Fusarium Tri4 encodes a multifunctional oxygenase required for trichothecene biosynthesis

Fusarium graminearum and Fusarium sporotrichioides produce the trichothecene mycotoxins 15-acetyldeoxynivalenol and T-2 toxin, respectively. In both species, disruption of the P450 monooxygenase-encoding gene, Tri4, blocks production of the mycotoxins and leads to the accumulation of the trichothecene precursor trichodiene. To further characterize its function, the F. graminearum Tri4 (FgTri4) was heterologously expressed in the trichothecene-nonproducing species Fusarium verticillioides. Transgenic F. verticillioides carrying the FgTri4 converted exogenous trichodiene to the trichothecene biosynthetic intermediates isotrichodermin and trichothecene. Conversion of trichodiene to isotrichodermin requires seven biochemical steps. The fifth and sixth steps can occur nonenzymatically. Precursor feeding studies done in the current study indicate that wild-type F. verticillioides has the enzymatic activity necessary to carry out the seventh step, the C-3 acetylation of isotrichodermol to form isotrichodermin. Together, the results of this study indicate that the Tri4 protein catalyzes the remaining four steps and is therefore a multifunctional monooxygenase required for trichothecene biosynthesis.     

Global molecular surveillance reveals novel Fusarium head blight species and trichothecene toxin diversity

To expand our knowledge of Fusarium head blight (FHB) pathogen and trichothecene toxin diversity, a global collection of 2100 isolates was screened for novel genetic variation, resulting in the identification of 16 phylogenetically divergent FHB isolates. The affinities and taxonomic status of these novel isolates were evaluated via phylogenetic analyses of multilocus DNA sequence data (13 genes; 16.3 kb/strain) together with analyses of their morphology, pathogenicity to wheat, and trichothecene toxin potential. Based on the results of these analyses, we formally describe two novel species (Fusarium vorosii and Fusarium gerlachii) within the Fusarium graminearum species complex (Fg complex), and provide the first published report of Fg complex isolates with either a nivalenol or 3-acetyldeoxynivalenol chemotype within the U.S. In addition, we describe a highly divergent population of F. graminearum from the Gulf Coast of the U.S., and divergent isolates of F. acaciae-mearnsii from Australia and South Africa.     

Tri1 encodes the cytochrome P450 monooxygenase for C-8 hydroxylation during trichothecene biosynthesis in Fusarium sporotrichioides and resides upstream of another new Tri gene

Many Fusarium species produce one or more agriculturally important trichothecene mycotoxins, and the relative level of toxicity of these compounds is determined by the pattern of oxygenations and acetylations or esterifications on the core trichothecene structure. Previous studies with UV-induced Fusarium sporotrichioides NRRL 3299 trichothecene mutants defined the Tri1 gene and demonstrated that it was required for addition of the oxygen at the C-8 position during trichothecene biosynthesis. We have cloned and characterized the Tri1 gene from NRRL 3299 and found that it encodes a cytochrome P450 monooxygenase. The disruption of Tri1 blocks production of C-8-oxygenated trichothecenes and leads to the accumulation of 4,15-diacetoxyscirpenol, the same phenotype observed in the tri1 UV-induced mutants MB1716 and MB1370. The Tri1 disruptants and the tri1 UV-induced mutants do not complement one another when coinoculated, and the Tri1 gene sequence restores T-2 toxin production in both MB1716 and MB1370. The DNA sequence flanking Tri1 contains another new Tri gene. Thus, Tri1 encodes a C-8 hydroxylase and is located either in a new distal portion of the trichothecene gene cluster or in a second separate trichothecene gene cluster.     

Mechanisms of DNA-binding specificity and functional gene regulation by transcription factors

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