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Title: ACTIVATION OF THE PHZ OPERON OF PSEUDOMONAS FLUORESCENS 2-79 REQUIRES THE LUXR HOMOLOG PHZR, N-(3-OH-HEXANOYL)-L-HOMOSERINE LACTONE PRODUCED BY PHZI, AND A CIS-ACTING PHZ-BOX.

Author
item KHAN, SHARIK - UNIV. OF ILL-URBANA
item MAVRODI, DMITRI - WASHINGTON STATE UNIV.
item JOG, GEETANJALI - UNIV. OF BUFFALO
item SUGA, HIROAKI - UNIV. OF BUFFALO
item Thomashow, Linda
item FARRAND, STEPHEN - UNIV. OF ILL-URBANA

Submitted to: Journal of Bacteriology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/20/2005
Publication Date: 9/20/2005
Citation: Khan, S.R., Mavrodi, D.V., Jog, G.J., Suga, H., Thomashow, L.S., Farrand, S.K. 2005. Activation of the phz operon of pseudomonas fluorescens 2-79 requires the luxr homolog phzr, n-(3-oh-hexanoyl)-l-homoserine lactone produced by phzi, and a cis-acting phz-box.. Journal of Bacteriology. (in press).

Interpretive Summary: We previously reported that phenazine-producing biocontrol strain P. fluorescens 2-79 synthesizes at least six acyl-homoserine lactones including the 3-hydroxy forms, N-(3-hydroxy-hexanoyl)-L-homoserine lactone (3OH-C6-HSL), N-(3-hydroxy-octanoyl)-L-homoserine lactone (3OH-C8-HSL), and N-(3-hydroxy-decanoyl)-Lhomoserine lactone (3OH-C10-HSL), the alkanoyl-forms hexanoyl-homoserine lactone (C6-HSL) and octanoyl-homoserine lactone (C8-HSL), and an active signal compound of unknown structure. However, P. aureofaciens 30-84 and P. chlororaphis PCL1391, the two other phenazine producers, both have been reported to produce and utilize as their signal the alkanoylsignal hexanoyl-homoserine lactone. Moreover, the two strains have not been reported to produce 3-hydroxy acyl-HSLs. Given the very close amino acid sequence relatedness of the PhzI and PhzR proteins from these three strains it became important to determine what acyl-HSL signals are produced by PhzI of P. fluorescens 2-79 and which of these signals activates the PhzR transcription factor of this strain. In this report we show that P. fluorescens 2-79 does indeed produce 3-hydroxy and alkanoyl-HSLs and that the PhzI synthase is responsible for the synthesis of all of the detectable acyl-HSL produced by this strain,. We also show that among the acyl-HSLs produced by strain 2-79, 3-OH-C6 accumulates to the highest concentration and that PhzR from strain 2-79 responds with greatest sensitivity to this signal. In addition, we show that activation of the phz operon by PhzR is dependent upon an intact inverted repeat, the phz box, in the phzA promoter region and that PhzR autoregulates its own transcription, also in a phz box-dependent manner.

Technical Abstract: We previously reported that phenazine-producing biocontrol strain Pseudomonas fluorescens 2-79 synthesizes at least six acyl-homoserine lactones including the 3-hydroxy forms, N-(3-hydroxy-hexanoyl)-L-homoserine lactone (3OH-C6-HSL), N-(3-hydroxy-octanoyl)-L-homoserine lactone (3OH-C8-HSL), and N-(3-hydroxy-decanoyl)-Lhomoserine lactone (3OH-C10-HSL), the alkanoyl-forms hexanoyl-homoserine lactone (C6-HSL) and octanoyl-homoserine lactone (C8-HSL), and an active signal compound of unknown structure. However, P. aureofaciens 30-84 and P. chlororaphis PCL1391, two other phenazine producers, both have been reported to produce and utilize as their signal the alkanoylsignal hexanoyl-homoserine lactone. Moreover, the latter two strains have not been reported to produce 3-hydroxy acyl-HSLs. Given the very close amino acid sequence relatedness of the PhzI and PhzR proteins from these three strains, it became important to determine what acyl-HSL signals are produced by PhzI of P. fluorescens 2-79 and which of these signals activates the PhzR transcription factor of this strain. In this report we show that P. fluorescens 2-79 does indeed produce 3-hydroxy and alkanoyl-HSLs and that the PhzI synthase is responsible for the synthesis of all of the detectable acyl-HSL produced by this strain. We also show that among the acyl-HSLs produced by strain 2-79, 3-OH-C6 accumulates to the highest concentration and that PhzR from strain 2-79 responds with greatest sensitivity to this signal. In addition, we show that activation of the phz operon by PhzR is dependent upon an intact inverted repeat, the phz box, in the phzA promoter region and that PhzR autoregulates its own transcription, also in a phz box-dependent manner.