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. 1987 Nov;6(11):3233–3237. doi: 10.1002/j.1460-2075.1987.tb02640.x

The molecular basis for rRNA-dependent spectinomycin resistance in Nicotiana chloroplasts

Hillel Fromm 1, Marvin Edelman 1, Dvora Aviv 1, Esra Galun 1
PMCID: PMC553774  PMID: 16453804

Abstract

The chloroplast genes coding for the 16S ribosomal RNA from several spectinomycin-resistant Nicotiana mutants were analyzed. Two classes of mutants were identified. In one class, a G to A base transition is found at position 1140 of the tobacco-chloroplast 16S rRNA gene, which eliminates an AatII restriction endonuclease site. This base transition is proximal to a mutation previously described for spectinomycin resistance in Escherichia coli. In the other class, a novel G to A transition is found at position 1012 of the 16S rRNA gene. Although the mutations in the two classes are 128 nucleotides apart, the secondary structure model for 16S rRNA suggests that the two mutated nucleotides are in spatial proximity on opposite sides of a conserved stem structure in the 3' region of the molecule. Phylogenetic evidence is presented linking this conserved stem with spectinomycin resistance in chloroplasts. Perturbation of the stem is proposed to be the molecular-genetic basis for rRNA-dependent spectinomycin resistance.

