title: Isolation of a phytoalexin-detoxification gene from the plant pathogenic fungus Nectria haematococca by detecting its expression in Aspergillus nidulans.
abstract: Detoxification of the pea phytoalexin pisatin via demethylation, mediated by a cytochrome P-450 monooxygenase, is thought to be important for pathogenicity of the fungus Nectria haematococca on pea. To isolate a fungal gene encoding pisatin demethylating activity (pda), we transformed Aspergillus nidulans with a genomic library of N. haematococca DNA constructed in a cosmid which carried the A. nidulans trpC gene. Transformants were selected for Trp+ and then screened for pda. One transformant among 1250 tested was Pda+ and was less sensitive to pisatin in culture than Pda- A. nidulans. The cosmid containing the gene (PDA) conferring this activity was recovered by phage lambda packaging of transformant genomic DNA. When A. nidulans was transformed with the cloned cosmid, 98% of the Trp+ transformants were Pda+. RNA blots probed with a 3.35 kb subclone carrying PDA indicated that the gene is expressed constitutively in A. nidulans but is inducible by pisatin in N. haematococca.
authors.affiliation: Department of Plant Pathology, Cornell University, Ithaca, NY 14853.
authors.forename: K M
authors.initials: KM
authors.lastname: Weltring
authors.forename: B G
authors.initials: BG
authors.lastname: Turgeon
authors.forename: O C
authors.initials: OC
authors.lastname: Yoder
authors.forename: H D
authors.initials: HD
authors.lastname: VanEtten
mesh_terms: D001203:Ascomycota / Q000187:drug effects / Q000201:enzymology / Q000235:genetics*; D001233:Aspergillus nidulans / Q000235:genetics*; D001578:Benzopyrans / Q000378:metabolism* / Q000494:pharmacology; D003001:Cloning, Molecular; D003360:Cosmids; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005796:Genes; D005800:Genes, Fungal; D008658:Inactivation, Metabolic; D010088:Oxidoreductases / Q000235:genetics*; D010090:Oxidoreductases, O-Demethylating / Q000235:genetics* / Q000378:metabolism; D036343:Pterocarpans; D015183:Restriction Mapping; D014158:Transcription, Genetic; D000094162:Phytoalexins
chemical_list: D001578:Benzopyrans; D003577:Cytochrome P-450 Enzyme System; D010088:Oxidoreductases; D010090:Oxidoreductases, O-Demethylating; D036343:Pterocarpans; C057976:pisatin demethylase; C003161:pisatin; D000094162:Phytoalexins