Skip to main content
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1985 Dec;5(12):3436–3442. doi: 10.1128/mcb.5.12.3436

Posttranscriptional regulation and assembly into ribosomes of a Saccharomyces cerevisiae ribosomal protein-beta-galactosidase fusion.

L Gritz, N Abovich, J L Teem, M Rosbash
PMCID: PMC369173  PMID: 3939318

Abstract

To study the regulation of ribosomal protein genes, we constructed a 'lacZ fusion of the Saccharomyces cerevisiae RP51A gene, containing the first 64 codons of RP51A. In a strain lacking an intact RP51A gene (cells are viable due to the presence of an active RP51B gene), beta-galactosidase activity is 10-fold greater than in a strain containing RP51A. RP51A-lacZ mRNA levels are equal in the two strains, indicating that regulation is posttranscriptional. In the absence of the RP51A gene, the fusion protein is predominantly cytoplasmic and associated with polysomes, whereas in the presence of RP51A, the fusion protein is predominantly nuclear, and none is associated with polysomes. Deletions were made in the RP51A-coding portion of the fusion gene. The most extensively deleted gene, containing only the first seven RP51A codons fused to lacZ, produced a high level of beta-galactosidase activity in both the presence and the absence of the RP51A gene. In both cases, little or none of this shorter fusion protein was found associated with polysomes. Thus, a regulatory site (or sites) lies in the protein-coding region of RP51A. We suggest that posttranscriptional regulation of the rp51 fusion protein is related to assembly of the protein into ribosomes.

Full text

PDF
3436

Images in this article

Selected References

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

  1. Abovich N., Gritz L., Tung L., Rosbash M. Effect of RP51 gene dosage alterations on ribosome synthesis in Saccharomyces cerevisiae. Mol Cell Biol. 1985 Dec;5(12):3429–3435. doi: 10.1128/mcb.5.12.3429. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Abovich N., Rosbash M. Two genes for ribosomal protein 51 of Saccharomyces cerevisiae complement and contribute to the ribosomes. Mol Cell Biol. 1984 Sep;4(9):1871–1879. doi: 10.1128/mcb.4.9.1871. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bollen G. H., Cohen L. H., Mager W. H., Klaassen A. W., Planta R. J. Isolation of cloned ribosomal protein genes from the yeast Saccharomyces carlsbergensis. Gene. 1981 Sep;14(4):279–287. doi: 10.1016/0378-1119(81)90160-8. [DOI] [PubMed] [Google Scholar]
  4. 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]
  5. Botstein D., Falco S. C., Stewart S. E., Brennan M., Scherer S., Stinchcomb D. T., Struhl K., Davis R. W. Sterile host yeasts (SHY): a eukaryotic system of biological containment for recombinant DNA experiments. Gene. 1979 Dec;8(1):17–24. doi: 10.1016/0378-1119(79)90004-0. [DOI] [PubMed] [Google Scholar]
  6. Bozzoni I., Beccari E., Luo Z. X., Amaldi F. Xenopus laevis ribosomal protein genes: isolation of recombinant cDNA clones and study of the genomic organization. Nucleic Acids Res. 1981 Mar 11;9(5):1069–1086. doi: 10.1093/nar/9.5.1069. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. D'Eustachio P., Meyuhas O., Ruddle F., Perry R. P. Chromosomal distribution of ribosomal protein genes in the mouse. Cell. 1981 May;24(2):307–312. doi: 10.1016/0092-8674(81)90320-2. [DOI] [PubMed] [Google Scholar]
  8. Davies J., Jacob F. Genetic mapping of the regulator and operator genes of the lac operon. J Mol Biol. 1968 Sep 28;36(3):413–417. doi: 10.1016/0022-2836(68)90165-4. [DOI] [PubMed] [Google Scholar]
  9. Fallon A. M., Jinks C. S., Strycharz G. D., Nomura M. Regulation of ribosomal protein synthesis in Escherichia coli by selective mRNA inactivation. Proc Natl Acad Sci U S A. 1979 Jul;76(7):3411–3415. doi: 10.1073/pnas.76.7.3411. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Fried H. M., Pearson N. J., Kim C. H., Warner J. R. The genes for fifteen ribosomal proteins of Saccharomyces cerevisiae. J Biol Chem. 1981 Oct 10;256(19):10176–10183. [PubMed] [Google Scholar]
  11. Gray M. R., Colot H. V., Guarente L., Rosbash M. Open reading frame cloning: identification, cloning, and expression of open reading frame DNA. Proc Natl Acad Sci U S A. 1982 Nov;79(21):6598–6602. doi: 10.1073/pnas.79.21.6598. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gritz L., Davies J. Plasmid-encoded hygromycin B resistance: the sequence of hygromycin B phosphotransferase gene and its expression in Escherichia coli and Saccharomyces cerevisiae. Gene. 1983 Nov;25(2-3):179–188. doi: 10.1016/0378-1119(83)90223-8. [DOI] [PubMed] [Google Scholar]
  13. Guarente L., Lauer G., Roberts T. M., Ptashne M. Improved methods for maximizing expression of a cloned gene: a bacterium that synthesizes rabbit beta-globin. Cell. 1980 Jun;20(2):543–553. doi: 10.1016/0092-8674(80)90640-6. [DOI] [PubMed] [Google Scholar]
  14. Guarente L., Yocum R. R., Gifford P. A GAL10-CYC1 hybrid yeast promoter identifies the GAL4 regulatory region as an upstream site. Proc Natl Acad Sci U S A. 1982 Dec;79(23):7410–7414. doi: 10.1073/pnas.79.23.7410. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hereford L. M., Rosbash M. Number and distribution of polyadenylated RNA sequences in yeast. Cell. 1977 Mar;10(3):453–462. doi: 10.1016/0092-8674(77)90032-0. [DOI] [PubMed] [Google Scholar]
  16. Johnsen M., Christensen T., Dennis P. P., Fiil N. P. Autogenous control: ribosomal protein L10-L12 complex binds to the leader sequence of its mRNA. EMBO J. 1982;1(8):999–1004. doi: 10.1002/j.1460-2075.1982.tb01284.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. 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]
  18. Lindahl L., Archer R., Zengel J. M. Transcription of the S10 ribosomal protein operon is regulated by an attenuator in the leader. Cell. 1983 May;33(1):241–248. doi: 10.1016/0092-8674(83)90353-7. [DOI] [PubMed] [Google Scholar]
  19. Monk R. J., Meyuhas O., Perry R. P. Mammals have multiple genes for individual ribosomal proteins. Cell. 1981 May;24(2):301–306. doi: 10.1016/0092-8674(81)90319-6. [DOI] [PubMed] [Google Scholar]
  20. Mortimer R. K., Hawthorne D. C. Yeast genetics. Annu Rev Microbiol. 1966;20:151–168. doi: 10.1146/annurev.mi.20.100166.001055. [DOI] [PubMed] [Google Scholar]
  21. 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]
  22. Pearson N. J., Fried H. M., Warner J. R. Yeast use translational control to compensate for extra copies of a ribosomal protein gene. Cell. 1982 Jun;29(2):347–355. doi: 10.1016/0092-8674(82)90151-9. [DOI] [PubMed] [Google Scholar]
  23. 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]
  24. Rose M., Casadaban M. J., Botstein D. Yeast genes fused to beta-galactosidase in Escherichia coli can be expressed normally in yeast. Proc Natl Acad Sci U S A. 1981 Apr;78(4):2460–2464. doi: 10.1073/pnas.78.4.2460. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Silver P. A., Keegan L. P., Ptashne M. Amino terminus of the yeast GAL4 gene product is sufficient for nuclear localization. Proc Natl Acad Sci U S A. 1984 Oct;81(19):5951–5955. doi: 10.1073/pnas.81.19.5951. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Stahl H. D., Coppel R. L., Brown G. V., Saint R., Lingelbach K., Cowman A. F., Anders R. F., Kemp D. J. Differential antibody screening of cloned Plasmodium falciparum sequences expressed in Escherichia coli: procedure for isolation of defined antigens and analysis of human antisera. Proc Natl Acad Sci U S A. 1984 Apr;81(8):2456–2460. doi: 10.1073/pnas.81.8.2456. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Teem J. L., Rosbash M. Expression of a beta-galactosidase gene containing the ribosomal protein 51 intron is sensitive to the rna2 mutation of yeast. Proc Natl Acad Sci U S A. 1983 Jul;80(14):4403–4407. doi: 10.1073/pnas.80.14.4403. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Woolford J. L., Jr, Hereford L. M., Rosbash M. Isolation of cloned DNA sequences containing ribosomal protein genes from Saccharomyces cerevisiae. Cell. 1979 Dec;18(4):1247–1259. doi: 10.1016/0092-8674(79)90236-8. [DOI] [PubMed] [Google Scholar]
  31. Yates J. L., Nomura M. Feedback regulation of ribosomal protein synthesis in E. coli: localization of the mRNA target sites for repressor action of ribosomal protein L1. Cell. 1981 Apr;24(1):243–249. doi: 10.1016/0092-8674(81)90520-1. [DOI] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES