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Volume 156, Issue 10

Functional roles for the GerE-family carboxyl-terminal domains of nitrate response regulators NarL and NarP of K-12 Free

Abstract

NarL and NarP are paralogous response regulators that control anaerobic gene expression in response to the favoured electron acceptors nitrate and nitrite. Their DNA-binding carboxyl termini are in the widespread GerE–LuxR–FixJ subfamily of tetrahelical helix–turn–helix domains. Previous biochemical and crystallographic studies with NarL suggest that dimerization and DNA binding by the carboxyl-terminal domain (CTD) is inhibited by the unphosphorylated amino-terminal receiver domain. We report here that NarL-CTD and NarP-CTD, liberated from their receiver domains, activated transcription from the class II and operon control regions, but failed to activate from the class I and operon control regions. Alanine substitutions were made to examine requirements for residues in the NarL DNA recognition helix. Substitutions for Val-189 and Arg-192 blocked DNA binding as assayed both and , whereas substitution for Arg-188 had a strong effect only . Similar results were obtained with the corresponding residues in NarP. Finally, Ala substitutions identified residues within the NarL CTD as important for transcription activation. Overall, results are congruent with those obtained for other GerE-family members, including GerE, TraR, LuxR and FixJ.

  • Received:
  • Accepted:
  • Revised:
  • Published Online:
Keyword(s): CTD, carboxyl-terminal domain , EMSA, electrophoretic mobility shift assay , HTH, helix–turn–helix and PC, positive control
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2010-10-01
2024-04-19
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References