Keywords: antibiotic resistance, chloroplast mutation, 16S ribosomal RNA

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Selected References

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

  1. Anderson P. Sensitivity and Resistance to Spectinomycin in Escherichia coli. J Bacteriol. 1969 Nov;100(2):939–947. doi: 10.1128/jb.100.2.939-947.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bollen A., Davies J., Ozaki M., Mizushima S. Ribosomal Protein Conferring Sensitivity to the Antibiotic Spectinomycin in Escherichia coli. Science. 1969 Jul 4;165(3888):85–86. doi: 10.1126/science.165.3888.85. [DOI] [PubMed] [Google Scholar]
  3. Brosius J., Dull T. J., Sleeter D. D., Noller H. F. Gene organization and primary structure of a ribosomal RNA operon from Escherichia coli. J Mol Biol. 1981 May 15;148(2):107–127. doi: 10.1016/0022-2836(81)90508-8. [DOI] [PubMed] [Google Scholar]
  4. Davies J., Anderson P., Davis B. D. Inhibition of protein synthesis by spectinomycin. Science. 1965 Sep 3;149(3688):1096–1098. doi: 10.1126/science.149.3688.1096. [DOI] [PubMed] [Google Scholar]
  5. Dente L., Cesareni G., Cortese R. pEMBL: a new family of single stranded plasmids. Nucleic Acids Res. 1983 Mar 25;11(6):1645–1655. doi: 10.1093/nar/11.6.1645. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Dron M., Rahire M., Rochaix J. D. Sequence of the chloroplast 16S rRNA gene and its surrounding regions of Chlamydomonas reinhardii. Nucleic Acids Res. 1982 Dec 11;10(23):7609–7620. doi: 10.1093/nar/10.23.7609. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Fluhr R., Aviv D., Galun E., Edelman M. Efficient induction and selection of chloroplast-encoded antibiotic-resistant mutants in Nicotiana. Proc Natl Acad Sci U S A. 1985 Mar;82(5):1485–1489. doi: 10.1073/pnas.82.5.1485. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Fluhr R., Fromm H., Edelman M. Clone bank of Nicotiana tabacum chloroplast DNA: mapping of the alpha, beta and epsilon subunits of the ATPase coupling factor, the large subunit of ribulosebisphosphate carboxylase, and the 32-kDal membrane protein. Gene. 1983 Nov;25(2-3):271–280. doi: 10.1016/0378-1119(83)90231-7. [DOI] [PubMed] [Google Scholar]
  9. Funatsu G., Schiltz E., Wittmann H. G. Ribosomal proteins. XXVII. Localization of the amino acid exchanges in protein S5 from two Escherichia coli mutants resistant to spectinomycin. Mol Gen Genet. 1972;114(2):106–111. doi: 10.1007/BF00332781. [DOI] [PubMed] [Google Scholar]
  10. Gillham N. W. Induction of chromosomal and nonchromosomal mutations in Chlamydomonas reinhardi with N-methyl-N'-nitro-N-nitrosoguanidine. Genetics. 1965 Sep;52(3):529–537. doi: 10.1093/genetics/52.3.529. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Goldthwaite C., Dubnau D., Smith I. Genetic mapping of antibiotic resistance in markers Bacillus subtilis. Proc Natl Acad Sci U S A. 1970 Jan;65(1):96–103. doi: 10.1073/pnas.65.1.96. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Graf L., Roux E., Stutz E., Kössel H. Nucleotide sequence of a Euglena gracilis chloroplast gene coding for the 16S rRNA: homologies to E. coli and Zea mays chloroplast 16S rRNA. Nucleic Acids Res. 1982 Oct 25;10(20):6369–6381. doi: 10.1093/nar/10.20.6369. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kramer W., Schughart K., Fritz H. J. Directed mutagenesis of DNA cloned in filamentous phage: influence of hemimethylated GATC sites on marker recovery from restriction fragments. Nucleic Acids Res. 1982 Oct 25;10(20):6475–6485. doi: 10.1093/nar/10.20.6475. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Moazed D., Noller H. F. Interaction of antibiotics with functional sites in 16S ribosomal RNA. Nature. 1987 Jun 4;327(6121):389–394. doi: 10.1038/327389a0. [DOI] [PubMed] [Google Scholar]
  15. Montandon P. E., Nicolas P., Schürmann P., Stutz E. Streptomycin-resistance of Euglena gracilis chloroplasts: identification of a point mutation in the 16S rRNA gene in an invariant position. Nucleic Acids Res. 1985 Jun 25;13(12):4299–4310. doi: 10.1093/nar/13.12.4299. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Noller H. F. Structure of ribosomal RNA. Annu Rev Biochem. 1984;53:119–162. doi: 10.1146/annurev.bi.53.070184.001003. [DOI] [PubMed] [Google Scholar]
  17. Orozco E. M., Jr, Mullet J. E., Hanley-Bowdoin L., Chua N. H. In vitro transcription of chloroplast protein genes. Methods Enzymol. 1986;118:232–253. doi: 10.1016/0076-6879(86)18076-1. [DOI] [PubMed] [Google Scholar]
  18. Roberts B. E., Paterson B. M. Efficient translation of tobacco mosaic virus RNA and rabbit globin 9S RNA in a cell-free system from commercial wheat germ. Proc Natl Acad Sci U S A. 1973 Aug;70(8):2330–2334. doi: 10.1073/pnas.70.8.2330. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. 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]
  20. Schlanger G., Sager R. Localization of five antibiotic resistances at the subunit level in chloroplast ribosomes of Chlamydomonas. Proc Natl Acad Sci U S A. 1974 May;71(5):1715–1719. doi: 10.1073/pnas.71.5.1715. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Shinozaki K., Ohme M., Tanaka M., Wakasugi T., Hayashida N., Matsubayashi T., Zaita N., Chunwongse J., Obokata J., Yamaguchi-Shinozaki K. The complete nucleotide sequence of the tobacco chloroplast genome: its gene organization and expression. EMBO J. 1986 Sep;5(9):2043–2049. doi: 10.1002/j.1460-2075.1986.tb04464.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Sigmund C. D., Ettayebi M., Morgan E. A. Antibiotic resistance mutations in 16S and 23S ribosomal RNA genes of Escherichia coli. Nucleic Acids Res. 1984 Jun 11;12(11):4653–4663. doi: 10.1093/nar/12.11.4653. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Spangler E. A., Blackburn E. H. The nucleotide sequence of the 17S ribosomal RNA gene of Tetrahymena thermophila and the identification of point mutations resulting in resistance to the antibiotics paromomycin and hygromycin. J Biol Chem. 1985 May 25;260(10):6334–6340. [PubMed] [Google Scholar]
  24. Tohdoh N., Sugiura M. The complete nucleotide sequence of 16S ribosomal RNA gene from tobacco chloroplasts. Gene. 1982 Feb;17(2):213–218. doi: 10.1016/0378-1119(82)90074-9. [DOI] [PubMed] [Google Scholar]
  25. Tomioka N., Sugiura M. The complete nucleotide sequence of a 16S ribosomal RNA gene from a blue-green alga, Anacystis nidulans. Mol Gen Genet. 1983;191(1):46–50. doi: 10.1007/BF00330888. [DOI] [PubMed] [Google Scholar]
  26. Yamada T., Davies J. A genetic and biochemical study of streptomycin- and spectinomycin-resistance in Salmonella typhimurium. Mol Gen Genet. 1971;110(3):197–210. doi: 10.1007/BF00337833. [DOI] [PubMed] [Google Scholar]

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