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The objective of this study was to determine if acetosyringone affected the expression of aflatoxin biosynthetic genes.Two genes, nor1 and ver1, representing genes whose products are involved in early and late steps in aflatoxin biosynthesis, were examined. Two GUS (β-glucuronidase) reporter constructs, nor1::GUS (pGAP12) and ver1::GUS (pGAP13), were used to study the effect of acetosyringone on expression of aflatoxin biosynthetic (AF) genes, nor1 and ver1. The product of nor1 is involved in the formation of norsolorinic acid, the first stable intermediate in the aflatoxin pathway. The ver1 gene codes for the enzyme catalyzing the formation of demethylsterigmatocystin, an intermediate late in the AF pathway. GUS activities of these two reporter constructs were inhibited by 80% in the presence of 2 m mol l−1 acetosyringone.Aflatoxin production in a toxigenic strain 42-12 was also shown to be inhibited by acetosyringone to the same level. The levels of inhibition in aflatoxin production and gene transcription are congruous in these three strains.Recent studies have indicated that some phenolics act as signal molecules in plant microbial interactions. Concentration of acetosyringone is shown to increase about ten fold when certain metabolically active plant tissues are wounded. The knowledge gained can be applied to develop strategies in plant breeding programs. The compound may be useful for studying molecular mechanism of modulating aflatoxin biosynthesis.