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. 1983 Apr;154(1):152–160. doi: 10.1128/jb.154.1.152-160.1983

Expression of the gene for ribosomal protein S20: effects of gene dosage.

G D Parsons, G A Mackie
PMCID: PMC217442  PMID: 6187728

Abstract

We have exploited the properties of three different plasmids which carry the gene for Escherichia coli ribosomal protein S20 (rpsT) to test the effects of gene dosage on the expression of rpsT. Over a range of total copies of rpsT of 1 to 58 per haploid genome equivalent, the rate of incorporation of uridine during a 30-s pulse into RNA annealing to either of two specific probes for S20 mRNA increased essentially in proportion to copy number. In contrast, the rate of synthesis of S20 protein increased no more than 2.1-fold at the highest copy number. We conclude, in contrast to an earlier report (D. Geyl, and A. Böck, Mol. Gen. Genet. 154:327-334, 1977), that the synthesis of S20 is regulated at a posttranscriptional step. We propose that S20 itself is the regulatory agent and that binding of S20 to its own mRNA in regions homologous in structure with 16S rRNA can account for our results.

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Selected References

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

  1. Bachmann B. J., Low K. B. Linkage map of Escherichia coli K-12, edition 6. Microbiol Rev. 1980 Mar;44(1):1–56. doi: 10.1128/mr.44.1.1-56.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bolivar F., Rodriguez R. L., Greene P. J., Betlach M. C., Heyneker H. L., Boyer H. W., Crosa J. H., Falkow S. Construction and characterization of new cloning vehicles. II. A multipurpose cloning system. Gene. 1977;2(2):95–113. [PubMed] [Google Scholar]
  3. Branlant C., Krol A., Ebel J. P. Characterization of the Escherichia coli 23S Ribosomal RNA region associated with ribosomal protein L1. Evidence for homologies with the 5'-end region of the L11 operon. Nucleic Acids Res. 1980 Dec 11;8(23):5567–5577. doi: 10.1093/nar/8.23.5567. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Clewell D. B., Helinski D. R. Supercoiled circular DNA-protein complex in Escherichia coli: purification and induced conversion to an opern circular DNA form. Proc Natl Acad Sci U S A. 1969 Apr;62(4):1159–1166. doi: 10.1073/pnas.62.4.1159. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Cohen S. N., Hurwitz J. Genetic transcription in bacteriophage lambda: studies of lambda m RNA synthesis in vivo. J Mol Biol. 1968 Nov 14;37(3):387–406. doi: 10.1016/0022-2836(68)90110-1. [DOI] [PubMed] [Google Scholar]
  6. 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]
  7. 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]
  8. Dennis P. P., Fill N. P. Transcriptional and post-transcriptional control of RNA polymerase and ribosomal protein genes cloned on composite ColE1 plasmids in the bacterium Escherichia coli. J Biol Chem. 1979 Aug 25;254(16):7540–7547. [PubMed] [Google Scholar]
  9. Dennis P. P. In vivo stability, maturation and relative differential synthesis rates of individual ribosomal proteins in Escherichia coli B/r. J Mol Biol. 1974 Sep 5;88(1):25–41. doi: 10.1016/0022-2836(74)90293-9. [DOI] [PubMed] [Google Scholar]
  10. Dennis P. P., Nomura M. Regulation of the expression of ribosomal protein genes in Escherichia coli. J Mol Biol. 1975 Sep 5;97(1):61–76. doi: 10.1016/s0022-2836(75)80022-2. [DOI] [PubMed] [Google Scholar]
