Peptide synthetase gene in Trichoderma virens.

S E Wilhite,D C Straney,R D Lumsden

doi:10.1128/aem.67.11.5055-5062.2001

Abstract

Trichoderma virens (synonym, Gliocladium virens), a deuteromycete fungus, suppresses soilborne plant diseases caused by a number of fungi and is used as a biocontrol agent. Several traits that may contribute to the antagonistic interactions of T. virens with disease-causing fungi involve the production of peptide metabolites (e.g., the antibiotic gliotoxin and siderophores used for iron acquisition). We cloned a 5,056-bp partial cDNA encoding a putative peptide synthetase (Psy1) from T. virens using conserved motifs found within the adenylate domain of peptide synthetases. Sequence similarities with conserved motifs of the adenylation domain, acyl transfer, and two condensation domains support identification of the Psy1 gene as a gene that encodes a peptide synthetase. Disruption of the native Psy1 gene through gene replacement was used to identify the function of this gene. Psy1 disruptants produced normal amounts of gliotoxin but grew poorly under low-iron conditions, suggesting that Psy1 plays a role in siderophore production. Psy1 disruptants cannot produce the major T. virens siderophore dimerum acid, a dipetide of acylated N(delta)-hydroxyornithine. Biocontrol activity against damping-off diseases caused by Pythium ultimum and Rhizoctonia solani was not reduced by the Psy1 disruption, suggesting that iron competition through dimerum acid production does not contribute significantly to disease suppression activity under the conditions used.

Highlights

Trichoderma virens is an effective biological control agent for plant diseases caused by soilborne fungi [26]
We used degenerate oligonucleotides corresponding to portions of conserved cores C and F of peptide synthetases to prime a PCR performed with T. virens genomic DNA
Sequence similarity was observed in core consensus regions (Fig. 1) found in the adenylate domains of bacterial peptide synthetases, including the C motif believed to be characteristic of peptide synthetase adenylate domains [36]

Summary

Introduction

Trichoderma virens (synonym, Gliocladium virens [30]) is an effective biological control agent for plant diseases caused by soilborne fungi [26]. Production of additional antibiotics, competition for resources in the soil or rhizosphere, and hyperparasitism of target fungi are traits that may account for other antagonistic interactions [7]. Cloning of genes associated with antagonistic traits could permit manipulation of the genes in T. virens and could increase the biocontrol activity of this organism. Small peptides are associated with a number of the traits that might contribute to T. virens biocontrol activity. The enzymes are organized into repetitive synthase units, each of which has the functions required to complete a different single amino acid elongation step in the synthesis of the peptide product [20]. The number and linear order of the repetitive synthase units correspond to the amino acid sequence of the peptide product. In this study our objectives were to clone a peptide synthetase from T. virens and to use specific gene replacement to identify the function of the cloned gene and its role, if any, in biocontrol activity

Objectives

Methods

Results

Conclusion