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The filamentous hemiascomycete Ashbya gossypii is a strong riboflavin overproducer. A striking but as yet uninvestigated phenomenon is the fact that the overproduction of this vitamin starts when growth rate declines, which means that most of the riboflavin is produced in the stationary phase, the so-called production phase. The specific activity of 3,4-dihydroxy-2-butanone 4-phosphate (DHBP) synthase, the first enzyme in the biosynthetic pathway for riboflavin, was determined during cultivation and an increase during the production phase was found. Furthermore, an increase of RIB3 mRNA, encoding DHBP synthase, was observed by competitive RT-PCR in the production phase. The mRNAs of two housekeeping genes, ACT1 (encoding actin) and TEF (encoding translation elongation factor-1 alpha), served as standards in the RT-PCR. Reporter studies with a RIB3 promoter-lacZ fusion showed an increase of beta-galactosidase specific activity in the production phase. This investigation verified that the increase of RIB3 mRNA in the production phase is caused by an induction of promoter activity. These data suggest that the time course of riboflavin overproduction of A. gossypii is correlated with a transcriptional regulation of the DHBP synthase.

Transcriptional regulation of 3,4-dihydroxy-2-butanone 4-phosphate synthase