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. 1985 Apr;5(4):816–822. doi: 10.1128/mcb.5.4.816

Isolation of the SUP45 omnipotent suppressor gene of Saccharomyces cerevisiae and characterization of its gene product.

H J Himmelfarb, E Maicas, J D Friesen
PMCID: PMC366786  PMID: 3887137

Abstract

The Saccharomyces cerevisiae SUP45+ gene has been isolated from a genomic clone library by genetic complementation of paromomycin sensitivity, which is a property of a mutant strain carrying the sup45-2 allele. This plasmid complements all phenotypes associated with the sup45-2 mutation, including nonsense suppression, temperature sensitivity, osmotic sensitivity, and paromomycin sensitivity. Genetic mapping with a URA3+-marked derivative of the complementing plasmid that was integrated into the chromosome by homologous recombination demonstrated that the complementing fragment contained the SUP45+ gene and not an unlinked suppressor. The SUP45+ gene is present as a single copy in the haploid genome and is essential for viability. In vitro translation of the hybrid-selected SUP45+ transcript yielded a protein of Mr = 54,000, which is larger than any known ribosomal protein. RNA blot hybridization analysis showed that the steady-state level of the SUP45+ transcript is less than 10% of that for ribosomal protein L3 or rp59 transcripts. When yeast cells are subjected to a mild heat shock, the synthesis rate of the SUP45+ transcript was transiently reduced, approximately in parallel with ribosomal protein transcripts. Our data suggest that the SUP45+ gene does not encode a ribosomal protein. We speculate that it codes for a translation-related function whose precise nature is not yet known.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bollen A., Cabezón T., de Wilde M., Villarroel R., Herzog A. Alteration of ribosomal protein S17 by mutation linked to neamine resistance in Escherichia coli. I. General properties of neaA mutants. J Mol Biol. 1975 Dec 25;99(4):795–806. doi: 10.1016/s0022-2836(75)80185-9. [DOI] [PubMed] [Google Scholar]
  2. Bollen G. H., Molenaar C. M., Cohen L. H., van Raamsdonk-Duin M. M., Mager W. H., Planta R. J. Ribosomal protein genes of yeast contain intervening sequences. Gene. 1982 Apr;18(1):29–37. doi: 10.1016/0378-1119(82)90053-1. [DOI] [PubMed] [Google Scholar]
  3. Cabezón T., Herzog A., De Wilde M., Villarroel R., Bollen A. Cooperative control of translational fidelity by ribosomal proteins in Escherichia coli. III. A ram mutation in the structural gene for protein S5 (rpx E). Mol Gen Genet. 1976 Feb 27;144(1):59–62. doi: 10.1007/BF00277305. [DOI] [PubMed] [Google Scholar]
  4. Chattoo B. B., Palmer E., Ono B., Sherman F. Patterns of Genetic and Phenotypic Suppression of lys2 Mutations in the Yeast SACCHAROMYCES CEREVISIAE. Genetics. 1979 Sep;93(1):67–79. doi: 10.1093/genetics/93.1.67. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Davies J., Gorini L., Davis B. D. Misreading of RNA codewords induced by aminoglycoside antibiotics. Mol Pharmacol. 1965 Jul;1(1):93–106. [PubMed] [Google Scholar]
  6. Davis R. W., Thomas M., Cameron J., St John T. P., Scherer S., Padgett R. A. Rapid DNA isolations for enzymatic and hybridization analysis. Methods Enzymol. 1980;65(1):404–411. doi: 10.1016/s0076-6879(80)65051-4. [DOI] [PubMed] [Google Scholar]
  7. Edelmann P., Gallant J. Mistranslation in E. coli. Cell. 1977 Jan;10(1):131–137. doi: 10.1016/0092-8674(77)90147-7. [DOI] [PubMed] [Google Scholar]
  8. Erhart E., Hollenberg C. P. The presence of a defective LEU2 gene on 2 mu DNA recombinant plasmids of Saccharomyces cerevisiae is responsible for curing and high copy number. J Bacteriol. 1983 Nov;156(2):625–635. doi: 10.1128/jb.156.2.625-635.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Fried H. M., Warner J. R. Cloning of yeast gene for trichodermin resistance and ribosomal protein L3. Proc Natl Acad Sci U S A. 1981 Jan;78(1):238–242. doi: 10.1073/pnas.78.1.238. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Gerlach W. L. Genetic properties of some amber-ochre supersuppressors in Saccharomyces cerevisiae. Mol Gen Genet. 1975;138(1):53–63. doi: 10.1007/BF00268827. [DOI] [PubMed] [Google Scholar]
