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Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1975 Aug;72(8):2900–2904. doi: 10.1073/pnas.72.8.2900

Very stable prokaryotic messenger RNA in chromosomeless Escherichia coli minicells.

S B Levy
PMCID: PMC432886  PMID: 1103125

Abstract

E. coli minicells lack DNA, yet they make protein, the synthesis of which is sensitive to chloramphenicol but insensitive to rifamycin. This protein is coded for by very stable cellular mRNA with an estimated half-life of 40-80 min. In an R factor-containing minicell, two very different species of mRNA are observed: (i) R factor-specific mRNA with a short half-life whose synthesis is rifamycin-sensitive and (ii) cellular mRNA with a long half-life whose synthesis is rifamycin-insensitive. These findings indicate that minicells contain normal degradative mechanisms for mRNA and point out the existence of a unique class of very stable cellular mRNA. Greater than 80% of the rifamycin-insensitive protein synthesized goes into the outer minicell membrane. Relatively stable mRNA, half-life 5.5-11.5 min, for outer membrane protein in whole cells has been reported [Hirashima et al. (1973) J. Mol. Biol. 79, 373-389]. The stability of minicell mRNA is significantly greater. This and other observations suggest that there are two functional species of mRNA for outer membrane protein perhaps in different sites in the cell. Furthermore, these studies suggest that a class of cellular proteins is synthesized in bacteria without concomitant transcription and in the absence of association with chromosomal DNA.

