Skip to main content
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1988 Nov;170(11):5042–5050. doi: 10.1128/jb.170.11.5042-5050.1988

Effects of induction of rRNA overproduction on ribosomal protein synthesis and ribosome subunit assembly in Escherichia coli.

M Yamagishi 1, M Nomura 1
PMCID: PMC211569  PMID: 3053641

Abstract

Overproduction of rRNA was artificially induced in Escherichia coli cells to test whether the synthesis of ribosomal protein (r-protein) is normally repressed by feedback regulation. When rRNA was overproduced more than twofold from a hybrid plasmid carrying the rrnB operon fused to the lambda pL promoter (pL-rrnB), synthesis of individual r-proteins increased by an average of about 60%. This demonstrates that the synthesis of r-proteins is repressed under normal conditions. The increase of r-protein production, however, for unknown reasons, was not as great as the increase in rRNA synthesis and resulted in an imbalance between the amounts of rRNA and r-protein synthesis. Therefore, only a small (less than 20%) increase in the synthesis of complete 30S and 50S ribosome subunits was detected, and a considerable fraction of the excess rRNA was degraded. Lack of complete cooperativity in the assembly of ribosome subunits in vivo is discussed as a possible explanation for the absence of a large stimulation of ribosome synthesis observed under these conditions. In addition to the induction of intact rRNA overproduction from the pL-rrnB operon, the effects of unbalanced overproduction of each of the two large rRNAs, 16S rRNA and 23S rRNA, on r-protein synthesis were examined using pL-rrnB derivatives carrying a large deletion in either the 23S rRNA gene or the 16S rRNA gene. Operon-specific derepression after 23S or 16S rRNA overproduction correlated with the overproduction of rRNA containing the target site for the operon-specific repressor r-protein. These results are discussed to explain the apparent coupling of the assembly of one ribosomal subunit with that of the other which was observed in earlier studies on conditionally lethal mutants with defects in ribosome assembly.

Full text

PDF
5042

Selected References

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

  1. Baughman G., Nomura M. Translational regulation of the L11 ribosomal protein operon of Escherichia coli: analysis of the mRNA target site using oligonucleotide-directed mutagenesis. Proc Natl Acad Sci U S A. 1984 Sep;81(17):5389–5393. doi: 10.1073/pnas.81.17.5389. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Brot N., Caldwell P., Weissbach H. Autogenous control of Escherichia coli ribosomal protein L10 synthesis in vitro. Proc Natl Acad Sci U S A. 1980 May;77(5):2592–2595. doi: 10.1073/pnas.77.5.2592. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cole J. R., Nomura M. Translational regulation is responsible for growth-rate-dependent and stringent control of the synthesis of ribosomal proteins L11 and L1 in Escherichia coli. Proc Natl Acad Sci U S A. 1986 Jun;83(12):4129–4133. doi: 10.1073/pnas.83.12.4129. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Cole J. R., Olsson C. L., Hershey J. W., Grunberg-Manago M., Nomura M. Feedback regulation of rRNA synthesis in Escherichia coli. Requirement for initiation factor IF2. J Mol Biol. 1987 Dec 5;198(3):383–392. doi: 10.1016/0022-2836(87)90288-9. [DOI] [PubMed] [Google Scholar]
  5. Dean D., Nomura M. Feedback regulation of ribosomal protein gene expression in Escherichia coli. Proc Natl Acad Sci U S A. 1980 Jun;77(6):3590–3594. doi: 10.1073/pnas.77.6.3590. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Dean D., Yates J. L., Nomura M. Escherichia coli ribosomal protein S8 feedback regulates part of spc operon. Nature. 1981 Jan 1;289(5793):89–91. doi: 10.1038/289089a0. [DOI] [PubMed] [Google Scholar]
  7. Dean D., Yates J. L., Nomura M. Identification of ribosomal protein S7 as a repressor of translation within the str operon of E. coli. Cell. 1981 May;24(2):413–419. doi: 10.1016/0092-8674(81)90331-7. [DOI] [PubMed] [Google Scholar]
  8. Freedman L. P., Zengel J. M., Archer R. H., Lindahl L. Autogenous control of the S10 ribosomal protein operon of Escherichia coli: genetic dissection of transcriptional and posttranscriptional regulation. Proc Natl Acad Sci U S A. 1987 Sep;84(18):6516–6520. doi: 10.1073/pnas.84.18.6516. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Fukuda R. Autogenous regulation of the synthesis of ribosomal proteins, L10 and L7/12, in Escherichia coli. Mol Gen Genet. 1980;178(2):483–486. doi: 10.1007/BF00270505. [DOI] [PubMed] [Google Scholar]
  10. Gourse R. L., Nomura M. Level of rRNA, not tRNA, synthesis controls transcription of rRNA and tRNA operons in Escherichia coli. J Bacteriol. 1984 Dec;160(3):1022–1026. doi: 10.1128/jb.160.3.1022-1026.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Gourse R. L., Takebe Y., Sharrock R. A., Nomura M. Feedback regulation of rRNA and tRNA synthesis and accumulation of free ribosomes after conditional expression of rRNA genes. Proc Natl Acad Sci U S A. 1985 Feb;82(4):1069–1073. doi: 10.1073/pnas.82.4.1069. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Guthrie C., Nashimoto H., Nomura M. Structure and function of E. coli ribosomes. 8. Cold-sensitive mutants defective in ribosome assembly. Proc Natl Acad Sci U S A. 1969 Jun;63(2):384–391. doi: 10.1073/pnas.63.2.384. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Howard G. A., Traut R. R. Separation and radioautography of microgram quantities of ribosomal proteins by two-dimensional polyacrylamide gel electrophoresis. FEBS Lett. 1973 Jan 15;29(2):177–180. doi: 10.1016/0014-5793(73)80555-1. [DOI] [PubMed] [Google Scholar]
  14. Hui A., Jhurani P., de Boer H. A. Directing ribosomes to a single mRNA species: a method to study ribosomal RNA mutations and their effects on translation of a single messenger in Escherichia coli. Methods Enzymol. 1987;153:432–452. doi: 10.1016/0076-6879(87)53070-1. [DOI] [PubMed] [Google Scholar]
  15. Hui A., de Boer H. A. Specialized ribosome system: preferential translation of a single mRNA species by a subpopulation of mutated ribosomes in Escherichia coli. Proc Natl Acad Sci U S A. 1987 Jul;84(14):4762–4766. doi: 10.1073/pnas.84.14.4762. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Jinks-Robertson S., Gourse R. L., Nomura M. Expression of rRNA and tRNA genes in Escherichia coli: evidence for feedback regulation by products of rRNA operons. Cell. 1983 Jul;33(3):865–876. doi: 10.1016/0092-8674(83)90029-6. [DOI] [PubMed] [Google Scholar]
