A trichodermin-resistant mutant of Saccharomyces cerevisiae with an abnormal distribution of native ribosomal...

1. A yeast mutant (CLP-8), resistant at the ribosome level to the trichothecene antibiotic trichodermin, differs from its parent in having an unusual distribution of native ribosomal subunits. Sucrose gradient analysis of cytoplasmic extracts from this mutant revealed a large excess of material sedimenting at 60 S with little or no material sedimenting at 40 S. 2. The excess 609-S material consists predominantly of functionally active 60-S ribosomal subunits, as indicated by both analysis of ribosomal RNA and studies in vitro using a poly(U)-directed protein-synthesizing system. 3. Using the poly(U) system it was found that high-salt-washed particles derived from either the excess 60-S peak or 80-S ribosomes of CLP-8 exhibited very similar levels of resistance to the antibiotic fusarenon-X, a drug closely related chemically to trichodermin. The same level of resistance to fusarenon-X was also shown by high-salt-washed 60-S ribosomal particles obtained from a further trichodermin-resistant yeast strain (TR-1), although this strain has a normal distribution of native ribosomal subunits. In addition, both CLP-8 and TR-1 are equally resistant to inhibition of protein synthesis by trichothecene antibiotics, as assayed in vivo. 4. Genetic analysis of CLP-8 indicates that the trichodermin-resistant trait can be segregated from the lesion responsible for the inbalance of native ribosomal subunits. However, the latter defect is only expressed phenotypically in cells that retain the trichodermin-resistant character. 5. CLP-8 has a further defect in that both in vivo and in vitro it fails to generate native 40-S ribosomal subunits from 80-S particles. There may be a lesion in the protein factor normally required for this process.