Skip to main content
NCBI home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1995 May;177(10):2737–2743. doi: 10.1128/jb.177.10.2737-2743.1995

Characterization of the umu-complementing operon from R391.

O I Kulaeva 1, J C Wootton 1, A S Levine 1, R Woodgate 1
PMCID: PMC176944  PMID: 7751283

Abstract

In addition to conferring resistances to antibiotics and heavy metals, certain R factors carry genes involved in mutagenic DNA repair. These plasmid-encoded genes are structurally and functionally related to the chromosomally encoded umuDC genes of Escherichia coli and Salmonella typhimurium. Three such plasmid operons, mucAB, impCAB, and samAB, have been characterized at the molecular level. Recently, we have identified three additional umu-complementing operons from IncJ plasmid R391 and IncL/M plasmids R446b and R471a. We report here the molecular characterization of the R391 umu-complementing operon. The nucleotide sequence of the minimal R plasmid umu-complementing (rum) region revealed an operon of two genes, rumA(R391) and rumB(R391), with an upstream regulatory signal strongly resembling LexA-binding sites. Phylogenetic analysis revealed that the RumAB(R391) proteins are approximately equally diverged in sequence from the chromosomal UmuDC proteins and the other plasmid-encoded Umu-like proteins and represent a new subfamily. Genetic characterization of the rumAB(R391) operon revealed that in recA+ and recA1730 backgrounds, the rumAB(R391) operon was phenotypically indistinguishable from mucAB. In contrast, however, the rumAB(R391) operon gave levels of mutagenesis that were intermediate between those given by mucAB and umuDC in a recA430 strain. The latter phenotype was shown to correlate with the reduced posttranslational processing of the RumA(R391) protein to its mutagenically active form, RumA'(R391). Thus, the rumAB(R391) operon appears to possess characteristics that are reminiscent of both chromosome and plasmid-encoded umu-like operons.

Full Text

The Full Text of this article is available as a PDF (458.1 KB).

Selected References

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

  1. Bailone A., Sommer S., Knezević J., Dutreix M., Devoret R. A RecA protein mutant deficient in its interaction with the UmuDC complex. Biochimie. 1991 Apr;73(4):479–484. doi: 10.1016/0300-9084(91)90115-h. [DOI] [PubMed] [Google Scholar]
  2. Ben-Bassat A., Bauer K., Chang S. Y., Myambo K., Boosman A., Chang S. Processing of the initiation methionine from proteins: properties of the Escherichia coli methionine aminopeptidase and its gene structure. J Bacteriol. 1987 Feb;169(2):751–757. doi: 10.1128/jb.169.2.751-757.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Blanco M., Herrera G., Aleixandre V. Different efficiency of UmuDC and MucAB proteins in UV light induced mutagenesis in Escherichia coli. Mol Gen Genet. 1986 Nov;205(2):234–239. doi: 10.1007/BF00430433. [DOI] [PubMed] [Google Scholar]
  4. Blanco M., Herrera G., Collado P., Rebollo J. E., Botella L. M. Influence of RecA protein on induced mutagenesis. Biochimie. 1982 Aug-Sep;64(8-9):633–636. doi: 10.1016/s0300-9084(82)80102-8. [DOI] [PubMed] [Google Scholar]
  5. Blanco M., Rebollo J. E. Plasmid pKM101-dependent repair and mutagenesis in Escherichia coli cells with mutations lexB30 tif and zab-53 in the recA gene. Mutat Res. 1981 May;81(3):265–275. doi: 10.1016/0027-5107(81)90115-9. [DOI] [PubMed] [Google Scholar]
  6. Bridges B. A., Woodgate R. Mutagenic repair in Escherichia coli: products of the recA gene and of the umuD and umuC genes act at different steps in UV-induced mutagenesis. Proc Natl Acad Sci U S A. 1985 Jun;82(12):4193–4197. doi: 10.1073/pnas.82.12.4193. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Bridges B. A., Woodgate R. The two-step model of bacterial UV mutagenesis. Mutat Res. 1985 Jun-Jul;150(1-2):133–139. doi: 10.1016/0027-5107(85)90110-1. [DOI] [PubMed] [Google Scholar]
  8. Brunner D. P., Traxler B. A., Holt S. M., Crose L. L. Enhancement of UV survival, UV- and MMS-mutability, precise excision of Tn10 and complementation of umuC by plasmids of different incompatibility groups. Mutat Res. 1986 Jul;166(1):29–37. doi: 10.1016/0167-8817(86)90038-6. [DOI] [PubMed] [Google Scholar]
  9. Burckhardt S. E., Woodgate R., Scheuermann R. H., Echols H. UmuD mutagenesis protein of Escherichia coli: overproduction, purification, and cleavage by RecA. Proc Natl Acad Sci U S A. 1988 Mar;85(6):1811–1815. doi: 10.1073/pnas.85.6.1811. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Coetzee J. N., Datta N., Hedges R. W. R factors from Proteus rettgeri. J Gen Microbiol. 1972 Oct;72(3):543–552. doi: 10.1099/00221287-72-3-543. [DOI] [PubMed] [Google Scholar]
  11. Dutreix M., Burnett B., Bailone A., Radding C. M., Devoret R. A partially deficient mutant, recA1730, that fails to form normal nucleoprotein filaments. Mol Gen Genet. 1992 Apr;232(3):489–497. doi: 10.1007/BF00266254. [DOI] [PubMed] [Google Scholar]
  12. Ennis D. G., Fisher B., Edmiston S., Mount D. W. Dual role for Escherichia coli RecA protein in SOS mutagenesis. Proc Natl Acad Sci U S A. 1985 May;82(10):3325–3329. doi: 10.1073/pnas.82.10.3325. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. FALKOW S., RYMAN I. R., WASHINGTON O. Deoxyribonucleic acid base composition of Proteus and Providence organisms. J Bacteriol. 1962 Jun;83:1318–1321. doi: 10.1128/jb.83.6.1318-1321.1962. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Felsenstein J. Phylogenies from molecular sequences: inference and reliability. Annu Rev Genet. 1988;22:521–565. doi: 10.1146/annurev.ge.22.120188.002513. [DOI] [PubMed] [Google Scholar]
  15. Frank E. G., Hauser J., Levine A. S., Woodgate R. Targeting of the UmuD, UmuD', and MucA' mutagenesis proteins to DNA by RecA protein. Proc Natl Acad Sci U S A. 1993 Sep 1;90(17):8169–8173. doi: 10.1073/pnas.90.17.8169. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hauser J., Levine A. S., Ennis D. G., Chumakov K. M., Woodgate R. The enhanced mutagenic potential of the MucAB proteins correlates with the highly efficient processing of the MucA protein. J Bacteriol. 1992 Nov;174(21):6844–6851. doi: 10.1128/jb.174.21.6844-6851.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Ho C., Kulaeva O. I., Levine A. S., Woodgate R. A rapid method for cloning mutagenic DNA repair genes: isolation of umu-complementing genes from multidrug resistance plasmids R391, R446b, and R471a. J Bacteriol. 1993 Sep;175(17):5411–5419. doi: 10.1128/jb.175.17.5411-5419.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Kato T., Shinoura Y. Isolation and characterization of mutants of Escherichia coli deficient in induction of mutations by ultraviolet light. Mol Gen Genet. 1977 Nov 14;156(2):121–131. doi: 10.1007/BF00283484. [DOI] [PubMed] [Google Scholar]
  19. Kitagawa Y., Akaboshi E., Shinagawa H., Horii T., Ogawa H., Kato T. Structural analysis of the umu operon required for inducible mutagenesis in Escherichia coli. Proc Natl Acad Sci U S A. 1985 Jul;82(13):4336–4340. doi: 10.1073/pnas.82.13.4336. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Koch W. H., Cebula T. A., Foster P. L., Eisenstadt E. UV mutagenesis in Salmonella typhimurium is umuDC dependent despite the presence of samAB. J Bacteriol. 1992 May;174(9):2809–2815. doi: 10.1128/jb.174.9.2809-2815.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Langer P. J., Perry K. L., Walker G. C. Complementation of a pKM101 derivative that decreases resistance to UV killing but increases susceptibility to mutagenesis. Mutat Res. 1985 Jun-Jul;150(1-2):147–158. doi: 10.1016/0027-5107(85)90112-5. [DOI] [PubMed] [Google Scholar]
  22. Little J. W. Autodigestion of lexA and phage lambda repressors. Proc Natl Acad Sci U S A. 1984 Mar;81(5):1375–1379. doi: 10.1073/pnas.81.5.1375. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Lodwick D., Owen D., Strike P. DNA sequence analysis of the imp UV protection and mutation operon of the plasmid TP110: identification of a third gene. Nucleic Acids Res. 1990 Sep 11;18(17):5045–5050. doi: 10.1093/nar/18.17.5045. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Lodwick D., Strike P. Distribution of sequences homologous to the impCAB operon of TP110 among bacterial plasmids of different incompatibility groups. Mol Gen Genet. 1991 Sep;229(1):27–30. doi: 10.1007/BF00264209. [DOI] [PubMed] [Google Scholar]
  25. MacPhee D. G. Spontaneous, ultraviolet and ionizing radiation mutagenesis in two auxotrophic strains of Salmonella typhimurium carrying an R plasmid. Mutat Res. 1977 Oct;45(1):1–6. doi: 10.1016/0027-5107(77)90036-7. [DOI] [PubMed] [Google Scholar]
  26. Murli S., Walker G. C. SOS mutagenesis. Curr Opin Genet Dev. 1993 Oct;3(5):719–725. doi: 10.1016/s0959-437x(05)80089-9. [DOI] [PubMed] [Google Scholar]
  27. Nohmi T., Battista J. R., Dodson L. A., Walker G. C. RecA-mediated cleavage activates UmuD for mutagenesis: mechanistic relationship between transcriptional derepression and posttranslational activation. Proc Natl Acad Sci U S A. 1988 Mar;85(6):1816–1820. doi: 10.1073/pnas.85.6.1816. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Nohmi T., Hakura A., Nakai Y., Watanabe M., Murayama S. Y., Sofuni T. Salmonella typhimurium has two homologous but different umuDC operons: cloning of a new umuDC-like operon (samAB) present in a 60-megadalton cryptic plasmid of S. typhimurium. J Bacteriol. 1991 Feb;173(3):1051–1063. doi: 10.1128/jb.173.3.1051-1063.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Nohmi T., Yamada M., Watanabe M., Murayama S. Y., Sofuni T. Roles of Salmonella typhimurium umuDC and samAB in UV mutagenesis and UV sensitivity. J Bacteriol. 1992 Nov;174(21):6948–6955. doi: 10.1128/jb.174.21.6948-6955.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Orrego C., Eisenstadt E. An inducible pathway is required for mutagenesis in Salmonella typhimurium LT2. J Bacteriol. 1987 Jun;169(6):2885–2888. doi: 10.1128/jb.169.6.2885-2888.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Overbye K. M., Margolin P. Role of the supX gene in ultraviolet light-induced mutagenesis in Salmonella typhimurium. J Bacteriol. 1981 Apr;146(1):170–178. doi: 10.1128/jb.146.1.170-178.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Pembroke J. T., Stevens E. The effect of plasmid R391 and other IncJ plasmids on the survival of Escherichia coli after UV irradiation. J Gen Microbiol. 1984 Jul;130(7):1839–1844. doi: 10.1099/00221287-130-7-1839. [DOI] [PubMed] [Google Scholar]
  33. Perry K. L., Elledge S. J., Mitchell B. B., Marsh L., Walker G. C. umuDC and mucAB operons whose products are required for UV light- and chemical-induced mutagenesis: UmuD, MucA, and LexA proteins share homology. Proc Natl Acad Sci U S A. 1985 Jul;82(13):4331–4335. doi: 10.1073/pnas.82.13.4331. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Perry K. L., Walker G. C. Identification of plasmid (pKM101)-coded proteins involved in mutagenesis and UV resistance. Nature. 1982 Nov 18;300(5889):278–281. doi: 10.1038/300278a0. [DOI] [PubMed] [Google Scholar]
  35. Pinney R. J. Distribution among incompatibility groups of plasmids that confer UV mutability and UV resistance. Mutat Res. 1980 Aug;72(1):155–159. doi: 10.1016/0027-5107(80)90232-8. [DOI] [PubMed] [Google Scholar]
  36. Rajagopalan M., Lu C., Woodgate R., O'Donnell M., Goodman M. F., Echols H. Activity of the purified mutagenesis proteins UmuC, UmuD', and RecA in replicative bypass of an abasic DNA lesion by DNA polymerase III. Proc Natl Acad Sci U S A. 1992 Nov 15;89(22):10777–10781. doi: 10.1073/pnas.89.22.10777. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. 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]
  38. Sedgwick S. G., Bridges B. A. Survival, mutation and capacity to repair single-strand DNA breaks after gamma irradiation in different Exr - strains of Escherichia coli. Mol Gen Genet. 1972;119(2):93–102. doi: 10.1007/BF00269129. [DOI] [PubMed] [Google Scholar]
  39. Sedgwick S. G., Goodwin P. A. Differences in mutagenic and recombinational DNA repair in enterobacteria. Proc Natl Acad Sci U S A. 1985 Jun;82(12):4172–4176. doi: 10.1073/pnas.82.12.4172. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Sedgwick S. G., Lodwick D., Doyle N., Crowne H., Strike P. Functional complementation between chromosomal and plasmid mutagenic DNA repair genes in bacteria. Mol Gen Genet. 1991 Oct;229(3):428–436. doi: 10.1007/BF00267466. [DOI] [PubMed] [Google Scholar]
  41. Shinagawa H., Iwasaki H., Kato T., Nakata A. RecA protein-dependent cleavage of UmuD protein and SOS mutagenesis. Proc Natl Acad Sci U S A. 1988 Mar;85(6):1806–1810. doi: 10.1073/pnas.85.6.1806. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Skavronskaya A. G., Stepanova N. F., Andreeva I. V. UV-mutable hybrids of Salmonella incorporating Escherichia coli region adjacent to tryptophan operon. Mol Gen Genet. 1982;185(2):315–318. doi: 10.1007/BF00330804. [DOI] [PubMed] [Google Scholar]
  43. Slilaty S. N., Little J. W. Lysine-156 and serine-119 are required for LexA repressor cleavage: a possible mechanism. Proc Natl Acad Sci U S A. 1987 Jun;84(12):3987–3991. doi: 10.1073/pnas.84.12.3987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Smith C. M., Eisenstadt E. Identification of a umuDC locus in Salmonella typhimurium LT2. J Bacteriol. 1989 Jul;171(7):3860–3865. doi: 10.1128/jb.171.7.3860-3865.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Smith C. M., Koch W. H., Franklin S. B., Foster P. L., Cebula T. A., Eisenstadt E. Sequence analysis and mapping of the Salmonella typhimurium LT2 umuDC operon. J Bacteriol. 1990 Sep;172(9):4964–4978. doi: 10.1128/jb.172.9.4964-4978.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Studier F. W., Rosenberg A. H., Dunn J. J., Dubendorff J. W. Use of T7 RNA polymerase to direct expression of cloned genes. Methods Enzymol. 1990;185:60–89. doi: 10.1016/0076-6879(90)85008-c. [DOI] [PubMed] [Google Scholar]
  47. Thomas S. M., Crowne H. M., Pidsley S. C., Sedgwick S. G. Structural characterization of the Salmonella typhimurium LT2 umu operon. J Bacteriol. 1990 Sep;172(9):4979–4987. doi: 10.1128/jb.172.9.4979-4987.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Thomas S. M., Sedgwick S. G. Cloning of Salmonella typhimurium DNA encoding mutagenic DNA repair. J Bacteriol. 1989 Nov;171(11):5776–5782. doi: 10.1128/jb.171.11.5776-5782.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Upton C., Pinney R. J. Expression of eight unrelated Muc+ plasmids in eleven DNA repair-deficient E. coli strains. Mutat Res. 1983 Oct;112(5):261–273. doi: 10.1016/0167-8817(83)90002-0. [DOI] [PubMed] [Google Scholar]
  50. Walker G. C. Mutagenesis and inducible responses to deoxyribonucleic acid damage in Escherichia coli. Microbiol Rev. 1984 Mar;48(1):60–93. doi: 10.1128/mr.48.1.60-93.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Woodgate R. Construction of a umuDC operon substitution mutation in Escherichia coli. Mutat Res. 1992 Mar;281(3):221–225. doi: 10.1016/0165-7992(92)90012-7. [DOI] [PubMed] [Google Scholar]
  52. Woodgate R., Sedgwick S. G. Mutagenesis induced by bacterial UmuDC proteins and their plasmid homologues. Mol Microbiol. 1992 Aug;6(16):2213–2218. doi: 10.1111/j.1365-2958.1992.tb01397.x. [DOI] [PubMed] [Google Scholar]
  53. Woodgate R., Singh M., Kulaeva O. I., Frank E. G., Levine A. S., Koch W. H. Isolation and characterization of novel plasmid-encoded umuC mutants. J Bacteriol. 1994 Aug;176(16):5011–5021. doi: 10.1128/jb.176.16.5011-5021.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES