Oxygen and carbon source‐regulated expression of PDC and ADH genes in the respiratory yeast Pichia anomala

Abstract

We amplified, sequenced and studied the transcriptional regulation of genes of the alcoholic fermentation pathway in the biocontrol and non-Saccharomyces wine yeast, Pichia anomala. Two ADH isogenes, PaADH1 and PaADH2, and one PDC gene, PaPDC1, were amplified from genomic P. anomala DNA by a two-step PCR approach, using degenerated primers against conserved regions of the respective genes for cloning core regions, and PCR-based gene walking for cloning the respective 5 and 3 -ends. According to sequence analysis, ADH1 and PDC1 are most likely cytoplasmatic proteins, while ADH2 is most probably localized in the mitochondria. PaADH1 was expressed during aerobic growth on glucose, ethanol and succinate, but was ninefold upregulated in response to oxygen limitation when grown on glucose. The gene seems to be involved in both production and consumption of ethanol. Only low expression of PaADH2 was detected during growth on glucose and ethanol, but it was highly expressed during growth on the non-fermentable carbon source succinate and repressed by the addition of glucose. PaPDC1 was expressed during aerobic growth on glucose and was upregulated four-fold in response to oxygen limitation. PaPDC1 expression was lower in cells grown on ethanol and succinate than on glucose and was up-regulated two-and four-fold, respectively, after glucose addition. Our results demonstrate that transcription of genes of the fermentative pathway is regulated by hypoxia and carbon source but posttranscriptional regulation may play a major role in regulating the metabolic flux.

"Glucose and ethanol-dependent transcriptional regulation of the astaxanthin biosynthesis pathway in Xanthophyllomyces dendrorhous"

“…The mRNA levels of the grg2 gene decreased dramatically when glucose was added to the culture. Moreover, the PDC gene was induced by glucose, as it is in the majority of phylogenetically related organisms [22-25]. In addition, we found that adding glucose to the media caused a decrease in the mRNA levels of all of the carotenogenesis genes involved in the synthesis of astaxanthin from GGPP.…”

“…This gene is highly repressed by glucose in N. crassa and in many other yeasts [20,21]. As a glucose induction control, we used the pyruvate decarboxylase gene PDC , which is induced by glucose in several fungi and yeasts [22-25]. For this experiment, genomic PDC and its cDNA were sequenced, its intron-exon structure was determined and its sequence was deposited in the database [GenBank: HQ694557 and HQ694558].…”

The Alcohol Dehydrogenase System in the Xylose-Fermenting Yeast Candida maltosa

“…Hence, the expression regulation and the physiological function of the CmADH1/CmADH2A system in C. maltosa were essentially similar to those of the ScADH1/ScADH2 system in S. cerevisiae during glucose metabolism [6], [33]. In contrast, the ancestral yeast species seems to contain one cytoplasmic ADH [18], and a dual function of an ADH gene responsible for both formation and consumption of ethanol have been described for PsADH1 of P. stipitis [7], PaADH1 of Pichia anomala [34] and CaADH1 of C. albicans [14]. Gene duplications lead to the modern ADH system composed of more than one ADH genes during yeast evolution (Figure 1).…”

Pichia anomala: cell physiology and biotechnology relative to other yeasts

“…Fredlund et al (2004a) reported that P. anomala exhibited growth rates of 0.22 and 0.056 h -1 and biomass yields of 0.59 and 0.11 g/g glucose under aerobic and anaerobic conditions, respectively. When shifted to oxygen limitation, P. anomala rapidly induced key fermentative enzymes (pyruvate decarboxylase and alcohol dehydrogenase- Fredlund et al 2006) and also lowered flux through the pentose phosphate pathway. S. cerevisiae is also regarded as a facultative yeast, but in this organism, glucose (rather than oxygen) governs central carbon metabolism.…”