Regulation and evaluation of five methanol-inducible promoters in the methylotrophic yeast Candida boidinii.
We isolated the promoter regions of five methanol-inducible genes (P(AOD1), alcohol oxidase; P(DAS1), dihydroxyacetone synthase; P(FDH1), formate dehydrogenase; P(PMP20), Pmp20; and P(PMP47), Pmp47) from the Candida boidinii genome, and evaluated their strength and studied their regulation using the acid phosphatase gene of Saccharomyces cerevisiae (ScPHO5) as the reporter. Of the five promoters, P(DAS1) was the strongest methanol-inducible promoter whose strength was approximately 1.5 times higher than that of the commonly used P(AOD1) in methanol-induced cells. Although the expression of P(AOD1) and P(DAS1) was completely repressed by the presence of glucose, formate-induced expression of P(FDH1) was not repressed by glucose. Expression under P(PMP47), another methanol-inducible promoter, was highly induced by oleate. The induction kinetics of P(PMP47) and P(DAS1) revealed that methanol induces the expression of peroxisome membrane protein Pmp47, earlier than the expression of matrix enzyme dihydroxyacetone synthase (Das1p), and that this information is contained in the promoter region of the respective gene. This is the first report which evaluates several methanol-inducible promoters in parallel in the methylotrophic yeast.
Yurimoto
H
H
Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-Oiwake, Sakyo-ku, Kyoto 606-8502, Japan.
Komeda
T
T
Lim
C R
CR
Nakagawa
T
T
Kondo
K
K
Kato
N
N
Sakai
Y
Y
D000135:Acid Phosphatase / Q000032:analysis / Q000235:genetics; D000429:Alcohol Oxidoreductases / Q000096:biosynthesis / Q000235:genetics; D019880:Aldehyde-Ketone Transferases / Q000096:biosynthesis / Q000235:genetics; D002175:Candida / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism; D016384:Consensus Sequence; D004790:Enzyme Induction / Q000187:drug effects; D005560:Formate Dehydrogenases; D005656:Fungal Proteins / Q000096:biosynthesis / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D017930:Genes, Reporter; D005947:Glucose; D008565:Membrane Proteins / Q000096:biosynthesis / Q000235:genetics; D000432:Methanol / Q000494:pharmacology*; D010544:Peroxidases; D054464:Peroxiredoxins; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic / Q000502:physiology*; D012441:Saccharomyces cerevisiae; D016415:Sequence Alignment
C499417:FDH1 protein, Candida; D005656:Fungal Proteins; D008565:Membrane Proteins; C066456:PMP47 protein, Candida boidinii; D000429:Alcohol Oxidoreductases; C019643:alcohol oxidase; D010544:Peroxidases; D054464:Peroxiredoxins; D005560:Formate Dehydrogenases; D019880:Aldehyde-Ketone Transferases; C031222:formaldehyde transketolase; D000135:Acid Phosphatase; D005947:Glucose; D000432:Methanol