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Selected References

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

  1. ARONSON A. I., ROSASDELVALLE M. RNA AND PROTEIN SYNTHESIS REQUIRED FOR BACTERIAL SPORE FORMATION. Biochim Biophys Acta. 1964 Jun 22;87:267–276. doi: 10.1016/0926-6550(64)90222-1. [DOI] [PubMed] [Google Scholar]
  2. Adler H. I., Fisher W. D., Cohen A., Hardigree A. A. MINIATURE escherichia coli CELLS DEFICIENT IN DNA. Proc Natl Acad Sci U S A. 1967 Feb;57(2):321–326. doi: 10.1073/pnas.57.2.321. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bosch V., Braun V. Distribution of murein-lipoprotein between the cytoplasmic and outer membrane of Escherichia coli. FEBS Lett. 1973 Aug 15;34(2):307–310. doi: 10.1016/0014-5793(73)80818-x. [DOI] [PubMed] [Google Scholar]
  4. Both G. W., McInnes J. L., Hanlon J. E., May B. K., Elliott W. H. Evidence for an accumulation of messenger RNA specific for extracellular protease and its relevance to the mechanism of enzyme secretion in bacteria. J Mol Biol. 1972 Jun 20;67(2):199–217. doi: 10.1016/0022-2836(72)90236-7. [DOI] [PubMed] [Google Scholar]
  5. Botstein D. Synthesis and maturation of phage P22 DNA. I. Identification of intermediates. J Mol Biol. 1968 Jun 28;34(3):621–641. doi: 10.1016/0022-2836(68)90185-x. [DOI] [PubMed] [Google Scholar]
  6. Braun V., Bosch V. In vivo biosynthesis of murein-lipoprotein of the outer membrane of E. coli. FEBS Lett. 1973 Aug 15;34(2):302–306. doi: 10.1016/0014-5793(73)80817-8. [DOI] [PubMed] [Google Scholar]
  7. Braun V., Rehn K. Chemical characterization, spatial distribution and function of a lipoprotein (murein-lipoprotein) of the E. coli cell wall. The specific effect of trypsin on the membrane structure. Eur J Biochem. 1969 Oct;10(3):426–438. doi: 10.1111/j.1432-1033.1969.tb00707.x. [DOI] [PubMed] [Google Scholar]
  8. Cancedda R., Schlesinger M. J. Localization of polyribosomes containing alkaline phosphatase nascent polypeptides on membranes of Escherichia coli. J Bacteriol. 1974 Jan;117(1):290–301. doi: 10.1128/jb.117.1.290-301.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Cohen S. N., Silver R. P., Sharp P. A., McCoubrey A. E. The problems of drug-resistant pathogenic bacteria. Studies on the molecular nature of R factors. Ann N Y Acad Sci. 1971 Jun 11;182:172–187. doi: 10.1111/j.1749-6632.1971.tb30655.x. [DOI] [PubMed] [Google Scholar]
  10. Dvorak H. F., Wetzel B. K., Heppel L. A. Biochemical and cytochemical evidence for the polar concentration of periplasmic enzymes in a "minicell" strain of Escherichia coli. J Bacteriol. 1970 Oct;104(1):543–548. doi: 10.1128/jb.104.1.543-548.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Filip C., Fletcher G., Wulff J. L., Earhart C. F. Solubilization of the cytoplasmic membrane of Escherichia coli by the ionic detergent sodium-lauryl sarcosinate. J Bacteriol. 1973 Sep;115(3):717–722. doi: 10.1128/jb.115.3.717-722.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. GROS F., HIATT H., GILBERT W., KURLAND C. G., RISEBROUGH R. W., WATSON J. D. Unstable ribonucleic acid revealed by pulse labelling of Escherichia coli. Nature. 1961 May 13;190:581–585. doi: 10.1038/190581a0. [DOI] [PubMed] [Google Scholar]
  13. Glew R. H., Heath E. C. Studies on the extracellular alkaline phosphatase of Micrococcus sodonensis. II. Factors affecting secretion. J Biol Chem. 1971 Mar 25;246(6):1566–1574. [PubMed] [Google Scholar]
  14. Goodell E. W., Schwarz U., Teather R. M. Cell envelope composition of Escherichia coli K12: a comparison of the cell poles and the lateral wall. Eur J Biochem. 1974 Sep 16;47(3):567–572. doi: 10.1111/j.1432-1033.1974.tb03727.x. [DOI] [PubMed] [Google Scholar]
  15. HARRIS H., SABATH L. D. INDUCED ENZYME SYNTHESIS IN THE ABSENCE OF CONCOMITANT RIBONUCLEIC ACID SYNTHESIS. Nature. 1964 Jun 13;202:1078–1080. doi: 10.1038/2021078a0. [DOI] [PubMed] [Google Scholar]
  16. Hirashima A., Childs G., Inouye M. Differential inhibitory effects of antibiotics on the biosynthesis of envelope proteins of Escherichia coli. J Mol Biol. 1973 Sep 15;79(2):373–389. doi: 10.1016/0022-2836(73)90012-0. [DOI] [PubMed] [Google Scholar]
  17. Inouye M., Shaw J., Shen C. The assembly of a structural lipoprotein in the envelope of Escherichia coli. J Biol Chem. 1972 Dec 25;247(24):8154–8159. [PubMed] [Google Scholar]
  18. Inselburg J. Segregation into and replication of plasmid deoxyribonucleic acid in chromosomeless segregants of Escherichia coli. J Bacteriol. 1970 Jun;102(3):642–647. doi: 10.1128/jb.102.3.642-647.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. KEPES A. KINETICS OF INDUCED ENZYME SYNTHESIS. DETERMINATION OF THE MEAN LIFE OF GALACTOSIDASE-SPECIFIC MESSENGER RNA. Biochim Biophys Acta. 1963 Oct 15;76:293–309. [PubMed] [Google Scholar]
  20. Kass L. R., Yarmolinsky M. B. Segregation of functional sex factor into minicells. Proc Natl Acad Sci U S A. 1970 Jul;66(3):815–822. doi: 10.1073/pnas.66.3.815. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Kool A. J., van Zeben M. S., Nijkamp H. J. Identification of messenger ribonucleic acids and proteins synthesized by the bacteriocinogenic factor Clo DF13 in purified minicells of Escherichia coli. J Bacteriol. 1974 Apr;118(1):213–224. doi: 10.1128/jb.118.1.213-224.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Lee N., Inouye M. Outer membrane proteins of Escherichia coli: biosynthesis and assembly. FEBS Lett. 1974 Feb 15;39(2):167–170. doi: 10.1016/0014-5793(74)80043-8. [DOI] [PubMed] [Google Scholar]
  23. Levy S. B., McMurry L., Palmer E. R factor proteins synthesized in Escherichia coli minicells: membrane-associated R factor proteins. J Bacteriol. 1974 Dec;120(3):1464–1471. doi: 10.1128/jb.120.3.1464-1471.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Levy S. B., Norman P. Segregation of transferable R factors into Escherichia coli minicells. Nature. 1970 Aug 8;227(5258):606–607. doi: 10.1038/227606a0. [DOI] [PubMed] [Google Scholar]
  25. Levy S. B. Physical and functional characteristics of R-factor deoxyribonucleic acid segregated into Escherichia coli minicells. J Bacteriol. 1971 Oct;108(1):300–308. doi: 10.1128/jb.108.1.300-308.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Levy S. B. R factor proteins synthesized in Escherichia coli minicells: incorporation studies with different R factors and detection of deoxyribonucleic acid-binding proteins. J Bacteriol. 1974 Dec;120(3):1451–1463. doi: 10.1128/jb.120.3.1451-1463.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Miller O. L., Jr, Hamkalo B. A., Thomas C. A., Jr Visualization of bacterial genes in action. Science. 1970 Jul 24;169(3943):392–395. doi: 10.1126/science.169.3943.392. [DOI] [PubMed] [Google Scholar]
  28. NAKADA D., MAGASANIK B. THE ROLES OF INDUCER AND CATABOLITE REPRESSOR IN THE SYNTHESIS OF BETA-GALACTOSIDASE BY ESCHERICHIA COLI. J Mol Biol. 1964 Jan;8:105–127. doi: 10.1016/s0022-2836(64)80153-4. [DOI] [PubMed] [Google Scholar]
  29. Osborn M. J., Gander J. E., Parisi E., Carson J. Mechanism of assembly of the outer membrane of Salmonella typhimurium. Isolation and characterization of cytoplasmic and outer membrane. J Biol Chem. 1972 Jun 25;247(12):3962–3972. [PubMed] [Google Scholar]
  30. Penman S. RNA metabolism in the HeLa cell nucleus. J Mol Biol. 1966 May;17(1):117–130. doi: 10.1016/s0022-2836(66)80098-0. [DOI] [PubMed] [Google Scholar]
  31. Roozen K. J., Fenwick R. G., Jr, Curtiss R., 3rd Synthesis of ribonucleic acid and protein in plasmid-containing minicells of Escherichia coli K-12. J Bacteriol. 1971 Jul;107(1):21–33. doi: 10.1128/jb.107.1.21-33.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. SCHAECHTER M., PREVIC E. P., GILLESPIE M. E. MESSENGER RNA AND POLYRIBOSOMES IN BACILLUS MEGATERIUM. J Mol Biol. 1965 May;12:119–129. doi: 10.1016/s0022-2836(65)80286-8. [DOI] [PubMed] [Google Scholar]
  33. Stent G. S. Genetic transcription. Proc R Soc Lond B Biol Sci. 1966 Mar 22;164(995):181–197. doi: 10.1098/rspb.1966.0022. [DOI] [PubMed] [Google Scholar]
  34. Studier F. W. Analysis of bacteriophage T7 early RNAs and proteins on slab gels. J Mol Biol. 1973 Sep 15;79(2):237–248. doi: 10.1016/0022-2836(73)90003-x. [DOI] [PubMed] [Google Scholar]
  35. Summers W. C. The process of infection with coliphage T7. IV. Stability of RNA in bacteriophage-infected cells. J Mol Biol. 1970 Aug;51(3):671–678. doi: 10.1016/0022-2836(70)90015-x. [DOI] [PubMed] [Google Scholar]
  36. Wehrli W., Nüesch J., Knüsel F., Staehelin M. Action of rifamycins on RNA polymerase. Biochim Biophys Acta. 1968 Mar 18;157(1):215–217. doi: 10.1016/0005-2787(68)90285-2. [DOI] [PubMed] [Google Scholar]
  37. Zubay G., Chambers D. A. A DNA-directed cell-free system for beta-galactosidase synthesis; characterization of the de novo synthesized enzyme and some aspects of the regulation of synthesis. Cold Spring Harb Symp Quant Biol. 1969;34:753–761. doi: 10.1101/sqb.1969.034.01.085. [DOI] [PubMed] [Google Scholar]

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