Elucidation of the enzyme involved in 2,3,5-triphenyl tetrazolium chloride (TTC) staining activity and the relationship between TTC staining activity and fermentation profiles in Saccharomyces cerevisiae
Section snippets
Yeast strains and culture medium
All S. cerevisiae strains used in this study are listed in Table 1. Yeast strains were maintained in YPD agar medium (2% glucose, 2% peptone, and 1% yeast extract). SC medium (2% glucose, 0.67% yeast nitrogen base w/o amino acids, 0.002% uracil, 0.006% leucine, 0.002% histidine, and 0.002% l-methionine) was used for the selection of gene disruptants, and uracil, leucine, histidine, and methionine were added to the SC medium when they were not used as selective markers. YM medium containing 10%
Gene disruptants of ubiquinol-cytochrome c oxidoreductase (complex III) and cytochrome c oxidase (complex IV) lost TTC staining activity
As shown in Fig. 1A, in S. cerevisiae, electrons are compiled to ubiquinone from each electron donor by enzymes such as inner mitochondrial NADH dehydrogenase (encoded by NDI1), succinate dehydrogenase (encoded by SDH1-4), outer mitochondrial NADH dehydrogenase (encoded by NDE1 and NDE2), glycerol-3-phosphate dehydrogenase (encoded by GUT2), and lactate dehydrogenase (encoded by DLD1 and CYB2) and subsequently, ubiquinol-cytochrome c oxidoreductase and cytochrome c oxidase transfer electrons
Discussion
TTC staining assays using gene disruptants involved in the electron transport system and a comparison of TTC reduction enzyme activities of ubiquinol-cytochrome c oxidoreductase and cytochrome c oxidase showed that ubiquinol-cytochrome c oxidoreductase is directly involved in TTC reduction.
Although Rich et al. (7) reported that mitochondrial NADH dehydrogenase (complex I) reduces TTC in C. reinhardtii by biochemical analysis, in our study, the disruptant of NDI1 corresponding to complex I did
Acknowledgments
The authors declare no conflict of interest.
References (25)
- et al.
The mitochondrial respiratory chain of yeast. Structure and biosynthesis and the role in cellular metabolism
Biochim. Biophys. Acta
(1987) - et al.
The sites of interaction of triphenyltetrazolium chloride with mitochondrial respiratory chains
FEMS Microbiol. Lett.
(2001) - et al.
Purification and subfractionation of mitochondria from the yeast Saccharomyces cerevisiae
Methods Cell Biol.
(2007) - et al.
The cytochrome bc1 and cytochrome c oxidase complexes associate to form a single supracomplex in yeast mitochondria
J. Biol. Chem.
(2000) - et al.
CTP synthetase and its role in phospholipid synthesis in the yeast Saccharomyces cerevisiae
Prog. Lipid Res.
(2008) - et al.
Evaluation of 2,3,5-triphenyltetrazolium chloride staining to delineate rat brain infarcts
Stroke
(1991) - et al.
Detection method for the ability of hemp (Cannabis sativa L.) seed germination by the use of 2,3,5-triphenyl-2H-tetrazolium chloride (TTC)
Yakugaku Zasshi: J. Pharmaceut. Soc. Jpn.
(2008) - et al.
Tetrazolium overlay technique for population studies of respiration deficiency in yeast
Science
(1957) - et al.
Advances in the study of respiration-deficient (RD) mutation in yeast and other microorganisms
Bacteriol. Rev.
(1961) - et al.
A microbial control of sakebrewing. Part IV. On the TTC agar overlay method for grouping of yeast (1)
Nippon Nogeikagaku Kaishi
(1963)
Studies on the reduction of tetrazolium salt by plant tissues: II. Effect of plasmolysis on the reduction of TTC in plant cell
Cytologia
Generalized indicator plate for genetic, metabolic, and taxonomic studies with microorganisms
Appl. Environ. Microbiol.
Cited by (17)
Physiological role of the EHL gene in sake yeast and its effects on quality of sake
2024, Journal of Bioscience and BioengineeringElucidating tetracycline degradation in a coupled system of photocatalysis combined with Bacillus cereus
2024, Journal of Water Process EngineeringThe bio3 mutation in sake yeast leads to changes in organic acid profiles and ester levels but not ethanol production
2023, Journal of Bioscience and BioengineeringOL-FS13 alleviates experimental cerebral ischemia-reperfusion injury
2022, Experimental NeurologyCitation Excerpt :The neuroprotective activity of OL-FS13 in vivo was demonstrated using I/R rats. TTC staining is commonly used to evaluate cerebral ischemic injury, with red indicating un-infarcted brain tissue (Tanaka et al., 2021). As shown in Fig. 3A&B, I/R of the brain on the left side resulted in severe brain infarction (P < 0.0001, I/R vs. control).
Recurrent evolutionary switches of mitochondrial cytochrome c maturation systems in Archaeplastida
2024, Nature CommunicationsJapanese sake making using wild yeasts isolated from natural environments
2024, Bioscience, Biotechnology and Biochemistry