Expression of the Gene for Resistance to Phaseolotoxin (argK) Depends on the Activity of Genes phtABC in Pseudomonas syringae pv. phaseolicola

Selene Aguilera,Susana De La Torre-Zavala,Jaime Bravo,José Luis Hernández-Flores,Ariel Alvarez-Morales,Jesús Murillo

doi:10.1371/journal.pone.0046815

Abstract

The bacterium Pseudomonas syringae pv. phaseolicola produces phaseolotoxin in a temperature dependent manner, being optimally produced between 18°C and 20°C, while no detectable amounts are present above 28°C. Phaseolotoxin is an effective inhibitor of ornithine carbamoyltransferase (OCTase) activity from plant, mammalian and bacterial sources and causes a phenotypic requirement for arginine. To protect the cell from its own toxin, P. syringae pv. phaseolicola synthesizes a phaseolotoxin-resistant OCTase (ROCT). The ROCT is the product of the argK gene and is synthesized only under conditions leading to phaseolotoxin synthesis. The argK gene is included in a chromosomal fragment named Pht cluster, which contains genes involved in the synthesis of phaseolotoxin. The aim of the present work was to investigate the possible involvement of other genes included in the Pht cluster in the regulation of gene argK. We conducted transcriptional analyses of argK in several mutants unable to produce phaseolotoxin, transcriptional fusions and electrophoretic mobility shift assays, which allowed us to determine that genes phtABC, located within the Pht cluster, participate in the transcriptional repression of gene argK at temperatures not permissive for phaseolotoxin biosynthesis. This repression is mediated by a protein present in both toxigenic and nontoxigenic strains of P. syringae and in E. coli, and requires the coordinated participation of phtA, phtB and phtC products in order to carry out an efficient argK repression.

Highlights

Production of phaseolotoxin, a non host specific toxin, has been described in Pseudomonas syringae pv. phaseolicola, which infects bean (Phaseolus vulgaris L), P. syringae pv. actinidiae, which infects kiwi (Actinidia chinensis), and in strain CFBP3388 of P. syringae pv. syringae, isolated from vetch (Vicia sativa) [1,2,3]
Sulfodiaminophosphinyl, and the L-ornithyl-alanyl-homoarginine tripeptide [1,11]. Targets of this toxin are the enzymes ornithine carbamoyltransferase (OCTase; EC 2.1.3.3) [12], which catalyzes the formation of citrulline from ornithine and carbamoylphosphate in the arginine biosynthetic pathway, and ornithine decarboxylase, which participates in the biosynthesis of polyamines [13]
In mutant YNorf1P, the argK gene showed an increased expression at 28uC, unlike what happens in strain NPS3121 at the same temperature (Figure 1B), indicating that a mutation on the phtA operon resulted in alleviation of the repression of argK at a nonpermissive temperature for phaseolotoxin synthesis

Summary

Introduction

Production of phaseolotoxin, a non host specific toxin, has been described in Pseudomonas syringae pv. phaseolicola, which infects bean (Phaseolus vulgaris L), P. syringae pv. actinidiae, which infects kiwi (Actinidia chinensis), and in strain CFBP3388 of P. syringae pv. syringae, isolated from vetch (Vicia sativa) [1,2,3]. Sulfodiaminophosphinyl, and the L-ornithyl-alanyl-homoarginine tripeptide [1,11] Targets of this toxin are the enzymes ornithine carbamoyltransferase (OCTase; EC 2.1.3.3) [12], which catalyzes the formation of citrulline from ornithine and carbamoylphosphate in the arginine biosynthetic pathway, and ornithine decarboxylase, which participates in the biosynthesis of polyamines [13]. Phaseolotoxin is an effective inhibitor of OCTase activity from plant, mammalian and bacterial sources and causes a phenotypic requirement for arginine. This property led to the development of a rapid bioassay that evaluates growth inhibition of a bacterial culture exposed to this toxin [14]

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Conclusion