Glutathione reductase from Brassica rapa affects tolerance and the redox state but not fermentation ability in response to oxidative stress in genetically modified Saccharomyces cerevisiae.
To determine whether the exogenous expression of glutathione reductase (GR) from Brassica rapa subsp. pekinensis (BrGR) can reduce the deleterious effects of unfavorable conditions, we constructed a transgenic Saccharomyces cerevisiae strain bearing the GR gene cloned into the yeast expression vector, pVTU260. BrGR expression was confirmed by semi reverse transcriptase-polymerase chain reaction (RT-PCR) analysis, immunoblotting analysis and an enzyme assay. Ectopic BrGR-expression improved cellular glutathione (GSH) homeostasis after higher GSH accumulation in the transgenic yeast than in the wild-type yeast under H(2)O(2)-induced oxidative stress. The BrGR-expressing yeast strain induced the activation of metabolic enzymes (Hxt, G6PDH, GAPDH and Ald), antioxidant systems (Gpx, Trx2, Trx3, Trr1, Tsa1 and porin) and molecular chaperones (Hsp104, Hsp90, Hsp70, Hsp42, Hsp26, Grp, Sti1 and Zpr1), which led to lower oxidative protein damage after a reduction in the level of cellular ROS in the BrGR-expressing yeast strain exposed to H(2)O(2) than in the wild-type yeast strain. BrGR-expression increased the ability to adapt and recover from H(2)O(2)-induced oxidative stress and various stressors, including heat shock, menadione, tert-butyl hydroperoxide, heavy metals, sodium dodecyl sulfate, ethanol and NaCl, but did not affect fermentation capacity. These results suggest that ectopic BrGR expression confers acquired tolerance by improving proteostasis and redox homeostasis through co-activation of various cell rescue proteins against ROS-induced oxidative stress in yeast cells.
Department of Biology, Kyungpook National University, #1370 Sankyuk-dong, Buk-gu, Daegu 702-701, Republic of Korea.
Ho-Sung
HS
Yoon
Sun-Young
SY
Shin
Young-Saeng
YS
Kim
Il-Sup
IS
Kim
D029741:Brassica rapa / Q000201:enzymology* / Q000235:genetics; D003001:Cloning, Molecular; D005285:Fermentation; D015870:Gene Expression; D005822:Genetic Vectors; D005978:Glutathione / Q000378:metabolism; D005980:Glutathione Reductase / Q000235:genetics / Q000378:metabolism*; D006861:Hydrogen Peroxide / Q000633:toxicity; D030781:Organisms, Genetically Modified / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D017382:Reactive Oxygen Species / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D013312:Stress, Physiological
D017382:Reactive Oxygen Species; D011994:Recombinant Proteins; D006861:Hydrogen Peroxide; D005980:Glutathione Reductase; D005978:Glutathione