title: Cloning and characterization of a melanin biosynthetic THR1 reductase gene essential for appressorial penetration of Colletotrichum lagenarium.
abstract: Melanin biosynthesis of Colletotrichum lagenarium is essential for appressorial penetration of the host plant. A melanin deficient mutant 9141 (Thr-) has a defect in the conversion of 1,3,8-trihydroxynaphthalene to vermelone in the melanin biosynthetic pathway. The mutant formed nonmelanized appressoria and had little infectivity on cucumber leaves. A cosmid clone pCR1 was selected from a cosmid library of wild-type C. lagenarium by means of a heterologous probe BRM2, one of the clustered genes involved in melanin biosynthesis of Alternaria alternata. pCR1 transformed the Thr- mutant 9141 to wild-type phenotype. A DNA fragment (THR1) homologous to BRM2 was subcloned from pCR1 and the nucleotide sequence determined. THR1 contains one open reading frame that encodes a protein of 282 amino acids. A transformant resulting from gene disruption showed a light brown phenotype different from the dark brown phenotype of the wild-type 104-T. The transformant formed nonmelanized appressoria and had little infectivity. The THR1 amino acid sequence contains a region highly similar to the Ver1 gene involved in the conversion of versicolorin A to sterigmatocystin in aflatoxin biosynthesis by Aspergillus parasiticus and to the T4HN reductase gene involved in the conversion of 1,3,6,8-tetrahydroxynaphthalene to scytalone and 1,3,8-trihydroxynaphthalene to vermelone in melanin biosynthesis by Magnaporthe grisea. Expression of the THR1 gene during spore germination of C. lagenarium was detected by RNA blotting. We propose that the C. lagenarium THR1 gene encodes a reductase involved in conversion of 1,3,8-trihydroxynaphthalene to vermelone.
authors.affiliation: Laboratory of Plant Pathology, Faculty of Agriculture, Kyoto University, Japan.
authors.forename: N S
authors.initials: NS
authors.lastname: Perpetua
authors.forename: Y
authors.initials: Y
authors.lastname: Kubo
authors.forename: N
authors.initials: N
authors.lastname: Yasuda
authors.forename: Y
authors.initials: Y
authors.lastname: Takano
authors.forename: I
authors.initials: I
authors.lastname: Furusawa
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D016384:Consensus Sequence; D017931:DNA Primers; D005656:Fungal Proteins; D015723:Gene Library; D005800:Genes, Fungal; D008543:Melanins / Q000096:biosynthesis; D003904:Mitosporic Fungi / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D005810:Multigene Family; D010088:Oxidoreductases / Q000096:biosynthesis* / Q000235:genetics; D044925:Oxidoreductases Acting on CH-CH Group Donors; D018515:Plant Leaves; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis; D015183:Restriction Mapping; D017386:Sequence Homology, Amino Acid; D014158:Transcription, Genetic; D014675:Vegetables / Q000382:microbiology
chemical_list: D017931:DNA Primers; D005656:Fungal Proteins; D008543:Melanins; D011994:Recombinant Proteins; D010088:Oxidoreductases; C086024:tetrahydroxynaphthalene reductase; D044925:Oxidoreductases Acting on CH-CH Group Donors