title: Insertional mutagenesis and cloning of the gene required for the biosynthesis of the non-host-specific toxin in Cochliobolus lunatus that causes maize leaf spot.
abstract: The maize pathotype Cochliobolus lunatus causes Curvularia leaf spot by producing a non-host-specific toxin known as methyl 5-(hydroxymethyl) furan-2-carboxylate (M5HF2C). However, related research that explores the genes that control the production of this toxin is rare. In the current work, Agrobacterium tumefaciens-mediated transformation (ATMT) was employed to tag the gene required for the biosynthesis of the toxin. Of the 3,000 ATMT transformants recovered, 4 showed a significant decline in pathogenicity on maize leaves; 1 transformant, T806, produced no detectable M5HF2C. Genomic DNA that flanks the integrated plasmid was recovered from one of the mutants. A cosmid clone of the wild-type strain was isolated using the recovered DNA as a probe. The results of the structural and functional analyses of the region corresponding to the tagged site were then used as a basis to successfully clone one gene, called Clt-1. Bioinformatics analysis showed that the gene coded a BTB domain-containing protein that comprises 745 amino acids. Southern analysis revealed that the gene was localized in the genome as a single copy. The essential roles of Clt-1 in both toxin production and pathogenicity were confirmed by gene disruption experiments. In summary, the novel gene Clt-1 is closely associated with toxin production and pathogen virulence in leaves of susceptible varieties.
authors.forename: Jin-Xin
authors.initials: JX
authors.lastname: Gao
authors.forename: Tong
authors.initials: T
authors.lastname: Liu
authors.forename: Jie
authors.initials: J
authors.lastname: Chen
mesh_terms: D016960:Agrobacterium tumefaciens / Q000235:genetics; D000595:Amino Acid Sequence; D001203:Ascomycota / Q000235:genetics* / Q000378:metabolism / Q000472:pathogenicity; D001483:Base Sequence; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D005663:Furans / Q000378:metabolism*; D008543:Melanins / Q000378:metabolism; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D009183:Mycotoxins / Q000378:metabolism; D010641:Phenotype; D010935:Plant Diseases / Q000382:microbiology*; D018515:Plant Leaves / Q000382:microbiology; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017384:Sequence Deletion; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014774:Virulence; D003313:Zea mays / Q000382:microbiology*
chemical_list: D004271:DNA, Fungal; D005656:Fungal Proteins; D005663:Furans; D008543:Melanins; D009183:Mycotoxins; D014157:Transcription Factors; C060089:2-furoic acid