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Yeast cycloheximide-resistant crl mutants are proteasome mutants defective in protein degradation.

In 1988 McCusker and Haber generated a series of mutants which are resistant to the minimum inhibitory concentration of the protein synthesis inhibitor cycloheximide. These cycloheximide-resistant, temperature-sensitive (crl) mutants, in addition, exhibited other pleiotropic phenotypes, e.g., incorrect response to starvation, hypersensitivity against amino acid analogues, and other protein synthesis inhibitors. Temperature sensitivity of one of these mutants, crl3-2, had been found to be suppressed by a mutation, SCL1-1, which resided in an alpha-type subunit of the 20S proteasome. We cloned the CRL3 gene by complementation and found CRL3 to be identical to the SUG1/CIM3 gene coding for a subunit of the 19S cap complex of the 26S proteasome. Another mutation, crl21, revealed to be allelic with the 20S proteasomal gene PRE3. crl3-2 and crl21 mutant cells show significant defects in proteasome-dependent proteolysis, whereas the SCL1-1 suppressor mutation causes partial restoration of crl3-2-induced proteolytic defects. Notably, cycloheximide resistance was also detected for other proteolytically deficient proteasome mutants (pre1-1, pre2-1, pre3-1, pre4-1). Moreover, proteasomal genes were found within genomic sequences of 9 of 13 chromosomal loci to which crl mutations had been mapped. We therefore assume that most if not all crl mutations reside in the proteasome and that phenotypes found are a result of defective protein degradation.

Pubmed ID: 9398670 RIS Download

Mesh terms: Adenosine Triphosphatases | Alleles | Amino Acid Sequence | Base Sequence | Blotting, Western | Cell Division | Chromosome Mapping | Chromosomes, Fungal | Cloning, Molecular | Cycloheximide | Cysteine Endopeptidases | Drug Resistance, Microbial | Endopeptidases | Fungal Proteins | Genes, Fungal | Microbial Sensitivity Tests | Molecular Sequence Data | Multienzyme Complexes | Mutation | Precipitin Tests | Proteasome Endopeptidase Complex | Repressor Proteins | Saccharomyces cerevisiae | Saccharomyces cerevisiae Proteins | Suppression, Genetic | Temperature | Ubiquitins