Skip to main content
NCBI home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1980 Dec 20;8(24):6081–6097. doi: 10.1093/nar/8.24.6081

Conformational alteration of mRNA structure and the posttranscriptional regulation of erythromycin-induced drug resistance.

T J Gryczan, G Grandi, J Hahn, R Grandi, D Dubnau
PMCID: PMC328074  PMID: 6162157

Abstract

The DNA sequence of the ermC gene of plasmid pE194 is presented. This determinant is responsible for erythromycin-induced resistance to the macrolide-lincosamide-streptogramin B group of antibiotics and specifies a 29,000 dalton inducible protein. The locations of the ermC promoter, as well as that of a probable transcriptional terminator, are established both from the sequence and by transcription mapping. The sequence contains an open reading frame sufficient to encode the previously identified 29,000 dalton ermC protein. Between the promoter and the putative ATG start codon is a 141 base pair leader sequence, within which several regulatory (constitutive) mutations have been mapped and sequenced. The leader has a second open reading frame, sufficient to encode a 19 amino acid peptide. It is suggested that induction by erythromycin involves a shift between alternative ribosome-bound mRNA conformations, so that the ribosome binding sequence and the start codon for synthesis of the 29K protein are unmasked in the presence of inducer. Possible active and inactive folded configuration of the leader sequence are presented, as well as the effects on these configurations of regulatory mutations.

