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Two novel classes of enzymes are required for the biosynthesis of aurofusarin in Fusarium graminearum.

ABSTRACT: Previous studies have reported the functional characterization of 9 out of 11 genes found in the gene cluster responsible for biosynthesis of the polyketide pigment aurofusarin in Fusarium graminearum. Here we reanalyze the function of a putative aurofusarin pump (AurT) and the two remaining orphan genes, aurZ and aurS. Targeted gene replacement of aurZ resulted in the discovery that the compound YWA1, rather than nor-rubrofusarin, is the primary product of F. graminearum polyketide synthase 12 (FgPKS12). AurZ is the first representative of a novel class of dehydratases that act on hydroxylated ?-pyrones. Replacement of the aurS gene resulted in accumulation of rubrofusarin, an intermediate that also accumulates when the GIP1, aurF, or aurO genes in the aurofusarin cluster are deleted. Based on the shared phenotype and predicted subcellular localization, we propose that AurS is a member of an extracellular enzyme complex (GIP1-AurF-AurO-AurS) responsible for converting rubrofusarin into aurofusarin. This implies that rubrofusarin, rather than aurofusarin, is pumped across the plasma membrane. Replacement of the putative aurofusarin pump aurT increased the rubrofusarin-to- aurofusarin ratio, supporting that rubrofusarin is normally pumped across the plasma membrane. These results provide functional information on two novel classes of proteins and their contribution to polyketide pigment biosynthesis.

SUBMITTER: Frandsen RJ

PROVIDER: S-EPMC3060495 | biostudies-literature | 2011 Mar

REPOSITORIES: biostudies-literature

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Two novel classes of enzymes are required for the biosynthesis of aurofusarin in Fusarium graminearum.

Frandsen Rasmus J N RJ Schütt Claes C Lund Birgitte W BW Staerk Dan D Nielsen John J Olsson Stefan S Giese Henriette H

The Journal of biological chemistry 20110204 12

Previous studies have reported the functional characterization of 9 out of 11 genes found in the gene cluster responsible for biosynthesis of the polyketide pigment aurofusarin in Fusarium graminearum. Here we reanalyze the function of a putative aurofusarin pump (AurT) and the two remaining orphan genes, aurZ and aurS. Targeted gene replacement of aurZ resulted in the discovery that the compound YWA1, rather than nor-rubrofusarin, is the primary product of F. graminearum polyketide synthase 12 ...[more]

PMID: 21296881

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Project description:3-isopropylmalate dehydrogenase (IPMD) encoded by LEU2 is a key enzyme in leucine (Leu) biosynthetic pathway. Analysis of the genome sequence of Fusarium graminearum revealed two paralogous LEU2 genes (designated as FgLEU2A and FgLEU2B) in this fungus and the deduced amino acid sequences of FgLeu2A and FgLeu2B share 45% identity. Targeted disruption of individual FgLEU2A/B gene in F. graminearum assigned a more crucial role of FgLeu2A in Leu biosynthesis as disruption of FgLEU2A resulted in mutant (?FgLeu2A-10) that was Leu-auxotrophic and could not grow in minimal medium limited for amino acids, whereas FgLEU2B deletion mutant ?FgLeu2B-2 was morphologically indistinguishable from the wild type strain PH-1. The growth defects of ?FgLeu2A-10 could be overcome by exogenous addition of Leu at 0.25 mM. Double deletion of FgLEU2A and FgLEU2B (?FgLeu2AB-8) caused a more severe Leu-auxotrophic phenotype as the concentration of Leu exogenously added to medium to rescue the growth defect of ?FgLeu2AB-8 should be raised to 1.25 mM, indicating a less important but nonnegligible role of FgLeu2B in Leu biosynthesis. Disturb of Leu biosynthesis caused by FgLEU2A deletion leads to slower growth rate, reduced aerial hyphal formation and red pigmentation on PDA plates and completely blocked conidial production and germination. All of the defects above could be overcome by Leu addition or complementation of the full-length FgLEU2A gene. ?FgLeu2A-10 also showed significantly increased sensitivity to osmotic and oxidative stresses. Pathogenicity assay results showed that virulence of mutants lacking FgLEU2A were dramatically impaired on wheat heads and non-host cherry tomatoes. Additionally, a low level of deoxynivalenol (DON) production of ?FgLeu2A-10 and ?FgLeu2AB-8 in wheat kernels was also detected. Taken together, results of this study indicated a crucial role of FgLeu2A and a less important role of FgLeu2B in Leu biosynthesis and fungal infection-related morphogenesis in F. graminearum and FgLeu2A may serve as a potential target for novel antifungal development.

Dataset Information

Two novel classes of enzymes are required for the biosynthesis of aurofusarin in Fusarium graminearum.

Publications

Two novel classes of enzymes are required for the biosynthesis of aurofusarin in Fusarium graminearum.

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