  11. Fallon A. M., Jinks C. S., Yamamoto M., Nomura M. Expression of ribosomal protein genes cloned in a hybrid plasmid in Escherichia coli: gene dosage effects on synthesis of ribosomal proteins and ribosomal protein messenger ribonucleic acid. J Bacteriol. 1979 May;138(2):383–396. doi: 10.1128/jb.138.2.383-396.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Friesen J. D., Parker J., Watson R. J., Fiil N. P., Pedersen S. Isolation of transducing phage carrying rps T, the structural gene for ribosomal protein S20. Mol Gen Genet. 1976 Feb 27;144(1):115–118. doi: 10.1007/BF00277314. [DOI] [PubMed] [Google Scholar]
  13. Geyl D., Böck A. Synthesis of ribosomal proteins in merodiploid strains and in minicells of Escherichia coli. Mol Gen Genet. 1977 Sep 9;154(3):327–334. doi: 10.1007/BF00571290. [DOI] [PubMed] [Google Scholar]
  14. Gillespie D., Spiegelman S. A quantitative assay for DNA-RNA hybrids with DNA immobilized on a membrane. J Mol Biol. 1965 Jul;12(3):829–842. doi: 10.1016/s0022-2836(65)80331-x. [DOI] [PubMed] [Google Scholar]
  15. Glotz C., Brimacombe R. An experimentally-derived model for the secondary structure of the 16S ribosomal RNA from Escherichia coli. Nucleic Acids Res. 1980 Jun 11;8(11):2377–2395. doi: 10.1093/nar/8.11.2377. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hines J. C., Ray D. S. Construction and characterization of new coliphage M13 cloning vectors. Gene. 1980 Nov;11(3-4):207–218. doi: 10.1016/0378-1119(80)90061-x. [DOI] [PubMed] [Google Scholar]
  17. Kennel D. Titration of the gene sites on DNA by DNA-RNA hybridization. II. The Escherichia coli chromosome. J Mol Biol. 1968 May 28;34(1):85–103. doi: 10.1016/0022-2836(68)90236-2. [DOI] [PubMed] [Google Scholar]
  18. 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]
  19. Mackie G. A. Cloning of fragments of lambda dapB2 DNA and identification of the dapB gene product. J Biol Chem. 1980 Sep 25;255(18):8928–8935. [PubMed] [Google Scholar]
  20. Mackie G. A. Nucleotide sequence of the gene for ribosomal protein S20 and its flanking regions. J Biol Chem. 1981 Aug 10;256(15):8177–8182. [PubMed] [Google Scholar]
  21. Mackie G. A., Parsons G. D. Identification of gene products from cloned fragments of the left arm of lambda dapB2. Can J Biochem. 1982 Mar;60(3):338–346. doi: 10.1139/o82-040. [DOI] [PubMed] [Google Scholar]
  22. Mackie G. A., Zimmermann R. A. RNA--protein interactions in the ribosome. IV. Structure and properties of binding sites for proteins S4, S16/S17 and S20 in the 16S RNA. J Mol Biol. 1978 May 5;121(1):17–39. doi: 10.1016/0022-2836(78)90260-7. [DOI] [PubMed] [Google Scholar]
  23. Messing J., Crea R., Seeburg P. H. A system for shotgun DNA sequencing. Nucleic Acids Res. 1981 Jan 24;9(2):309–321. doi: 10.1093/nar/9.2.309. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Mets L. J., Bogorad L. Two-dimensional polyacrylamide gel electrophoresis: an improved method for ribosomal proteins. Anal Biochem. 1974 Jan;57(1):200–210. doi: 10.1016/0003-2697(74)90065-7. [DOI] [PubMed] [Google Scholar]
  25. Miura A., Krueger J. H., Itoh S., de Boer H. A., Nomura M. Growth-rate-dependent regulation of ribosome synthesis in E. coli: expression of the lacZ and galK genes fused to ribosomal promoters. Cell. 1981 Sep;25(3):773–782. doi: 10.1016/0092-8674(81)90185-9. [DOI] [PubMed] [Google Scholar]
  26. Muto A., Ehresmann C., Fellner P., Zimmermann R. A. RNA-protein interactions in the ribosome. I. Characterization and ribonuclease digestion of 16 S RNA-ribosomal protein complexes. J Mol Biol. 1974 Jun 25;86(2):411–432. doi: 10.1016/0022-2836(74)90028-x. [DOI] [PubMed] [Google Scholar]
