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Cycloheximide-resistant temperature-sensitive lethal mutations of Saccharomyces cerevisiae
1Department of Biology, Brandeis University, Waltham, Massachusetts 02254.
Abstract:
We describe the isolation and preliminary characterization of a set of pleiotropic mutations resistant to the minimum inhibitory concentration of cycloheximide and screened for ts (temperature-sensitive) growth. These mutations fall into 22 complementation groups of cycloheximide resistant ts lethal mutations (crl). None of the crl mutations appears to be allelic with previously isolated mutations. Fifteen of the CRL loci have been mapped. At the nonpermissive temperature (37 degrees), these mutants arrest late in the cell cycle after several cell divisions. Half of these mutants are also unable to grow at very low temperatures (5 degrees). Although mutants from all of the 22 complementation groups exhibit similar temperature-sensitive phenotypes, an extragenic suppressor of the ts lethality of crl3 does not relieve the ts lethality of most other crl mutants. A second suppressor mutation allows crl10, crl12, and crl14 to grow at 37 degrees but does not suppress the ts lethality of the remaining crl mutants. We also describe two new methods for the enrichment of auxotrophic mutations from a wild-type yeast strain.
Insights
Researchers identified 22 new cycloheximide-resistant, temperature-sensitive lethal (crl) mutations in yeast. These mutations cause cell cycle arrest and some exhibit cold-sensitive growth, revealing new genetic pathways.
Area of Science:
- Molecular biology
- Yeast genetics
- Cell cycle regulation
Background:
- Cycloheximide resistance and temperature-sensitive (ts) mutations are valuable tools for studying essential genes.
- Identifying novel mutations aids in understanding fundamental cellular processes.
Purpose of the Study:
- To isolate and characterize new pleiotropic mutations conferring cycloheximide resistance and temperature-sensitive growth.
- To identify novel genes involved in yeast cell cycle progression and stress response.
Main Methods:
- Screening for cycloheximide-resistant, temperature-sensitive lethal (crl) mutants in yeast.
- Complementation analysis to establish 22 distinct complementation groups.
- Genetic mapping of 15 CRL loci.
- Phenotypic analysis at non-permissive (37°C) and low (5°C) temperatures.
- Analysis of extragenic suppressor mutations.
Main Results:
- Isolation of 22 complementation groups of novel crl mutations.
- Mutants arrest late in the cell cycle at 37°C after multiple divisions.
- Approximately half of the mutants show cold-sensitive growth at 5°C.
- Specific suppressor mutations partially or fully restored growth for subsets of crl mutants, indicating distinct genetic interactions.
Conclusions:
- The identified crl mutations represent novel genes essential for yeast cell viability and cell cycle progression.
- The diverse phenotypes suggest involvement of these genes in multiple cellular pathways.
- Further characterization of these CRL loci will elucidate new aspects of cell cycle control and stress response in eukaryotes.