The Saccharomyces cerevisiae YMR315W gene encodes an NADP(H)-specific oxidoreductase regulated by the transcription factor Stb5p in response to NADPH limitation.
Engineered xylose-metabolizing Saccharomyces cerevisiae cells grown on xylose show increased expression of YMR315W at both the mRNA and protein levels. Additionally, the YMR315W promoter contains a putative binding site for the transcription factor Stb5p, which has been shown to regulate genes involved in NADPH production such as ZWF1, GND1 and GND2. We hypothesized that Ymr315wp, a conserved protein of unknown function, is an additional source of NADPH in wild-type cells. In this study, we purified histidine-tagged enzyme and determined that Ymr315wp is an NADP(H)-specific oxidoreductase. We also showed that YMR315W transcription is regulated by Stb5p in response to diamide induced NADPH depletion. Overexpression of Ymr315wp in BY4727 cells resulted in elevated NADPH levels and increased resistance to diamide. However, the presence of Ymr315wp in cells lacking the oxidative branch of the pentose phosphate pathway resulted in decreased NADPH levels and increased diamide sensitivity. These results suggest that in BY4727 cells Ymr315wp contributes to NADPH production as an alternative source of NADPH.
Hector
Ronald E
RE
Fermentation Biotechnology Research Unit, United States Department of Agriculture, Agricultural Research Service, National Center for Agricultural Utilization Research, Peoria, IL 61604, USA. Ronald.Hector@ars.usda.gov
Bowman
Michael J
MJ
Skory
Christopher D
CD
Cotta
Michael A
MA
D001483:Base Sequence; D003958:Diamide / Q000494:pharmacology; D025141:Drug Resistance, Fungal / Q000187:drug effects; D015966:Gene Expression Regulation, Fungal / Q000187:drug effects; D005800:Genes, Fungal; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D009247:NADH, NADPH Oxidoreductases / Q000235:genetics* / Q000378:metabolism; D009249:NADP / Q000494:pharmacology*; D011401:Promoter Regions, Genetic / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000201:enzymology / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism; D013379:Substrate Specificity / Q000187:drug effects; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects; D015533:Transcriptional Activation / Q000187:drug effects
D029701:Saccharomyces cerevisiae Proteins; C488251:Stb5 protein, S cerevisiae; D014157:Transcription Factors; D003958:Diamide; D009249:NADP; D009247:NADH, NADPH Oxidoreductases; C546951:YMR315w protein, S cerevisiae