Cis-acting elements sufficient for induction of FDH1 expression by formate in the methylotrophic yeast Candida boidinii.
The FDH1 gene of Candida boidinii encodes an NAD+-dependent formate dehydrogenase, which catalyzes the last reaction in the methanol dissimilation pathway. FDH1 expression is strongly induced by methanol, as are the promoters of the genes AOD1 (alcohol oxidase) and DAS1 (dihydroxyacetone synthase). FDH1 expression can be induced by formate when cells are grown on a medium containing glucose as a carbon source, whereas expression of AOD1 and DAS1 is completely repressed in the presence of glucose. Using deletion analyses, we identified two cis-acting regulatory elements, termed UAS-FM and UAS-M, respectively, in the 5' non-coding region of the FDH1 gene. Both elements were necessary for full induction of the FDH1 promoter by methanol, while only the UAS-FM element was required for full induction by formate. Irrespective of whether induction was achieved with methanol or formate, the UAS-FM element enhanced the level of induction of the FDH1 promoter in a manner dependent on the number of copies, but independent of their orientation, and also converted the ACT1 promoter from a constitutive into an inducible element. Our results not only provide a powerful promoter for heterologous gene expression, but also yield insights into the mechanism of regulation of FDH1 expression at the molecular level.
Komeda
T
T
Central Laboratories for Key Technology, Kirin Brewery Co, Ltd, 1-13-5 Fukuura, Kanazawa-ku, 236-0004 Yokohama-shi, Kanagawa, Japan. t-komeda@kirin.co.jp
Yurimoto
H
H
Kato
N
N
Sakai
Y
Y
Kondo
K
K
D001483:Base Sequence; D002175:Candida / Q000187:drug effects / Q000235:genetics*; D017931:DNA Primers; D004352:Drug Resistance, Microbial / Q000235:genetics; D005560:Formate Dehydrogenases; D005561:Formates / Q000494:pharmacology*; D005656:Fungal Proteins / Q000235:genetics*; D015966:Gene Expression Regulation, Fungal / Q000235:genetics; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D015183:Restriction Mapping; D017384:Sequence Deletion
D017931:DNA Primers; C499417:FDH1 protein, Candida; D005561:Formates; D005656:Fungal Proteins; C030544:formic acid; D005560:Formate Dehydrogenases