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

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.

Una región interpromotora incrementa la transcripción del gen argK, que codifica para la ornitina carbamoiltransferasa resistente a la faseolotoxina en Pseudomonas syringae pv. phaseolicola

“…In both cases, we observed expression of argK only at 18°C, unlike the expression detected in CYL233(pSAK) at both temperatures. In all cases, argK expression is achieved without the need of any element coded by the Pht cluster, in agreement with previous reports (Aguilera et al, 2012).…”

“…Our work group has used the strain CYL233 to perform heterologous expression assays of genes from the Pht cluster in a genetic background similar to strain NPS3121. In those studies we have observed that in strain CYL233(pSAK), the introduced argK gene is expressed both 18°C and 28°C contrary to strain NPS3121wt in which argK is expressed only at 18°C (Aguilera et al, 2012;Aguilera et al, 2007).…”

“…To this, we obtained the strains NPS3121(pSAK) and NPS3121(pKD2), which contain both a chromosomal and the plasmids copies of argK and its regulatory sequences. The plasmid pKD2 occurs in multiple copies in a similar way to the plasmid pSAK (Aguilera et al, 2012).…”

“…Recently, it was demonstrated that the coordinated participation of phtA, phtB and phtC products, coded into the Pht cluster, are necessary to carry out an efficient argK repression at temperatures not permissive for phaseolotoxin biosynthesis. Also, argK transcriptional control is mediated by a protein present in both, toxigenic and nontoxigenic strains of P. syringae and in Escherichia coli (Aguilera et al, 2012). The promoters for argK and phtABC genes have been previously determined and shown to be a Pribnow-type (σ 70 ) promoters with appropriate -10 and -35 regions (Aguilera et al, 2007;Hatziloukas & Panopoulos, 1992).…”

Metagenomic screening of microbiomes identifies pathogen-enriched environments

“…syringae pv. phaseolicola [78]. ArgK plays an essential role in the survival and pathogenicity of Ps.…”