  1. Aravind L., Anantharaman V., Balaji S., Babu M. M., Iyer L. M. 2005; The many faces of the helix-turn-helix domain: transcription regulation and beyond. FEMS Microbiol Rev 29:231–262
     [Google Scholar]
  2. Baikalov I., Schröder I., Kaczor-Grzeskowiak M., Grzeskowiak K., Gunsalus R. P., Dickerson R. E. 1996; Structure of the Escherichia coli response regulator NarL. Biochemistry 35:11053–11061
     [Google Scholar]
  3. Barnard A., Wolfe A., Busby S. 2004; Regulation at complex bacterial promoters: how bacteria use different promoter organizations to produce different regulatory outcomes. Curr Opin Microbiol 7:102–108
     [Google Scholar]
  4. Bartolomé B., Jubete Y., Martinez E., de la Cruz F. 1991; Construction and properties of a family of pACYC184-derived cloning vectors compatible with pBR322 and its derivatives. Gene 102:75–78
     [Google Scholar]
  5. Browning D., Lee D., Green J., Busby S. 2002; Secrets of bacterial transcription initiation taught by the Escherichia coli FNR protein. In Signals, Switches, Regulons, and Cascades: Control of Bacterial Gene Expression, Society for General Microbiology Symposium series vol 61 pp 127–142 Edited by Hodgson D. A., Thomas C. M. Reading, UK: Society for General Microbiology;
     [Google Scholar]
  6. Chang A. C. Y., Cohen S. N. 1978; Construction and characterization of amplifiable multicopy DNA cloning vehicles derived from the P15A cryptic miniplasmid. J Bacteriol 134:1141–1156
     [Google Scholar]
  7. Choi S. H., Greenberg E. P. 1991; The C-terminal region of the Vibrio fischeri LuxR protein contains an inducer-independent lux gene activating domain. Proc Natl Acad Sci U S A 88:11115–11119
     [Google Scholar]
  8. Costa E. D., Cho H., Winans S. C. 2009; Identification of amino acid residues of the pheromone-binding domain of the transcription factor TraR that are required for positive control. Mol Microbiol 73:341–351
     [Google Scholar]
  9. Crater D. L., Moran C. P. Jr 2002; Two regions of GerE required for promoter activation in Bacillus subtilis. J Bacteriol 184:241–249
     [Google Scholar]
  10. Darwin A. J., Tyson K. L., Busby S. J., Stewart V. 1997; Differential regulation by the homologous response regulators NarL and NarP of Escherichia coli K-12 depends on DNA binding site arrangement. Mol Microbiol 25:583–595
     [Google Scholar]
  11. Datsenko K. A., Wanner B. L. 2000; One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci U S A 97:6640–6645
     [Google Scholar]
  12. Ducros V. M., Lewis R. J., Verma C. S., Dodson E. J., Leonard G., Turkenburg J. P., Murshudov G. N., Wilkinson A. J., Brannigan J. A. 2001; Crystal structure of GerE, the ultimate transcriptional regulator of spore formation in Bacillus subtilis. J Mol Biol 306:759–771
     [Google Scholar]
  13. Egan S. M., Stewart V. 1990; Nitrate regulation of anaerobic respiratory gene expression in narX deletion mutants of Escherichia coli K-12. J Bacteriol 172:5020–5029
     [Google Scholar]
  14. Egland K. A., Greenberg E. P. 2001; Quorum sensing in Vibrio fischeri: analysis of the LuxR DNA binding region by alanine-scanning mutagenesis. J Bacteriol 183:382–386
     [Google Scholar]
  15. Eldridge A. M., Kang H. S., Johnson E., Gunsalus R., Dahlquist F. W. 2002; Effect of phosphorylation on the interdomain interaction of the response regulator, NarL. Biochemistry 41:15173–15180
     [Google Scholar]
  16. Galperin M. Y. 2006; Structural classification of bacterial response regulators: diversity of output domains and domain combinations. J Bacteriol 188:4169–4182
     [Google Scholar]
  17. Haldimann A., Wanner B. L. 2001; Conditional-replication, integration, excision, and retrieval plasmid–host systems for gene structure–function studies of bacteria. J Bacteriol 183:6384–6393
     [Google Scholar]
  18. Henikoff S., Wallace J. C., Brown J. P. 1990; Finding protein similarities with nucleotide sequence databases. Methods Enzymol 183:111–132
     [Google Scholar]
  19. Kahn D., Ditta G. 1991; Modular structure of FixJ: homology of the transcriptional activator domain with the −35 binding domain of sigma factors. Mol Microbiol 5:987–997
     [Google Scholar]
  20. Kiley P. J., Beinert H. 1998; Oxygen sensing by the global regulator, FNR: the role of the iron–sulfur cluster. FEMS Microbiol Rev 22:341–352
     [Google Scholar]
  21. Kurashima-Ito K., Kasai Y., Hosono K., Tamura K., Oue S., Isogai M., Ito Y., Nakamura H., Shiro Y. 2005; Solution structure of the C-terminal transcriptional activator domain of FixJ from Sinorhizobium meliloti and its recognition of the fixK promoter. Biochemistry 44:14835–14844
     [Google Scholar]
  22. Lamberg K. E., Kiley P. J. 2000; FNR-dependent activation of the class II dmsA and narG promoters of Escherichia coli requires FNR-activating regions 1 and 3. Mol Microbiol 38:817–827
     [Google Scholar]
  23. Li J., Stewart V. 1992; Localization of upstream sequence elements required for nitrate and anaerobic induction of fdn (formate dehydrogenase-N) operon expression in Escherichia coli K-12. J Bacteriol 174:4935–4942
     [Google Scholar]
  24. Lin A. V. 2009; Factors influencing transcription activation by response regulators NarP and NarL of Escherichia coli. PhD dissertation University of California; Davis, USA:
  25. Lin H.-Y., Bledsoe P. J., Stewart V. 2007; Activation of yeaR–yoaG operon transcription by the nitrate-responsive regulator NarL is independent of oxygen-responsive regulator Fnr in Escherichia coli K-12. J Bacteriol 189:7539–7548
     [Google Scholar]
  26. Maloy S. R., Stewart V. J., Taylor R. K. 1996 Genetic Analysis of Pathogenic Bacteria: a Laboratory Manual Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
     [Google Scholar]
  27. Marchler-Bauer A., Anderson J. B., Chitsaz F., Derbyshire M. K., DeWeese-Scott C., Fong J. H., Geer L. Y., Geer R. C., Gonzales N. R. other authors 2009; CDD: specific functional annotation with the Conserved Domain Database. Nucleic Acids Res 37:D205–D210
     [Google Scholar]
  28. Maris A. E., Sawaya M. R., Kaczor-Grzeskowiak M., Jarvis M. R., Bearson S. M., Kopka M. L., Schröder I., Gunsalus R. P., Dickerson R. E. 2002; Dimerization allows DNA target site recognition by the NarL response regulator. Nat Struct Biol 9:771–778
