Mutations in CDC14 result in high sensitivity to cyclin gene dosage in Saccharomyces cerevisiae

Abstract

We screened for mutations that resulted in lethality when the G1 cyclin Cln2p was overexpressed throughout the cell cycle in Saccharomyces cerevisiae. Mutations in five complementation groups were found to give this phenotype, and three of the mutated genes were identified as MEC1, NUP170, and CDC14. Mutations in CDC14 may have been recovered in the screen because Cdc14p may reduce the cyclin B (Clb)-associated Cdc28 kinase activity in late mitosis, and Cln2p may normally activate Clb-Cdc28 kinase activity by related mechanisms. In agreement with the idea that cdc14 mutations elevate Clb-Cdc28 kinase activity, deletion of the gene for the Clb-Cdc28 inhibitor Sic1 caused synthetic lethality with cdc14-1, as did the deletion of HCT1, which is required for proteolysis of Clb2p. Surprisingly, deletion of the gene for the major B-type cyclin, CLB2, also caused synthetic lethality with the cdc14-1 mutation. The clb2 cdc14 strains arrested with replicated but unseparated DNA and unseparated spindle pole bodies; this phenotype is distinct from the late mitotic arrest of the sic1::TRP1 cdc14-1 and the cdc14-1 hct1::LEU2 double mutants and of the cdc14 CLN2 overexpressor. We found genetic interactions between CDC14 and the replication initiator gene CDC6, extending previous observations of interactions between the late mitotic function of Cdc14p and control of DNA replication. We also describe genetic interactions between CDC28 and CDC14.

Degradation of the Mitotic Cyclin Clb3 Is not Required for Mitotic Exit but Is Necessary for G1 Cyclin Control of the Succeeding Cell Cycle

“…GAL-SIC1 cdc14-1 CLB3 cells were viable up to 30°with GAL-SIC1 off (YEPD), and were rescued weakly at higher temperatures with GAL-SIC1 on (YEPG), consistent with previous observations (Jaspersen et al 1998;Yuste-Rojas and Cross 2000). GAL-SIC1 cdc14-1 CLB3Ddb cells were inviable at all tested temperatures with GAL-SIC1 off, and were rescued with GAL-SIC1 on up to 27°(with partial rescue at 30°).…”

“…To test for viability of CLB3Ddb cells with lowered Cdc14 activity, we used the temperature-sensitive allele cdc14-1 (Hartwell et al 1974), which can be partially rescued at nonpermissive temperature by overexpressed SIC1 (Jaspersen et al 1998;Yuste-Rojas and Cross 2000). We made 103 serial dilutions of CDC14 CLB3, CDC14 CLB3Ddb, cdc14-1 CLB3, and cdc14-1 CLB3Ddb cells (all bearing GAL-SIC1) on YEP-Glucose ("YEPD"; GAL off) and YEP-Galactose ("YEPG"; GAL on) plates, and placed them at various temperatures ( Figure 5A).…”

Cell Cycle Control of DNA Replication by Phosphorylation and Dephosphorylation of Replication-Initiation Proteins in Budding Yeast

“…Similar to all known initiation mutants reported, the plasmid loss of cdc14-1 cells could be suppressed by adding multiple copies of an ARS to the plasmid (Hogan & Koshland, 1992;Ma et al, 2010). Overexpression of the initiation protein Orc6p caused synthetic dosage lethality in cdc14-1 cells (Kroll et al, 1996), and synthetic lethality was observed when cdc14-1 was combined with cdc6-1, orc2-1 or orc5-1 (Loo et al, 1995;Kroll et al, 1996;Yuste-Rojas & Cross, 2000). Orc6p was found to be dephosphorylated by www.intechopen.com (Bloom & Cross, 2007).…”

The budding yeast Polo-like kinase localizes to distinct populations at centrosomes during mitosis

“…The cdc14-1 mutant arrests in telophase at its restrictive temperature of 37°C, but even at its permissive room temperature (25°C), this mutant is still hypomorphic because it shows synthetic lethal genetic interactions with various mutants, unlike CDC14 (Jaspersen et al. , 1998; Yuste-Rojas and Cross, 2000; Rossio et al. , 2010; Zhai et al.…”