Hydroxylations of trichothecene rings in the biosynthesis of Fusarium trichothecenes: evolution of alternative pathways in the nivalenol chemotype

Abstract

Fusarium sporotrichioides genes FsTri11, FsTri13, and FsTri1 encode cytochrome P450 monooxygenases (CYPs) responsible for hydroxylations at C-15, C-4, and C-8 of the trichothecene skeleton, respectively. However, the corresponding genes of nivalenol (NIV)-chemotype Fusarium graminearum remain to be functionally elucidated. In this study, we characterized the roles of these CYPs in NIV biosynthesis. Analyses of the metabolites of the F. graminearum Fgtri11 mutant, a disruptant of FgTri11 encoding isotrichodermin (ITD) C-15 hydroxylase, revealed a small amount of NIV-type trichothecenes suggesting that an alternative C-15 hydroxylase partially complemented FgTRI11p. In contrast, the C-7/C-8 hydroxylations depended solely on FgTRI1p, as suggested by the metabolite profiles of the Fgtri11 Fgtri1 double gene disruptant. Disruption of FgTri1 in both the wild-type and Fgtri13 mutant backgrounds revealed that FgTRI13p exhibits marginal activity toward calonectrin (CAL) and that it was the only C-4 hydroxylase. In addition, feeding experiments demonstrated that the C-4 hydroxylation of a 7-hydroxytrichothecene lacking C-8 ketone was extremely limited. The marginal activity of FgTRI13p toward CAL was advantageous for the C-7/C-8 hydroxylation steps in NIV biosynthesis, as transformation of a C-4 oxygenated trichothecene lacking C-7/C-8 modifications into NIV-type trichothecenes was quite inefficient. The significance of hydroxylation steps in the evolution of Fusarium trichothecenes is discussed.

Evolution of structural diversity of trichothecenes, a family of toxins produced by plant pathogenic and entomopathogenic fungi

“…graminearum and F . sporotrichioides have genes that can partially compensate for the absence of TRI genes in deletion mutants [ 46 , 47 ]. Such genes encode enzymes that can modify trichothecene structures, and suggest another possible origin of gained TRI genes.…”

Comparing genotype and chemotype of Fusarium graminearum from cereals in Ontario, Canada

“…This means that strain to strain variations in the accumulation of precursor secondary metabolites can result in a variety of end products along what came to be described as a metabolic grid [36–38]. Studies have shown that these differences in substrate specificity of biosynthetic enzymes means that several pathways exist even for the same compound [38–40]. Secondly, as with sequence differences in the TRI8 gene results in populations that produce either 3ADON or 15ADON, genetic variations that arise within other genes involved in secondary metabolism can lead to the production of different chemical end-products.…”

Reduced Toxicity of Trichothecenes, Isotrichodermol, and Deoxynivalenol, by Transgenic Expression of the Tri101 3-O-Acetyltransferase Gene in Cultured Mammalian FM3A Cells

“…Each trichothecene was obtained from rice medium [26] or rice flour liquid medium [27] inoculated with F. graminearum : Gibberella zeae JCM 9873 for DON [28] and F. graminearum DSM 4528 for 3-acetyldeoxynivalenol (3-ADON) [29]. Isotrichodermin was obtained from the culture medium of F. graminearum MAFF 111233 Fgtri11 disruptant ( Fgtri11 - , NBRC 113181) [27], while ITDol was obtained by the deacetylation of ITD in 2.8% ammonium solution. For purification, the ethyl acetate extract was applied to Purif-Rp2 equipped with Purif-Pack SI 30 µm SIZE (Shoko Scientific, Kanagawa, Japan), and the fraction containing a target trichothecene was concentrated.…”