     [Google Scholar]
  29. Maris A. E., Kaczor-Grzeskowiak M., Ma Z., Kopka M. L., Gunsalus R. P., Dickerson R. E. 2005; Primary and secondary modes of DNA recognition by the NarL two-component response regulator. Biochemistry 44:14538–14552
     [Google Scholar]
  30. Miller J. H. 1972 Experiments in Molecular Genetics Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
     [Google Scholar]
  31. Morrison T. B., Parkinson J. S. 1994; Liberation of an interaction domain from the phosphotransfer region of CheA, a signaling kinase of Escherichia coli. Proc Natl Acad Sci U S A 91:5485–5489
     [Google Scholar]
  32. Nasser W., Reverchon S. 2007; New insights into the regulatory mechanisms of the LuxR family of quorum sensing regulators. Anal Bioanal Chem 387:381–390
     [Google Scholar]
  33. Noriega C. E., Lin H.-Y., Chen L.-L., Williams S. B., Stewart V. 2010; Asymmetric cross-regulation between the nitrate-responsive NarX–NarL and NarQ–NarP two-component regulatory systems from Escherichia coli K-12. Mol Microbiol 75:394–412
     [Google Scholar]
  34. Pappas K. M., Weingart C. L., Winans S. C. 2004; Chemical communication in proteobacteria: biochemical and structural studies of signal synthases and receptors required for intercellular signalling. Mol Microbiol 53:755–769
     [Google Scholar]
  35. Qin Y., Keenan C., Farrand S. K. 2009; N- and C-terminal regions of the quorum-sensing activator TraR cooperate in interactions with the alpha and sigma-70 components of RNA polymerase. Mol Microbiol 74:330–346
     [Google Scholar]
  36. Rabin R. S., Stewart V. 1992; Either of two functionally redundant sensor proteins, NarX and NarQ, is sufficient for nitrate regulation in Escherichia coli K-12. Proc Natl Acad Sci U S A 89:8419–8423
     [Google Scholar]
  37. Schlax P. J., Capp M. W., Record M. T. Jr 1995; Inhibition of transcription initiation by lac repressor. J Mol Biol 245:331–350
     [Google Scholar]
  38. Squire D. J., Xu M., Cole J. A., Busby S. J., Browning D. F. 2009; Competition between NarL-dependent activation and Fis-dependent repression controls expression from the Escherichia coli yeaR and ogt promoters. Biochem J 420:249–257
     [Google Scholar]
  39. Stewart V., Bledsoe P. J. 2003; Synthetic lac operator substitutions to study the nitrate- and nitrite-responsive NarX–NarL and NarQ–NarP two-component regulatory systems of Escherichia coli K-12. J Bacteriol 185:2104–2111
     [Google Scholar]
  40. Stewart V., Bledsoe P. J. 2005; Fnr-, NarP- and NarL-dependent regulation of transcription initiation from the Haemophilus influenzae Rd napF (periplasmic nitrate reductase) promoter in Escherichia coli K-12. J Bacteriol 187:6928–6935
     [Google Scholar]
  41. Stewart V., Bledsoe P. J. 2008; Substitutions at auxiliary operator O3 enhance repression by nitrate-responsive regulator NarL at synthetic lac control regions in Escherichia coli K-12. J Bacteriol 190:428–433
     [Google Scholar]
  42. Stewart V., Parales J. 1988; Identification and expression of genes narL and narX of the nar (nitrate reductase) locus in Escherichia coli K-12. J Bacteriol 170:1589–1597
     [Google Scholar]
  43. Stewart V., Rabin R. S. 1995; Dual sensors and dual response regulators interact to control nitrate- and nitrite-responsive gene expression in Escherichia coli. In Two-Component Signal Transduction pp 233–252 Edited by Hoch J. A., Silhavy T. J. Washington, DC: American Society for Microbiology;
     [Google Scholar]
  44. Stewart V., Bledsoe P. J., Williams S. B. 2003; Dual overlapping promoters control napF (periplasmic nitrate reductase) operon expression in Escherichia coli K-12. J Bacteriol 185:5862–5870
     [Google Scholar]
  45. Ton-Hoang B., Salhi M., Schumacher J., Da Re S., Kahn D. 2001; Promoter-specific involvement of the FixJ receiver domain in transcriptional activation. J Mol Biol 312:583–589
     [Google Scholar]
  46. Vannini A., Volpari C., Gargioli C., Muraglia E., Cortese R., De Francesco R., Neddermann P., Marco S. D. 2002; The crystal structure of the quorum sensing protein TraR bound to its autoinducer and target DNA. EMBO J 21:4393–4401
     [Google Scholar]
  47. White C. E., Winans S. C. 2005; Identification of amino acid residues of the Agrobacterium tumefaciens quorum-sensing regulator TraR that are critical for positive control of transcription. Mol Microbiol 55:1473–1486
     [Google Scholar]
  48. White C. E., Winans S. C. 2007; The quorum-sensing transcription factor TraR decodes its DNA binding site by direct contacts with DNA bases and by detection of DNA flexibility. Mol Microbiol 64:245–256
     [Google Scholar]
  49. Xiao G., Cole D. L., Gunsalus R. P., Sigman D. S., Chen C. H. 2002; Site-specific DNA cleavage of synthetic NarL sites by an engineered Escherichia coli NarL protein-1,10-phenanthroline cleaving agent. Protein Sci 11:2427–2436
     [Google Scholar]
  50. Zhang X., Zhou Y., Ebright Y. W., Ebright R. H. 1992; Catabolite gene activator protein (CAP) is not an “acidic activating region” transcription activator protein. J Biol Chem 267:8136–8139
     [Google Scholar]
  51. Zhang R. G., Pappas T., Brace J. L., Miller P. C., Oulmassov T., Molyneaux J. M., Anderson J. C., Bashkin J. K., Winans S. C., Joachimiak A. 2002; Structure of a bacterial quorum-sensing transcription factor complexed with pheromone and DNA. Nature 417:971–974
     [Google Scholar]
  52. Zheng L., Halberg R., Roels S., Ichikawa H., Kroos L., Losick R. 1992; Sporulation regulatory protein GerE from Bacillus subtilis binds to and can activate or repress transcription from promoters for mother-cell-specific genes. J Mol Biol 226:1037–1050
     [Google Scholar]
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Functional roles for the GerE-family carboxyl-terminal domains of nitrate response regulators NarL and NarP of Escherichia coli K-12
Microbiology 156, 2933 (2010); https://doi.org/10.1099/mic.0.040469-0
/content/journal/micro/10.1099/mic.0.040469-0
/content/journal/micro/10.1099/mic.0.040469-0
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