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Mutations in CDC14 result in high sensitivity to cyclin gene dosage in Saccharomyces cerevisiae
1Rockefeller University, New York, NY 10021, USA.
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.
Insights
Screening Saccharomyces cerevisiae for mutations causing lethality with G1 cyclin Cln2p overexpression identified MEC1, NUP170, and CDC14. CDC14 mutations interact with cell cycle regulators, impacting DNA replication and mitosis.
Area of Science:
- * Molecular and Cellular Biology
- * Yeast Genetics
- * Cell Cycle Regulation
Background:
- * Overexpression of the G1 cyclin Cln2p in Saccharomyces cerevisiae can be lethal.
- * Identifying genes that cause lethality when Cln2p is overexpressed provides insights into cell cycle control.
- * The protein Cdc14p plays a role in regulating cyclin B-associated kinase activity during mitosis.
Purpose of the Study:
- * To identify genes that are essential for viability when G1 cyclin Cln2p is overexpressed.
- * To investigate the genetic interactions of CDC14 with other cell cycle genes.
- * To understand the role of Cdc14p in regulating cell division and DNA replication.
Main Methods:
- * Screening for mutations conferring lethality upon Cln2p overexpression in Saccharomyces cerevisiae.
- * Complementation analysis to identify distinct mutation groups.
- * Gene identification and characterization of genetic interactions through gene deletions and double mutant analysis.
Main Results:
- * Mutations in MEC1, NUP170, and CDC14 were identified as causing lethality with Cln2p overexpression.
- * cdc14 mutations showed synthetic lethality with deletions of SIC1 (inhibitor of Clb-Cdc28 kinase) and HCT1 (proteolysis factor for Clb2p).
- * Synthetic lethality was also observed between cdc14-1 and CLB2 (major B-type cyclin) deletion, leading to distinct cell cycle arrest phenotypes.
- * Genetic interactions between CDC14 and CDC6 (replication initiator) and CDC28 were established.
Conclusions:
- * MEC1, NUP170, and CDC14 are important for cell cycle progression when G1 cyclin Cln2p is overexpressed.
- * Cdc14p function is critical for regulating cyclin B-Cdc28 kinase activity and preventing aberrant cell division.
- * CDC14 exhibits complex genetic interactions with genes involved in cyclin regulation, proteolysis, and DNA replication, highlighting its central role in cell cycle control.