Skip to main content
SpringerLink
Log in

GacS-dependent regulation of enzymic and antifungal activities and synthesis of N-acylhomoserine lactones in rhizospheric strain Pseudomonas chlororaphis 449

  • Published:
Folia Microbiologica Aims and scope Submit manuscript

Abstract

Pseudomonas chlororaphis strain 449 isolated from the rhizosphere of maize suppresses numerous plant pathogens in vitro. The strain produces phenazine antibiotics and synthesizes at least three types of quorum sensing signaling molecules, N-acylhomoserine lactones. Here we have shown that the rhizospheric P. chlororaphis strains 449, well known strain 30–84 as well as two other P. chlororaphis strains exhibit polygalacturonase activity. Using mini-Tn5 transposon mutagenesis, four independent mutants of strain P. chlororaphis 449 with insertion of mini-Tn5 Km2 in gene gacS of two-component GacA-GacS system of global regulation were selected. All these mutant strains were deficient in production of extracellular proteinase(s), phenazines, N-acylhomoserine lactones synthesis, and did not inhibit the growth of G+ bacteria in comparison with the wild type strain. The P. chlororaphis 449-06 gacS − mutant studied in greater detail was deficient in polygalacturonase, pectin methylesterase activities, swarming motility and antifungal activity. It is the first time the involvement of GacA-GacS system in the regulation of enzymes of pectin metabolism, polygalacturonase and pectin methylesterase, was demonstrated in fluorescent pseudomonads.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

Abbreviations

AHL(s):

N-acylhomoserine lactone(s)

Ap:

ampicillin

Cbr :

carbenicillin-resistant

Clm:

chloramphenicol

EPR(s):

extracellular proteinase(s)

Gm:

gentamicin

Gmr :

gentamicin-resistant

Hgr :

mercury-resistant

HSL:

homoserine lactone(s)

IS:

insertion sequence

Km:

kanamycin

Kmr :

kanamycin-resistant

LA:

LB agarized

LB:

Luria broth

PCA:

phenazine-1-carboxylic acid

2-OH-PCA:

2-hydroxyphenazine-PCA

PCR:

polymerase chain reaction

PDA:

potato dextrose agar

PG:

polygalacturonase (EC 3.2.1.15)

PHZ(s):

phenazine(s)

2-OH-PHZ:

2-phenazinol

PME:

pectin methylesterase (EC 3.1.1.11)

PNPP:

4-nitrophenyl phosphate

QS:

quorum sensing

Tcr :

tetracycline-resistant

References

  • Ausubel F.M., Brendt R., Kingston R.E., Moore D.D., Seidman J.G., Smith J.A., Struhl K. (Eds): Current Protocols in Molecular Biology. John Wiley & Sons, New York 1994.

    Google Scholar 

  • Bradford M.: A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein binding. Anal.Biochem. 72, 248–254 (1976).

    Article  CAS  PubMed  Google Scholar 

  • Brencic A., Winans S.: Detection of and response to signals involved in host-microbe interactions by plant-associated bacteria. Microbiol.Mol.Biol.Rev. 69, 155–194 (2005).

    Article  CAS  PubMed  Google Scholar 

  • Brummell D.A., Harpster M.H.: Cell wall metabolism in fruit softening and quality and its manipulation in transgenic plants. Plant Mol.Biol. 47, 311–340 (2001).

    Article  CAS  PubMed  Google Scholar 

  • Chancey S.T., Wood D.W., Pierson L.S. 3rd: Two-component transcriptional regulation of N-acylhomoserine lactone production in Pseudomonas aureofaciens. Appl.Environ.Microbiol. 65, 2294–2299 (1999).

    CAS  PubMed  Google Scholar 

  • Daniels R., Vanderleyden J., Michiels J.: Quorum sensing and swarming migration in bacteria. FEMS Microbiol.Rev. 28, 261–289 (2004).

    Article  CAS  PubMed  Google Scholar 

  • Dassa e., Cahu M., Desjoyaux-Cherel B., Boquet P.L.: The acid phosphatase with optimum pH of 2.5 of Escherichia coli. J.Biol. Chem. 257, 6669–6676 (1982).

    CAS  PubMed  Google Scholar 

  • Delaney S.M., Mavrodi D.V., Robert F., Bonsall R.F., Thomashow L.S.: phzO, a gene for biosynthesis of 2-hydroxylated phenazine compounds in Pseudomonas aureofaciens 30–84. J.Bacteriol. 183, 318–327 (2001).

    Article  CAS  PubMed  Google Scholar 

  • Dingle J., Reid W.W., Solomonos G.L.: The enzymatic degradation of pectin and other polysaccharides. II. The application of the cup-plate assay to the estimation of enzyme. J.Sci.Food Agric. 4, 149–153 (1953).

    Article  CAS  Google Scholar 

  • Girard G., van Rij E.T., Lugtenberg B.J.J., Bloemberg G.V.: Regulatory roles of psrA and rpoS in phenazine-1-carboxamide synthesis by Pseudomonas chlororaphis PCL1391. Microbiology 152, 43–58 (2006).

    Google Scholar 

  • Givskov M., de Nys R., Manefield M., Gram L., Maximilien R., Eberl L., Molin S., Steinberg P.D., Kjelleberg S.: Eukaryotic interference with homoserine lactone-mediated prokaryotic signalling. J.Bacteriol. 178, 6618–6622 (1996).

    CAS  PubMed  Google Scholar 

  • Gugi B., Orange N., Hellio F., Burini J., Guillou C., Leriche F., Guespin-Mishel J.: Effect of growth temperature on several exported enzyme activities in the psychrotrophic bacterium Pseudomonas fluorescens. J.Bacteriol. 173, 3814–3820 (1991).

    CAS  PubMed  Google Scholar 

  • Haas D., Defago G.: Biological control of soil-borne pathogens by fluorescent pseudomonads. Nat.Rev.Microbiol. 3, 307–319 (2005).

    Article  CAS  PubMed  Google Scholar 

  • Han S.H., Lee S.L., Moon J.H., Park K.H., Yang K.Y., Cho B.H., Kim K.Y., Kim Y.W., Lee M.C., Anderson A.J., Kim Y.C.: GacS-dependent production of 2R,3R-butanediol by Pseudomonas chlororaphis 06 is a major determinant for eliciting systemic resistance against Erwinia carotovora but not against Pseudomonas syringae pv. tabaci in tobacco. Mol. Plant-Microbe Interact. 19, 924–930 (2006).

    Article  CAS  PubMed  Google Scholar 

  • Klein S., Veselova M., Mayatskaya A., Khmel I., Chet I., Chernin L.: Biocontrol activity of phenazine-producing rhizobacterium Pseudomonas chlororaphis 449, pp. 145–150 in R. Sikora, S. Gowen, R. Hauschild, S. Kiewnick (Eds): Multitrophic Integrations in Soil. IOBC wprs Bull., Vol. 27 (2004).

  • Lapouge K., Schubert M., Allain F.H.-T., Haas D.: Gac/Rsm signal transduction pathway of γ-proteobacteria: from RNA recognition to regulation of social behaviour. Mol.Microbiol. 67, 241–253 (2008).

    CAS  PubMed  Google Scholar 

  • Lazdunski A., Murgier M., Lazdunski C.: Evidence for an aminoendopeptidase localized near the cell surface of Escherichia coli. Regulation of synthesis by inorganic phosphate. Eur.J.Biochem. 60, 349–355 (1975).

    Article  CAS  PubMed  Google Scholar 

  • Lonon M.K., Woods D.E., Straus D.C.: Production of lipase by clinical isolates of Pseudomonas cepacia. J.Clin.Microbiol. 26, 979–984 (1988).

    CAS  PubMed  Google Scholar 

  • de Lorenzo V., Timmis K.N.: Analysis and construction of stable phenotypes in Gram-negative bacteria with Tn5- and Tn10-derived minitransposons. Meth.Enzymol. 235, 386–405 (1994).

    Article  CAS  PubMed  Google Scholar 

  • de Lorenzo V., Herrero M., Jakubzik U., Timmis K.N.: Mini-Tn5 transposon derivatives for insertion mutagenesis, promoter probing, and chromosomal insertion of cloned DNA in Gram-negative eubacteria. J.Bacteriol. 172, 6568–6572 (1990).

    PubMed  Google Scholar 

  • Maddula V.S.R.K., Pierson E.A., Pierson L.S. III: Altering the ratio of phenazines in Pseudomonas chlororaphis (aureofaciens) strain 30–84: effects on biofilm formation and pathogen inhibition. J.Bacteriol. 190, 2759–2766 (2008).

    Article  CAS  PubMed  Google Scholar 

  • Massa C., Degrassi G., Devescovi G., Venturi V., Lamba D.: Isolation, heterologues expression and characterization of an endopolygalacturonase produced by the phytopathogen Burkholderia cepacia. Protein Expr.Purif. 54, 300–308 (2007).

    Article  CAS  PubMed  Google Scholar 

  • McClean K.H., Winson M.K., Fish L., Taylor A., Chabra S.R., Camara M., Daykin M., Lamb J.H., Swift S., Bycroft B.W., Stewart G.S.A.B., Williams P.: Quorum sensing in Chromobacterium violaceum: exploitation of violacein production and inhibition for the detection of N-acylhomoserine lactones. Microbiology 143, 3703–3711 (1997).

    Google Scholar 

  • McGuire R., Rodriguez-Palenzuela P., Collmer A., Burr T.: Polygalacturonase production by Agrobacterium tumefaciens biovar 3. Appl.Environ.Microbiol. 57, 660–664 (1991).

    CAS  PubMed  Google Scholar 

  • Micheli F.: Pectin methylesterases: cell wall enzymes with important roles in plant physiology. Trends Plant Sci. 6, 414–419 (2001).

    Article  CAS  PubMed  Google Scholar 

  • Miller J.H.: A Short Course in Molecular Genetics. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York 1992.

    Google Scholar 

  • Poritsanos N., Selin C., Fernando W.G.D., Nakkeeran S., De Kievit T.R.: A gacS deficiency does not affect Pseudomonas chlororaphis PA23 fitness when growing on canola, in aged batch culture or as a biofilm. Can.J.Microbiol. 52, 1177–1188 (2006).

    Article  CAS  PubMed  Google Scholar 

  • Schell M.A.: Purification and characterization of an endoglucanase from Pseudomonas solanacearum. Appl.Environ.Microbiol. 53, 2237–2241 (1987).

    CAS  PubMed  Google Scholar 

  • Schell M., Roberts D., Denny T.: Analysis of the Pseudomonas solanocearum polygalacturonase encoded by pglA and its involvement in phytopathogenicity. J.Bacteriol. 170, 4501–4508 (1988).

    CAS  PubMed  Google Scholar 

  • Shaw P.D., Ping G., Daly S.L., Cha C., Cronan J.E. Jr., RINEHART K.L., FARRAND S.K.: Detecting and characterizing N-acyl-homoserine lactone signal molecules by thin-layer chromatography. Proc.Nat.Acad.Sci.USA 94, 6036–6041 (1997).

  • Thomashow L.S., Weller D.M.: Current concepts in the use of introduced bacteria for biological disease control: mechanisms and antifungal metabolites, pp. 187–235 in Plant-Microbe Interactions (G. Stacey, N. Keen, Eds), Vol. 1. Chapman and Hall, New York 1996.

    Google Scholar 

  • Torriani A.: Influence of inorganic phosphate in the formation of phosphatases by Escherichia coli. Biochim.Biophys.Acta 38, 460–469 (1960).

    Google Scholar 

  • Veselova M., Kholmeckaya M., Klein S., Voronina E., Lipasova V., Metlitskaya A., Mayatskaya A., Lobanok E., Khmel I., Chernin L.: Production of N-acylhomoserine lactone signal molecules by Gram-negative soil-borne and plantassociated bacteria. Folia Microbiol. 48, 794–798 (2003).

    Article  CAS  Google Scholar 

  • Veselova M.A., Klein Sh., Bass I.A., Lipasova V.A., Metlitskaya A.Z., Ovadis M.I., Chernin L.S., Khmel I.A.: Quorum sensing systems of regulation, synthesis of phenazine antibiotics and antifungal activity in rhizospheric bacterium Pseudomonas chlororaphis 449. Russ.J.Genet. 44, 1617–1626 (2008).

    Article  CAS  Google Scholar 

  • Wood D.W., Gong F., Daykin M.M., Williams P., Pierson L.S. 3rd.: N-Acylhomoserine lactone-mediated regulation of phenazine gene expression by Pseudomonas aureofaciens 30–84 in the wheat rhizosphere. J.Bacteriol. 179, 7663–7670 (1997).

    CAS  PubMed  Google Scholar 

  • Zhang Z., Pierson L.S. III: A second quorum-sensing system regulates cell surface properties but not phenazine antibiotic production in Pseudomonas aureofaciens. Appl.Environ.Microbiol. 67, 4305–4315 (2001).

    Article  CAS  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Institute of Molecular Genetics, Russian Academy of Sciences, Moscow, Russia

    M. Veselova, V. Lipasova & I. A. Khmel

  2. A.N. Bach Institute of Biochemistry, Russian Academy of Sciences, Moscow, Russia

    M. A. Protsenko & N. Buza

Authors
  1. M. Veselova
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. V. Lipasova
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. M. A. Protsenko
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. N. Buza
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. I. A. Khmel
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to I. A. Khmel.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Veselova, M., Lipasova, V., Protsenko, M.A. et al. GacS-dependent regulation of enzymic and antifungal activities and synthesis of N-acylhomoserine lactones in rhizospheric strain Pseudomonas chlororaphis 449. Folia Microbiol 54, 401–408 (2009). https://doi.org/10.1007/s12223-009-0056-z

Download citation

  • Received:

  • Revised:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12223-009-0056-z

Keywords

Search

Navigation