  17. Jinks-Robertson S., Nomura M. Ribosomal protein S4 acts in trans as a translational repressor to regulate expression of the alpha operon in Escherichia coli. J Bacteriol. 1982 Jul;151(1):193–202. doi: 10.1128/jb.151.1.193-202.1982. [DOI] [PMC free article] [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. Lindahl L., Zengel J. M. Operon-specific regulation of ribosomal protein synthesis in Escherichia coli. Proc Natl Acad Sci U S A. 1979 Dec;76(12):6542–6546. doi: 10.1073/pnas.76.12.6542. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Lindahl L., Zengel J. M. Ribosomal genes in Escherichia coli. Annu Rev Genet. 1986;20:297–326. doi: 10.1146/annurev.ge.20.120186.001501. [DOI] [PubMed] [Google Scholar]
  21. Nashimoto H., Nomura M. Structure and function of bacterial ribosomes. XI. Dependence of 50S ribosomal assembly on simultaneous assembly of 30S subunits. Proc Natl Acad Sci U S A. 1970 Nov;67(3):1440–1447. doi: 10.1073/pnas.67.3.1440. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Neidhardt F. C., Bloch P. L., Smith D. F. Culture medium for enterobacteria. J Bacteriol. 1974 Sep;119(3):736–747. doi: 10.1128/jb.119.3.736-747.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Nomura M., Gourse R., Baughman G. Regulation of the synthesis of ribosomes and ribosomal components. Annu Rev Biochem. 1984;53:75–117. doi: 10.1146/annurev.bi.53.070184.000451. [DOI] [PubMed] [Google Scholar]
  24. Nomura M., Traub P., Guthrie C., Nashimoto H. The assembly of ribosomes. J Cell Physiol. 1969 Oct;74(2 Suppl):241+–241+. doi: 10.1002/jcp.1040740428. [DOI] [PubMed] [Google Scholar]
  25. Nowotny V., Nierhaus K. H. Initiator proteins for the assembly of the 50S subunit from Escherichia coli ribosomes. Proc Natl Acad Sci U S A. 1982 Dec;79(23):7238–7242. doi: 10.1073/pnas.79.23.7238. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Olsson M. O., Isaksson L. A. Analysis of rpsD mutations in Escherichia coli. III. Effects of rpsD mutations on expression of some ribosomal protein genes. Mol Gen Genet. 1979 Feb 1;169(3):271–278. doi: 10.1007/BF00382273. [DOI] [PubMed] [Google Scholar]
  27. Pardo D., Vola C., Rosset R. Assembly of ribosomal subunits affected in a ribosomal mutant of E. coli having an altered L22 protein. Mol Gen Genet. 1979 Jul 2;174(1):53–58. doi: 10.1007/BF00433305. [DOI] [PubMed] [Google Scholar]
  28. Siehnel R. J., Morgan E. A. Unbalanced rRNA gene dosage and its effects on rRNA and ribosomal-protein synthesis. J Bacteriol. 1985 Aug;163(2):476–486. doi: 10.1128/jb.163.2.476-486.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Steen R., Dahlberg A. E., Lade B. N., Studier F. W., Dunn J. J. T7 RNA polymerase directed expression of the Escherichia coli rrnB operon. EMBO J. 1986 May;5(5):1099–1103. doi: 10.1002/j.1460-2075.1986.tb04328.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Wirth R., Böck A. Regulation of synthesis of ribosomal protein S20 in vitro. Mol Gen Genet. 1980;178(2):479–481. doi: 10.1007/BF00270504. [DOI] [PubMed] [Google Scholar]
  31. Yamagishi M., Cole J. R., Nomura M., Studier F. W., Dunn J. J. Stringent control in Escherichia coli applies also to transcription by T7 RNA polymerase. J Biol Chem. 1987 Mar 25;262(9):3940–3943. [PubMed] [Google Scholar]
  32. Yamagishi M., de Boer H. A., Nomura M. Feedback regulation of rRNA synthesis. A mutational alteration in the anti-Shine-Dalgarno region of the 16 S rRNA gene abolishes regulation. J Mol Biol. 1987 Dec 5;198(3):547–550. doi: 10.1016/0022-2836(87)90299-3. [DOI] [PubMed] [Google Scholar]
  33. Yates J. L., Arfsten A. E., Nomura M. In vitro expression of Escherichia coli ribosomal protein genes: autogenous inhibition of translation. Proc Natl Acad Sci U S A. 1980 Apr;77(4):1837–1841. doi: 10.1073/pnas.77.4.1837. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Yates J. L., Dean D., Strycharz W. A., Nomura M. E. coli ribosomal protein L10 inhibits translation of L10 and L7/L12 mRNAs by acting at a single site. Nature. 1981 Nov 12;294(5837):190–192. doi: 10.1038/294190a0. [DOI] [PubMed] [Google Scholar]
  35. Yates J. L., Nomura M. E. coli ribosomal protein L4 is a feedback regulatory protein. Cell. 1980 Sep;21(2):517–522. doi: 10.1016/0092-8674(80)90489-4. [DOI] [PubMed] [Google Scholar]
  36. Zengel J. M., Archer R. H., Freedman L. P., Lindahl L. Role of attenuation in growth rate-dependent regulation of the S10 r-protein operon of E. coli. EMBO J. 1984 Jul;3(7):1561–1565. doi: 10.1002/j.1460-2075.1984.tb02011.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Zengel J. M., Lindahl L. Transcription of ribosomal genes during a nutritional shift-up of Escherichia coli. J Bacteriol. 1986 Sep;167(3):1095–1097. doi: 10.1128/jb.167.3.1095-1097.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Zengel J. M., Mueckl D., Lindahl L. Protein L4 of the E. coli ribosome regulates an eleven gene r protein operon. Cell. 1980 Sep;21(2):523–535. doi: 10.1016/0092-8674(80)90490-0. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES