Molecular Genetics and Genomics

, Volume 271, Issue 1, pp 40–49

LexA-binding sequences in Gram-positive and cyanobacteria are closely related

  • G. Mazón
  • J. M. Lucena
  • S. Campoy
  • A. R. Fernández de Henestrosa
  • P. Candau
  • J. Barbé
Original Paper

Abstract

The lexA gene of the cyanobacterium Anabaena sp. strain PCC7120 has been cloned by PCR amplification with primers designed after TBLASTN analysis of its genome sequence using the Escherichia coli LexA sequence as a probe. After over-expression in E. coli and subsequent purification, footprinting experiments demonstrated that the Anabaena LexA protein binds to the sequence TAGTACTAATGTTCTA, which is found upstream of its own coding gene. Directed mutagenesis and sequence comparison of promoters of other Anabaena genes, as well as those of several cyanobacteria, allowed us to define the motif RGTACNNNDGTWCB as the LexA box in this bacterial phylum. Substitution of a single nucleotide in this motif present in the Anabena lexA promoter is sufficient to enable it to bind the Bacillus subtilis LexA protein. These data indicate that Cyanobacteria and Gram-positive bacteria are phylogenetically closely related.

Keywords

Lex A box SOS response Cyanobacteria Bacterial phylogeny 

References

  1. Bonacossa de Almeida C, Coste G, Sommer S, Bailone A (2002) Quantification of RecA protein in Deinococcus radiodurans reveals involvement of RecA, but not LexA, in its regulation. Mol Genet Genomics 268:28–41Google Scholar
  2. Campoy S, Mazón G, Fernández de Henestrosa AR, Llagostera M, Brant-Monteiro P, Barbé J (2002) A new regulatory DNA motif of the gamma subclass Proteobacteria: identification of the LexA protein binding site of the plant pathogen Xylella fastidiosa. Microbiology 148:3583–3597PubMedGoogle Scholar
  3. Campoy S, Fontes M, Padmanabhan S, Cortés P, Llagostera M, Barbé J (2003) LexA-independent DNA damage-mediated induction of gene expression in Myxococcus xanthus. Mol Microbiol 49:769–781CrossRefPubMedGoogle Scholar
  4. Chávez S, Reyes JC, Chauvat F, Florencio FJ, Candau P (1995) The NADP-glutamate dehydrogenase of the cyanobacterium Synechocystis PCC6803: cloning, transcriptional analysis and disruption of the gdhA gene. Plant Mol Biol 28:173–188PubMedGoogle Scholar
  5. Davis EO, Dullaghan EM, Rand L (2002) Definition of the mycobacterial SOS box and its use to identify LexA-regulated genes in Mycobacterium tuberculosi s. J Bacteriol 184:3287–3295CrossRefPubMedGoogle Scholar
  6. Erill I, Escribano M, Campoy S, Barbé J (2003) In silico analysis reveals substantial variability in the gene contents of the Gamma Proteobacteria LexA regulon. Bioinformatics 19:2225–2236Google Scholar
  7. Fernández de Henestrosa AR, Ogi T, Aoyagi S, Chafin D, Hayes JJ, Ohmori H, Woodgate R (2000) Identification of additional genes belonging to the LexA regulon in Escherichia coli. Mol Microbiol 35:1560–1572CrossRefPubMedGoogle Scholar
  8. Fernández de Henestrosa AR, Cuñé J, Erill I, Magnuson JK, Barbé J (2002) A green nonsulfur bacterium, Dehalococcoides ethenogenes , with the LexA binding sequence found in gram-positive organisms. J Bacteriol 184:6073 –6080CrossRefPubMedGoogle Scholar
  9. Fogh RH, Ottleben G, Rüterjans H, Schnarr M, Boelens R, Kaptein R (1994) Solution structure of the LexA repressor DNA binding domain determined by 1H NMR spectroscopy. EMBO J 13:3936–3944PubMedGoogle Scholar
  10. Frías JE, Flores E, Herrero A (1997) Nitrate assimilation gene cluster from the heterocyst-forming cyanobacterium Anabaena sp. strain PCC7120. J Bacteriol 179:477–486PubMedGoogle Scholar
  11. Geoghegan CM, Houghton JA (1987) Molecular cloning and isolation of a cyanobacterial gene which increases the UV and methyl methanesulphonate survival of recA strains of Escherichia coli K12. J Gen Microbiol 133:119–126PubMedGoogle Scholar
  12. Gupta RS, Griffiths E (2002) Critical issues in bacterial phylogeny. Theor Popul Biol 61:423–434CrossRefPubMedGoogle Scholar
  13. Jara M, Núñez C, Campoy S, Fernández de Henestrosa AR, Lovley DR, Barbé J (2003) Geobacter sulfurreducens has two autoregulated lexA genes whose products do not bind the recA promoter: differing responses of lexA and recA to DNA damage. J Bacteriol 185:2493–2502CrossRefPubMedGoogle Scholar
  14. Knegtel RMA, Fogh RH, Ottleben G, Rüterjans H, Dumoulin P, Schnarr M, Boelens R, Kaptein R (1995) A model for the LexA repressor DNA complex. Proteins 21:226–236PubMedGoogle Scholar
  15. Luo Y, Pfuetzner RA, Mosimann S, Paetzel M, Frey EA, Cherney M, Kim B, Little JW, Strynadka NC (2001) Crystal structure of LexA: a conformational switch for regulation of self-cleavage. Cell 106:585–594PubMedGoogle Scholar
  16. Miller JH (1992) A short course in bacterial genetics. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.Google Scholar
  17. Murphy RC, Bryant DA, Porter RD, Tandeau de Marsac N (1987) Molecular cloning and characterization of the recA gene from the cyanobacterium Synechoccus sp. strain PCC7002. J Bacteriol 169:2739–2747PubMedGoogle Scholar
  18. Narumi I, Satoh K, Kikuchi M, Funayama T, Yanagisawa T, Kobayashi Y, Watanabe H, Yamamoto K (2001) The LexA protein from Deinococcus radiodurans is not involved in RecA induction following gamma irradiation. J Bacteriol 183:6951–6956PubMedGoogle Scholar
  19. Owttrim GW, Coleman JR (1987) Molecular cloning of a recA -like gene from the cyanobacterium Anabaena variabilis. J Bacteriol 169:1824–1829PubMedGoogle Scholar
  20. Patzer SI, Hantke K (2001) Dual repression by Fe2+-Fur and Mn2+-MntR of the mntH gene, encoding an NRAMP-like Mn2+ transporter in Escherichia coli. J Bacteriol 183:4806–4813CrossRefPubMedGoogle Scholar
  21. Sambrook J, Russell DW (2001) Molecular cloning: a laboratory manual (3rd edn). Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.Google Scholar
  22. Sanger F, Nicklen S, Coulson A (1977) DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci USA 74:5463–5467PubMedGoogle Scholar
  23. Sassanfar M, Roberts JW (1990) Nature of the SOS-inducing signal in Escherichia coli. The involvement of DNA replication. J Mol Biol 212:79–96PubMedGoogle Scholar
  24. Tapias A, Barbé J (1998) Mutational analysis of the Rhizobium etli recA operator. J Bacteriol 180:6325–6331PubMedGoogle Scholar
  25. Tapias A, Barbé J (1999) Regulation of divergent transcription from the uvrA-ssb promoters in Sinorhizobium meliloti. Mol Gen Genet 262:121–130CrossRefPubMedGoogle Scholar
  26. Walker GC (1984) Mutagenesis and inducible responses to deoxyribonucleic acid damage in Escherichia coli. Microbiol Rev 48:60–93PubMedGoogle Scholar
  27. Winterling KW, Chafin D, Hayes JJ, Sun J, Levine AS, Yasbin RE, Woodgate R (1998) The Bacillus subtilis DinR binding site: redefinition of the consensus sequence. J Bacteriol 180:2201–2211PubMedGoogle Scholar

Copyright information

© Springer-Verlag 2004

Authors and Affiliations

  • G. Mazón
    • 1
  • J. M. Lucena
    • 2
  • S. Campoy
    • 3
  • A. R. Fernández de Henestrosa
    • 1
    • 4
  • P. Candau
    • 2
  • J. Barbé
    • 1
  1. 1.Departament de Genètica i MicrobiologiaUniversitat Autònoma de BarcelonaBarcelonaSpain
  2. 2.Instituto de Bioquímica Vegetal y FotosíntesisUniversidad de Sevilla-CSICSevillaSpain
  3. 3.Institut de Recerca i Tecnologia Agroalimentària (UAB-IRTA), Centre de Recerca en Sanitat Animal (CReSA)Universitat Autònoma de BarcelonaBarcelonaSpain
  4. 4.Centro de Biología Molecular Severo Ochoa (CSIC)MadridSpain

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