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Characterization of the function of the ver-1A and ver-1B genes, involved in aflatoxin biosynthesis in Aspergillus parasiticus

Author:
Liang, S.H., Skory, C.D., Linz, J.E.
Source:
Applied and environmental microbiology 1996 v.62 no.12 pp. 4568
ISSN:
0099-2240
Subject:
Aspergillus parasiticus, structural genes, polypeptides, oxidoreductases, nucleotide sequences, amino acid sequences, aflatoxins, biosynthesis, genetic transformation, mutants, sterigmatocystin, restriction mapping, genetic complementation, aflatoxin B1
Abstract:
The ver-1A gene was cloned and its nucleotide sequence was determined as part of a previous study on aflatoxin B1(AFB1) biosynthesis in the filamentous fungus Aspergillus parasiticus SU-1. A second copy of this gene, ver-1B, was tentatively identified in this fungal strain. In this study, ver-1B was cloned by screening an A. parasiticus cosmic library with a ver-1A probe. The nucleotide sequence of ver-1B was determined. The predicted amino acid sequence of ver-1B had 95% identity with ver-1A. A translational stop codon, found in the ver-1B gene coding region, indicated that it encodes a truncated polypeptide. To confirm the function of the ver-1 genes in AFB1 synthesis, a plasmid (pDV-VA) was designed to disrupt ver-1A and/or ver-1B by transformation of the AFB, producer A. parasiticus NR-1. One disruptant, VAD-102, which accumulated the pathway intermediate versicolorin A was obtained. Southern hybridization analysis of VAD-102 revealed that ver-1A but not ver-1B was disrupted. A functional ver-1A gene was transformed back into strain VAD-102. Transformants which received ver-1A produced AFB1, confirming that ver-1A is the only functional ver-1 gene in A. parasiticus SU-1 and that its gene product is involved in the conversion of versicolorin A to sterigmatocystin in AFB1 biosynthesis. A duplicated chromosomal region (approximately 12 kb) was identified upstream from ver-1A and ver-1B by Southern hybridization analysis. This duplicated region contained the aflR gene, which is proposed to be one regulator of AFB1 synthesis. A similar gene duplication was also identified in several other strains of A. parasiticus.
Agid:
25169
Handle:
10113/25169