

Abstract

We have isolated a multicopy suppressor of the temperature-sensitive growth phenotype of organisms carrying mutations of DBF4, a gene that is required for the initiation of chromosomal DNA replication in Saccharomyces cerevisiae and that interacts with the CDC7 protein kinase. periodically during the cell cycle, peaking at the G2/M boundary. CDCS on a multicopy plasmid also suppresses temperature-sensitive cdc15, cdc2O, and dbf2 mutations which affect mitosis during the cell cycle.Protein phosphorylation by protein kinases plays an important role in regulating both the mitotic cell cycle and meiosis in eukaryotes (28, 49). In the yeast Saccharomyces cerevisiae, genes encoding more than 30 protein kinases have been identified (16). Among these, at least four protein kinase genes, CDC28, CDC7, DBF2, and HRR25, are associated with DNA metabolism in the mitotic cell cycle. CDC28 is essential for cell growth and is required both for entry into the S phase and for the G2/M transition (35). Although the cellular abundance of the Cdc28 protein remains constant throughout the cell cycle (30), its protein kinase is periodically active and is regulated by a physical interaction with G, and G2 cyclins (8, 37, 45). CDC7 is also essential for cell growth and is required for the GJIS transition. It has recently been shown that the Cdc7 polypeptide has an associated protein kinase activity and is phosphorylated in vivo (18). Its execution point is just before the initiation of DNA replication (12). After the CDC7 execution point, no protein synthesis is required for the initiation of DNA replication (15). The abundance of the CDC7 transcript remains constant throughout the cell cycle (42), and this is likely to be true for the Cdc7 polypeptide also. However, no direct relationship between the Cdc28 and Cdc7 protein kinases has been elucidated to date.Deletion of DBF2 is not lethal because a homolog ofDBF2 (DBF20) is able to substitute for DBF2 function (48). However, in dbf2 mutants DNA synthesis is transiently delayed and the cell cycle is blocked in late nuclear division at the restrictive temperature, suggesting that the Dbf2 protein is required for completion of the S and M phases (20). The DBF2 transcript accumulates periodically during the cell * Corresponding author.cycle (20), and it is likely that this is also true of the Dbf2 protein. HRR25 is not essential for cell growth, but deletion of HRR25 results in a delay at the G2/M boundary (17), suggesting an important role in the cell cycle. Mutant cells carrying hrr25 exhibit sensitivity to methyl methanesulfonate and X rays (17), suggesting that the gene is also required for DNA repair.In an attempt to identify the various proteins that might interact either with the Cdc7 protein kinase or with both the Cdc28 and Cdc7 protein kinases, we isolated a multicopy suppressor of the temperature-sensitive growth phenotype of organisms carrying cdc7 mutations (24). The suppressor was identified as the DBF4 gene, whose execution point is just before initiation of DNA replication ...

Mining DNA microarray data using a novel approach based on graph theory

“…Proteins posited to be critical in the S phase or phosphate metabolism are thus hypothesized to be associated with an inviable null phenotype. In agreement with this hypothesis, the null phenotypes for the DN detected were indeed inviable [14,15]: all DN predicted in these proof-of-principle experiments displayed inviable null phenotypes. Conversely, all proteins not predicted to be DN displayed viable null phenotypes, implying that these are not required for survival, with a very few exceptions^two of 15 for the cell cycle study, and zero of eight for the phosphate metabolism study^exceptions that proved to be informative (Table 1): CDC20 was the only protein predicted to be neither a DN nor a SN yet still displayed a non-viable null phenotype, but importantly the function of CDC20 is not restricted to the cell cycle.…”

Adhesion induced expression of the serine/threonine kinase Fnk in human macrophages

“…The Drosophila gene polo and the yeast cell cycle genes Cdc5 and plo1 encode related protein kinases which are required for progression through mitosis (Sunkel and Glover, 1988;Llamazares et al, 1991;Fenton and Glover, 1993;Kitada et al, 1993;Ohkura et al, 1995;Toczyski et al, 1997). In recent studies mammalian protein kinases have been identi®ed which are homologous to the Drosophila gene polo (Simmons et al, 1992;Clay et al, 1993;Lake and Jelinek, 1993;Golsteyn et al, 1994;Hamanaka et al, 1994;Holtrich et al, 1994).…”

The physical association and phosphorylation of Cdc25C protein phosphatase by Prk

“…In the past few years, an emerging family of protein kinases (the polo kinase family) has been described in yeast (Kitada et al, 1993), Drosophila (Llamazares et al, 1991), Xenopus (Kumagai and Dunphy, 1996), mouse (Simmons et al, 1992;Golsteyn et al, 1995;Donohue et al, 1995), and human (Hamanaka et al, 1994;Li et al, 1996; also see recent review Nigg, 1998). The Drosophila polo gene, homologous to the budding yeast Cdc5, encodes a serine/threonine (Ser/Thr) kinase and is required for mitosis in this species; mutations in this gene result in abnormal mitotic and meiotic division (Fenton and Glover, 1993).…”