Functional roles of FgLaeA in controlling secondary metabolism, sexual development, and virulence in Fusarium graminearum.

Hee-Kyoung Kim,Seunghoon Lee,Robert H Proctor,Robert A E Butchko,Sung-Hwan Yun,Susan P Mccormick,Seong-Mi Jo,Jae-Hyuk Yu

doi:10.1371/journal.pone.0068441

Abstract

Fusarium graminearum, the causal agent of Fusarium head blight in cereal crops, produces mycotoxins such as trichothecenes and zearalenone in infected plants. Here, we focused on the function of FgLaeA in F. graminearum, a homolog of Aspergillus nidulans LaeA encoding the global regulator for both secondary metabolism and sexual development. Prior to gene analysis, we constructed a novel luciferase reporter system consisting of a transgenic F. graminearum strain expressing a firefly luciferase gene under control of the promoter for either TRI6 or ZEB2 controlling the biosynthesis of these mycotoxins. Targeted deletion of FgLaeA led to a dramatic reduction of luminescence in reporter strains, indicating that FgLaeA controls the expression of these transcription factors in F. graminearum; reduced toxin accumulation was further confirmed by GC-MS analysis. Overexpression of FgLaeA caused the increased production of trichothecenes and additional metabolites. RNA seq-analysis revealed that gene member(s) belonging to ∼70% of total tentative gene clusters, which were previously proposed, were differentially expressed in the ΔFgLaeA strain. In addition, ΔFgLaeA strains exhibited an earlier induction of sexual fruiting body (perithecia) formation and drastically reduced disease symptoms in wheat, indicating that FgLaeA seems to negatively control perithecial induction, but positively control virulence toward the host plant. FgLaeA was constitutively expressed under both mycotoxin production and sexual development conditions. Overexpression of a GFP-FgLaeA fusion construct in the ΔFgLaeA strain restored all phenotypic changes to wild-type levels and led to constitutive expression of GFP in both nuclei and cytoplasm at different developmental stages. A split luciferase assay demonstrated that FgLaeA was able to interact with FgVeA, a homolog of A. nidulans veA. Taken together, these results demonstrate that FgLaeA, a member of putative FgVeA complex, controls secondary metabolism, sexual development, and virulence in F. graminearum, although the specific regulation pattern differs from that of LaeA in A. nidulans.

Highlights

Fusarium graminearum, a homothallic ascomycetous fungus, is an important pathogen of major cereal plants that causes diseases such as head blight, stalk rot, and ear rot [1,2,3]
AAQ95166.1) and F. fujikuroi FfLae1 (FN548141), identified two open reading frames (ORFs) as possible LaeA homologs based on sequence homology, both carrying a conserved methyltransferase domain, in F. graminearum
The first ORF, encodes for 317 amino acids, interrupted by 6 introns, which was confirmed by reverse transcription-polymerase chain reaction (RT-PCR), but different from the previous annotation in the F. graminearum database

Summary

Introduction

Fusarium graminearum (teleomorph: Gibberella zeae), a homothallic ascomycetous fungus, is an important pathogen of major cereal plants that causes diseases such as head blight (scab), stalk rot, and ear rot [1,2,3]. The ability of F. graminearum to produce sexual progeny (ascospores) on overwintering cereal debris is essential for completion of the recurrent cycle of the plant diseases [10]. In addition to direct yield losses, F. graminearum produces mycotoxins such as trichothecenes and zearalenone in host plants, threatening human and animal health [2]. Trichothecenes appear to contribute to the virulence of F. graminearum on host plants [12]. Zearalenone causes estrogenic disorders in laboratory rats, mice, and farm-raised swine [2]

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Results

Conclusion