  11. Gorenstein C., Warner J. R. Coordinate regulation of the synthesis of eukaryotic ribosomal proteins. Proc Natl Acad Sci U S A. 1976 May;73(5):1547–1551. doi: 10.1073/pnas.73.5.1547. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hardy S. J. The stoichiometry of the ribosomal proteins of Escherichia coli. Mol Gen Genet. 1975 Oct 3;140(3):253–274. doi: 10.1007/BF00334270. [DOI] [PubMed] [Google Scholar]
  13. Hartwell L. H. Macromolecule synthesis in temperature-sensitive mutants of yeast. J Bacteriol. 1967 May;93(5):1662–1670. doi: 10.1128/jb.93.5.1662-1670.1967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hartwell L. H., McLaughlin C. S., Warner J. R. Identification of ten genes that control ribosome formation in yeast. Mol Gen Genet. 1970;109(1):42–56. doi: 10.1007/BF00334045. [DOI] [PubMed] [Google Scholar]
  15. Hawthorne D. C., Leupold U. Suppressors in yeast. Curr Top Microbiol Immunol. 1974;64(0):1–47. doi: 10.1007/978-3-642-65848-8_1. [DOI] [PubMed] [Google Scholar]
  16. Hawthorne D. C., Mortimer R. K. Genetic mapping of nonsense suppressors in yeast. Genetics. 1968 Dec;60(4):735–742. doi: 10.1093/genetics/60.4.735. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Himmelfarb H. J., Vassarotti A., Friesen J. D. Molecular cloning and biosynthetic regulation of cry1 gene of Saccharomyces cerevisiae. Mol Gen Genet. 1984;195(3):500–506. doi: 10.1007/BF00341453. [DOI] [PubMed] [Google Scholar]
  18. Hopfield J. J. Kinetic proofreading: a new mechanism for reducing errors in biosynthetic processes requiring high specificity. Proc Natl Acad Sci U S A. 1974 Oct;71(10):4135–4139. doi: 10.1073/pnas.71.10.4135. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Kim C. H., Warner J. R. Mild temperature shock alters the transcription of a discrete class of Saccharomyces cerevisiae genes. Mol Cell Biol. 1983 Mar;3(3):457–465. doi: 10.1128/mcb.3.3.457. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Kühberger R., Piepersberg W., Petzet A., Buckel P., Böck A. Alteration of ribosomal protein L6 in gentamicin-resistant strains of Escherichia coli. Effects on fidelity of protein synthesis. Biochemistry. 1979 Jan 9;18(1):187–193. doi: 10.1021/bi00568a028. [DOI] [PubMed] [Google Scholar]
  21. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  22. Larkin J. C., Woolford J. L., Jr Molecular cloning and analysis of the CRY1 gene: a yeast ribosomal protein gene. Nucleic Acids Res. 1983 Jan 25;11(2):403–420. doi: 10.1093/nar/11.2.403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Ma C., Mortimer R. K. Empirical equation that can be used to determine genetic map distances from tetrad data. Mol Cell Biol. 1983 Oct;3(10):1886–1887. doi: 10.1128/mcb.3.10.1886. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Masurekar M., Palmer E., Ono B. I., Wilhelm J. M., Sherman F. Misreading of the ribosomal suppressor SUP46 due to an altered 40 S subunit in yeast. J Mol Biol. 1981 Apr 15;147(3):381–390. doi: 10.1016/0022-2836(81)90490-3. [DOI] [PubMed] [Google Scholar]
  25. McNeil J. B., Friesen J. D. Expression of the Herpes simplex virus thymidine kinase gene in Saccharomyces cerevisiae. Mol Gen Genet. 1981;184(3):386–393. doi: 10.1007/BF00352510. [DOI] [PubMed] [Google Scholar]
  26. Ng R., Abelson J. Isolation and sequence of the gene for actin in Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1980 Jul;77(7):3912–3916. doi: 10.1073/pnas.77.7.3912. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. O'Farrell P. H. High resolution two-dimensional electrophoresis of proteins. J Biol Chem. 1975 May 25;250(10):4007–4021. [PMC free article] [PubMed] [Google Scholar]
  28. O'Farrell P. Z., Goodman H. M., O'Farrell P. H. High resolution two-dimensional electrophoresis of basic as well as acidic proteins. Cell. 1977 Dec;12(4):1133–1141. doi: 10.1016/0092-8674(77)90176-3. [DOI] [PubMed] [Google Scholar]
  29. Ono B. I., Stewart J. W., Sherman F. Serine insertion caused by the ribosomal suppressor SUP46 in yeast. J Mol Biol. 1981 Apr 15;147(3):373–379. doi: 10.1016/0022-2836(81)90489-7. [DOI] [PubMed] [Google Scholar]