Full text

PDF
6081

Images in this article

6088Image
on p.6088
6089Image
on p.6089

Selected References

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

  1. Alwine J. C., Kemp D. J., Parker B. A., Reiser J., Renart J., Stark G. R., Wahl G. M. Detection of specific RNAs or specific fragments of DNA by fractionation in gels and transfer to diazobenzyloxymethyl paper. Methods Enzymol. 1979;68:220–242. doi: 10.1016/0076-6879(79)68017-5. [DOI] [PubMed] [Google Scholar]
  2. Barnes W. M. DNA sequence from the histidine operon control region: seven histidine codons in a row. Proc Natl Acad Sci U S A. 1978 Sep;75(9):4281–4285. doi: 10.1073/pnas.75.9.4281. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Berk A. J., Sharp P. A. Sizing and mapping of early adenovirus mRNAs by gel electrophoresis of S1 endonuclease-digested hybrids. Cell. 1977 Nov;12(3):721–732. doi: 10.1016/0092-8674(77)90272-0. [DOI] [PubMed] [Google Scholar]
  4. Carmichael G. G., McMaster G. K. The analysis of nucleic acids in gels using glyoxal and acridine orange. Methods Enzymol. 1980;65(1):380–391. doi: 10.1016/s0076-6879(80)65049-6. [DOI] [PubMed] [Google Scholar]
  5. Cerná J., Rychlík I., Pulkrábek P. The effect of antibiotics on the coded binding of peptidyl-tRNA to the ribosome and on the transfer of the peptidyl residue to puromycin. Eur J Biochem. 1969 May 1;9(1):27–35. doi: 10.1111/j.1432-1033.1969.tb00571.x. [DOI] [PubMed] [Google Scholar]
  6. Contente S., Dubnau D. Marker rescue transformation by linear plasmid DNA in Bacillus subtilis. Plasmid. 1979 Oct;2(4):555–571. doi: 10.1016/0147-619x(79)90054-4. [DOI] [PubMed] [Google Scholar]
  7. Di Nocera P. P., Blasi F., Di Lauro R., Frunzio R., Bruni C. B. Nucleotide sequence of the attenuator region of the histidine operon of Escherichia coli K-12. Proc Natl Acad Sci U S A. 1978 Sep;75(9):4276–4280. doi: 10.1073/pnas.75.9.4276. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Favaloro J., Treisman R., Kamen R. Transcription maps of polyoma virus-specific RNA: analysis by two-dimensional nuclease S1 gel mapping. Methods Enzymol. 1980;65(1):718–749. doi: 10.1016/s0076-6879(80)65070-8. [DOI] [PubMed] [Google Scholar]
  9. Gardner J. F. Regulation of the threonine operon: tandem threonine and isoleucine codons in the control region and translational control of transcription termination. Proc Natl Acad Sci U S A. 1979 Apr;76(4):1706–1710. doi: 10.1073/pnas.76.4.1706. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Gemmill R. M., Wessler S. R., Keller E. B., Calvo J. M. leu operon of Salmonella typhimurium is controlled by an attenuation mechanism. Proc Natl Acad Sci U S A. 1979 Oct;76(10):4941–4945. doi: 10.1073/pnas.76.10.4941. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Gryczan T. J., Contente S., Dubnau D. Characterization of Staphylococcus aureus plasmids introduced by transformation into Bacillus subtilis. J Bacteriol. 1978 Apr;134(1):318–329. doi: 10.1128/jb.134.1.318-329.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gryczan T., Shivakumar A. G., Dubnau D. Characterization of chimeric plasmid cloning vehicles in Bacillus subtilis. J Bacteriol. 1980 Jan;141(1):246–253. doi: 10.1128/jb.141.1.246-253.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Haldenwang W. G., Losick R. A modified RNA polymerase transcribes a cloned gene under sporulation control in Bacillus subtilis. Nature. 1979 Nov 15;282(5736):256–260. doi: 10.1038/282256a0. [DOI] [PubMed] [Google Scholar]
  14. Iordănescu S. Three distinct plasmids originating in the same Staphylococcus aureus strain. Arch Roum Pathol Exp Microbiol. 1976 Jan-Jun;35(1-2):111–118. [PubMed] [Google Scholar]
  15. Lai C. J., Dahlberg J. E., Weisblum B. Structure of an inducibly methylatable nucleotide sequence in 23S ribosomal ribonucleic acid from erythromycin-resistant Staphylococcus aureus. Biochemistry. 1973 Jan 30;12(3):457–460. doi: 10.1021/bi00727a015. [DOI] [PubMed] [Google Scholar]
  16. Lai C. J., Weisblum B. Altered methylation of ribosomal RNA in an erythromycin-resistant strain of Staphylococcus aureus. Proc Natl Acad Sci U S A. 1971 Apr;68(4):856–860. doi: 10.1073/pnas.68.4.856. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Lai C. J., Weisblum B., Fahnestock S. R., Nomura M. Alteration of 23 S ribosomal RNA and erythromycin-induced resistance to lincomycin and spiramycin in Staphylococcus aureus. J Mol Biol. 1973 Feb 15;74(1):67–72. doi: 10.1016/0022-2836(73)90355-0. [DOI] [PubMed] [Google Scholar]
  18. Maniatis T., Efstratiadis A. Fractionation of low molecular weight DNA or RNA in polyacrylamide gels containing 98% formamide or 7 M urea. Methods Enzymol. 1980;65(1):299–305. doi: 10.1016/s0076-6879(80)65040-x. [DOI] [PubMed] [Google Scholar]
  19. Maniatis T., Jeffrey A., Kleid D. G. Nucleotide sequence of the rightward operator of phage lambda. Proc Natl Acad Sci U S A. 1975 Mar;72(3):1184–1188. doi: 10.1073/pnas.72.3.1184. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
  21. Maxwell I. H., Maxwell F., Hahn W. E. Removal of RNase activity from DNase by affinity chromatography on agarose coupled aminophenylphosphoryl-uridine-2' (3')-phosphate. Nucleic Acids Res. 1977 Jan;4(1):241–246. doi: 10.1093/nar/4.1.241. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
  23. Rosenberg M., Court D. Regulatory sequences involved in the promotion and termination of RNA transcription. Annu Rev Genet. 1979;13:319–353. doi: 10.1146/annurev.ge.13.120179.001535. [DOI] [PubMed] [Google Scholar]
  24. Sanger F., Coulson A. R. The use of thin acrylamide gels for DNA sequencing. FEBS Lett. 1978 Mar 1;87(1):107–110. doi: 10.1016/0014-5793(78)80145-8. [DOI] [PubMed] [Google Scholar]
  25. 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]
  26. Shine J., Dalgarno L. The 3'-terminal sequence of Escherichia coli 16S ribosomal RNA: complementarity to nonsense triplets and ribosome binding sites. Proc Natl Acad Sci U S A. 1974 Apr;71(4):1342–1346. doi: 10.1073/pnas.71.4.1342. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Shivakumar A. G., Gryczan T. J., Kozlov Y. I., Dubnau D. Organization of the pE194 genome. Mol Gen Genet. 1980;179(2):241–252. doi: 10.1007/BF00425450. [DOI] [PubMed] [Google Scholar]
  28. Shivakumar A. G., Hahn J., Dubnau D. Studies on the synthesis of plasmid-coded proteins and their control in Bacillus subtilis minicells. Plasmid. 1979 Apr;2(2):279–289. doi: 10.1016/0147-619x(79)90046-5. [DOI] [PubMed] [Google Scholar]
  29. Shivakumar A. G., Hahn J., Grandi G., Kozlov Y., Dubnau D. Posttranscriptional regulation of an erythromycin resistance protein specified by plasmic pE194. Proc Natl Acad Sci U S A. 1980 Jul;77(7):3903–3907. doi: 10.1073/pnas.77.7.3903. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Sprague K. U., Steitz J. A., Grenley R. M., Stocking C. E. 3' terminal sequences of 16S rRNA do not explain translational specificity differences between E. coli and B. stearothermophilus ribosomes. Nature. 1977 Jun 2;267(5610):462–465. doi: 10.1038/267462a0. [DOI] [PubMed] [Google Scholar]
  31. Sutcliffe J. G. pBR322 restriction map derived from the DNA sequence: accurate DNA size markers up to 4361 nucleotide pairs long. Nucleic Acids Res. 1978 Aug;5(8):2721–2728. doi: 10.1093/nar/5.8.2721. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Talkington C., Pero J. Distinctive nucleotide sequences of promoters recognized by RNA polymerase containing a phage-coded "sigma-like" protein. Proc Natl Acad Sci U S A. 1979 Nov;76(11):5465–5469. doi: 10.1073/pnas.76.11.5465. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Weisblum B., Graham M. Y., Gryczan T., Dubnau D. Plasmid copy number control: isolation and characterization of high-copy-number mutants of plasmid pE194. J Bacteriol. 1979 Jan;137(1):635–643. doi: 10.1128/jb.137.1.635-643.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Woese C., Sogin M., Stahl D., Lewis B. J., Bonen L. A comparison of the 16S ribosomal RNAs from mesophilic and thermophilic bacilli: some modifications in the Sanger method for RNA sequencing. J Mol Evol. 1976 Apr 9;7(3):197–213. doi: 10.1007/BF01731489. [DOI] [PubMed] [Google Scholar]
  35. Zurawski G., Brown K., Killingly D., Yanofsky C. Nucleotide sequence of the leader region of the phenylalanine operon of Escherichia coli. Proc Natl Acad Sci U S A. 1978 Sep;75(9):4271–4275. doi: 10.1073/pnas.75.9.4271. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Zurawski G., Elseviers D., Stauffer G. V., Yanofsky C. Translational control of transcription termination at the attenuator of the Escherichia coli tryptophan operon. Proc Natl Acad Sci U S A. 1978 Dec;75(12):5988–5992. doi: 10.1073/pnas.75.12.5988. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES