The Pbo Cluster from Pseudomonas syringae pv. Phaseolicola NPS3121 Is Thermoregulated and Required for Phaseolotoxin Biosynthesis

Lizeth Guardado-Valdivia,Jesús Murillo,José Luis Hernández-Flores,Luis Xoca-Orozco,Selene Aguilera,Jorge Poveda,Alejandra Chacón-López

doi:10.3390/toxins13090628

Abstract

The bean (Phaseolus vulgaris) pathogen Pseudomonas syringae pv. phaseolicola NPS3121 synthesizes phaseolotoxin in a thermoregulated way, with optimum production at 18 °C. Gene PSPPH_4550 was previously shown to be thermoregulated and required for phaseolotoxin biosynthesis. Here, we established that PSPPH_4550 is part of a cluster of 16 genes, the Pbo cluster, included in a genomic island with a limited distribution in P. syringae and unrelated to the possession of the phaseolotoxin biosynthesis cluster. We identified typical non-ribosomal peptide synthetase, and polyketide synthetase domains in several of the pbo deduced products. RT-PCR and the analysis of polar mutants showed that the Pbo cluster is organized in four transcriptional units, including one monocistronic and three polycistronic. Operons pboA and pboO are both essential for phaseolotoxin biosynthesis, while pboK and pboJ only influence the amount of toxin produced. The three polycistronic units were transcribed at high levels at 18 °C but not at 28 °C, whereas gene pboJ was constitutively expressed. Together, our data suggest that the Pbo cluster synthesizes secondary metabolite(s), which could participate in the regulation of phaseolotoxin biosynthesis.

Highlights

The chromosomal fragment containing the gene PSPPH_4550 from P. syringae pv. phaseolicola NPS3121 is part of a large cluster, the Pbo cluster, composed of 16 genes organized in four transcriptional units, three polycistronic and one monocistronic
We identified a 34.8 kb genomic island in P. syringae pv. phaseolicola NPS3121 containing the newly described Pbo cluster
16 genes organized in four transcriptional units and is involved in the biosynthesis of the antimetabolite toxin phaseolotoxin

Summary

Introduction

The typical disease symptoms are small brown spots surrounded by a light-green or yellow halo [1] This halo is produced by phaseolotoxin, a non-specific toxin that inhibits the ornithine carbamoyltransferase (OCTase) activity from plants, mammalian, and bacterial sources, causing a phenotypic requirement for arginine and polyamines [2,3]. This property led to the development of a rapid bioassay for evaluating the growth inhibition of an Escherichia coli indicator strain caused by the toxin contained in the pseudomonas

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Conclusion