  30. Orr-Weaver T. L., Szostak J. W., Rothstein R. J. Yeast transformation: a model system for the study of recombination. Proc Natl Acad Sci U S A. 1981 Oct;78(10):6354–6358. doi: 10.1073/pnas.78.10.6354. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Palmer E., Wilhelm J. M., Sherman F. Phenotypic suppression of nonsense mutants in yeast by aminoglycoside antibiotics. Nature. 1979 Jan 11;277(5692):148–150. doi: 10.1038/277148a0. [DOI] [PubMed] [Google Scholar]
  32. Paterson B. M., Bishop J. O. Changes in the mRNA population of chick myoblasts during myogenesis in vitro. Cell. 1977 Nov;12(3):751–765. doi: 10.1016/0092-8674(77)90275-6. [DOI] [PubMed] [Google Scholar]
  33. Piepersberg W., Böck A., Wittmann H. G. Effect of different mutations in ribosomal protein S5 of Escherichia coli on translational fidelity. Mol Gen Genet. 1975 Sep 29;140(2):91–100. doi: 10.1007/BF00329777. [DOI] [PubMed] [Google Scholar]
  34. Reeh S., Pedersen S., Friesen J. D. Biosynthetic regulation of individual proteins in relA+ and relA strains of Escherichia coli during amino acid starvation. Mol Gen Genet. 1976 Dec 22;149(3):279–289. doi: 10.1007/BF00268529. [DOI] [PubMed] [Google Scholar]
  35. Rosbash M., Harris P. K., Woolford J. L., Jr, Teem J. L. The effect of temperature-sensitive RNA mutants on the transcription products from cloned ribosomal protein genes of yeast. Cell. 1981 Jun;24(3):679–686. doi: 10.1016/0092-8674(81)90094-5. [DOI] [PubMed] [Google Scholar]
  36. Rosset R., Gorini L. A ribosomal ambiguity mutation. J Mol Biol. 1969 Jan 14;39(1):95–112. doi: 10.1016/0022-2836(69)90336-2. [DOI] [PubMed] [Google Scholar]
  37. Schultz L. D., Friesen J. D. Nucleotide sequence of the tcml gene (ribosomal protein L3) of Saccharomyces cerevisiae. J Bacteriol. 1983 Jul;155(1):8–14. doi: 10.1128/jb.155.1.8-14.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Shulman R. W., Sripati C. E., Warner J. R. Noncoordinated transcription in the absence of protein synthesis in yeast. J Biol Chem. 1977 Feb 25;252(4):1344–1349. [PubMed] [Google Scholar]
  39. Singh A., Ursic D., Davies J. Phenotypic suppression and misreading Saccharomyces cerevisiae. Nature. 1979 Jan 11;277(5692):146–148. doi: 10.1038/277146a0. [DOI] [PubMed] [Google Scholar]
  40. Surguchov A. P., Beretetskaya Y. V., Fominykch E. S., Pospelova E. M., SmirnovVN, Ter-Avanesyan M. D., Inge-Vechtomov S. G. Recessive suppression in yeast Saccharomyces cerevisiae is mediated by a ribosomal mutation. FEBS Lett. 1980 Feb 25;111(1):175–178. doi: 10.1016/0014-5793(80)80786-1. [DOI] [PubMed] [Google Scholar]
  41. Surguchov A. P., Fominykch E. S., Smirnov V. N., Ter-Avanesyan M. D., Mironova L. N., Inge-Vechtomov S. G. Further characterization of recessive suppression in yeast. Isolation of the low-temperature sensitive mutant of Saccharomyces cerevisiae defective in the assembly of 60 S ribosomal subunit. Biochim Biophys Acta. 1981 Jun 26;654(1):149–155. doi: 10.1016/0005-2787(81)90148-9. [DOI] [PubMed] [Google Scholar]
  42. Surguchov A. P., Pospelova E. M., Smirnov V. N. Synergistic action of genetic and phenotypic suppression of nonsense mutations in yeast Saccharomyces cerevisiae. Mol Gen Genet. 1981;183(1):197–198. doi: 10.1007/BF00270162. [DOI] [PubMed] [Google Scholar]
  43. Ter-Avanesyan M. D., Zimmermann J., Inge-Vechtomov S. G., Sudarikov A. B., Smirnov V. N., Surguchov A. P. Ribosomal recessive suppressors cause a respiratory deficiency in yeast Saccharomyces cerevisiae. Mol Gen Genet. 1982;185(2):319–323. doi: 10.1007/BF00330805. [DOI] [PubMed] [Google Scholar]
  44. Thompson R. C., Karim A. M. The accuracy of protein biosynthesis is limited by its speed: high fidelity selection by ribosomes of aminoacyl-tRNA ternary complexes containing GTP[gamma S]. Proc Natl Acad Sci U S A. 1982 Aug;79(16):4922–4926. doi: 10.1073/pnas.79.16.4922. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Warner J. R., Gorenstein C. The synthesis of eucaryotic ribosomal proteins in vitro. Cell. 1977 May;11(1):201–212. doi: 10.1016/0092-8674(77)90331-2. [DOI] [PubMed] [Google Scholar]

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