  27. Noller H. F., Woese C. R. Secondary structure of 16S ribosomal RNA. Science. 1981 Apr 24;212(4493):403–411. doi: 10.1126/science.6163215. [DOI] [PubMed] [Google Scholar]
  28. Nomura M., Morgan E. A. Genetics of bacterial ribosomes. Annu Rev Genet. 1977;11:297–347. doi: 10.1146/annurev.ge.11.120177.001501. [DOI] [PubMed] [Google Scholar]
  29. Nomura M., Yates J. L., Dean D., Post L. E. Feedback regulation of ribosomal protein gene expression in Escherichia coli: structural homology of ribosomal RNA and ribosomal protein MRNA. Proc Natl Acad Sci U S A. 1980 Dec;77(12):7084–7088. doi: 10.1073/pnas.77.12.7084. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Olins P. O., Nomura M. Regulation of the S10 ribosomal protein operon in E. coli: nucleotide sequence at the start of the operon. Cell. 1981 Oct;26(2 Pt 2):205–211. doi: 10.1016/0092-8674(81)90303-2. [DOI] [PubMed] [Google Scholar]
  31. Olins P. O., Nomura M. Translational regulation by ribosomal protein S8 in Escherichia coli: structural homology between rRNA binding site and feedback target on mRNA. Nucleic Acids Res. 1981 Apr 10;9(7):1757–1764. doi: 10.1093/nar/9.7.1757. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Rose J. K., Mosteller R. D., Yanofsky C. Tryptophan messenger ribonucleic acid elongation rates and steady-state levels of tryptophan operon enzymes under various growth conditions. J Mol Biol. 1970 Aug;51(3):541–550. doi: 10.1016/0022-2836(70)90007-0. [DOI] [PubMed] [Google Scholar]
  33. Schwarzbauer J., Craven G. R. Apparent association constants for E. coli ribosomal proteins S4, S7, S8, S15, S17 and S20 binding to 16S RNA. Nucleic Acids Res. 1981 May 11;9(9):2223–2237. doi: 10.1093/nar/9.9.2223. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Shepard H. M., Polisky B. Measurement of Plasmid copy number. Methods Enzymol. 1979;68:503–513. doi: 10.1016/0076-6879(79)68039-4. [DOI] [PubMed] [Google Scholar]
  35. Shine J., Dalgarno L. Determinant of cistron specificity in bacterial ribosomes. Nature. 1975 Mar 6;254(5495):34–38. doi: 10.1038/254034a0. [DOI] [PubMed] [Google Scholar]
  36. Vandekerckhove J., Rombauts W., Peeters B., Wittmann-Liebold B. Determination of the complete amino acid sequence of protein S21 from Escherichia coli ribosomes. Hoppe Seylers Z Physiol Chem. 1975 Dec;356(12):1955–1976. doi: 10.1515/bchm2.1975.356.2.1955. [DOI] [PubMed] [Google Scholar]
  37. 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]
  38. Wirth R., Kohles V., Böck A. Factors modulating transcription and translation in vitro of ribosomal protein S20 and isoleucyl-tRNA synthetase from Escherichia coli. Eur J Biochem. 1981 Feb;114(2):429–437. doi: 10.1111/j.1432-1033.1981.tb05164.x. [DOI] [PubMed] [Google Scholar]
  39. Wittmann-Liebold B., Marzinzig E., Lehmann A. Primary structure of protein S20 from the small ribosomal subunit of Escherichia coli. FEBS Lett. 1976 Sep 15;68(1):110–114. doi: 10.1016/0014-5793(76)80416-4. [DOI] [PubMed] [Google Scholar]
  40. 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]
  41. 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]
  42. Zimmermann R. A., Mackie G. A., Muto A., Garrett R. A., Ungewickell E., Ehresmann C., Stiegler P., Ebel J. P., Fellner P. Location and characteristics of ribosomal protein binding sites in the 16S RNA of Escherichia coli. Nucleic Acids Res. 1975 Feb;2(2):279–302. doi: 10.1093/nar/2.2.279. [DOI] [PMC free article] [PubMed] [Google Scholar]

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