Abstract
Genes required for ergot alkaloid biosynthesis are clustered in the genomes of several fungi. Several conserved ergot cluster genes have been hypothesized, and in some cases demonstrated, to encode early steps of the pathway shared among fungi that ultimately make different ergot alkaloid end products. The deduced amino acid sequence of one of these conserved genes (easC) indicates a catalase as the product, but a role for a catalase in the ergot alkaloid pathway has not been established. We disrupted easC of Aspergillus fumigatus by homologous recombination with a truncated copy of that gene. The resulting mutant (ΔeasC) failed to produce the ergot alkaloids typically observed in A. fumigatus, including chanoclavine-I, festuclavine, and fumigaclavines B, A, and C. The ΔeasC mutant instead accumulated N-methyl-4-dimethylallyltryptophan (N-Me-DMAT), an intermediate recently shown to accumulate in Claviceps purpurea strains mutated at ccsA (called easE in A. fumigatus) (Lorenz et al. Appl Environ Microbiol 76:1822–1830, 2010). A ΔeasE disruption mutant of A. fumigatus also failed to accumulate chanoclavine-I and downstream ergot alkaloids and, instead, accumulated N-Me-DMAT. Feeding chanoclavine-I to the ΔeasC mutant restored ergot alkaloid production. Complementation of either ΔeasC or ΔeasE mutants with the respective wild-type allele also restored ergot alkaloid production. The easC gene was expressed in Escherichia coli, and the protein product displayed in vitro catalase activity with H2O2 but did not act, in isolation, on N-Me-DMAT as substrate. The data indicate that the products of both easC (catalase) and easE (FAD-dependent oxidoreductase) are required for conversion of N-Me-DMAT to chanoclavine-I.
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Acknowledgments
We thank Christopher L. Schardl (University of Kentucky) for providing plasmid pKAES154 containing C. purpurea dmaW. This work was supported by grant number 2008-35318-04549 from the United States Department of Agriculture National Institute of Food and Agriculture and published with permission of the West Virginia Agriculture and Forestry Experiment Station as scientific article number 3902.
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Communicated by J. Heitman.
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Goetz, K.E., Coyle, C.M., Cheng, J.Z. et al. Ergot cluster-encoded catalase is required for synthesis of chanoclavine-I in Aspergillus fumigatus . Curr Genet 57, 201–211 (2011). https://doi.org/10.1007/s00294-011-0336-4
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DOI: https://doi.org/10.1007/s00294-011-0336-4