Comparative Polygenic Analysis of Maximal Ethanol Accumulation Capacity and Tolerance to High Ethanol Levels of Cell Proliferation in Yeast

English version České info

The yeast Saccharomyces cerevisiae is able to accumulate ≥17% ethanol (v/v) by fermentation in the absence of cell proliferation. The genetic basis of this unique capacity is unknown. Up to now, all research has focused on tolerance of yeast cell proliferation to high ethanol levels. Comparison of maximal ethanol accumulation capacity and ethanol tolerance of cell proliferation in 68 yeast strains showed a poor correlation, but higher ethanol tolerance of cell proliferation clearly increased the likelihood of superior maximal ethanol accumulation capacity. We have applied pooled-segregant whole-genome sequence analysis to identify the polygenic basis of these two complex traits using segregants from a cross of a haploid derivative of the sake strain CBS1585 and the lab strain BY. From a total of 301 segregants, 22 superior segregants accumulating ≥17% ethanol in small-scale fermentations and 32 superior segregants growing in the presence of 18% ethanol, were separately pooled and sequenced. Plotting SNP variant frequency against chromosomal position revealed eleven and eight Quantitative Trait Loci (QTLs) for the two traits, respectively, and showed that the genetic basis of the two traits is partially different. Fine-mapping and Reciprocal Hemizygosity Analysis identified ADE1, URA3, and KIN3, encoding a protein kinase involved in DNA damage repair, as specific causative genes for maximal ethanol accumulation capacity. These genes, as well as the previously identified MKT1 gene, were not linked in this genetic background to tolerance of cell proliferation to high ethanol levels. The superior KIN3 allele contained two SNPs, which are absent in all yeast strains sequenced up to now. This work provides the first insight in the genetic basis of maximal ethanol accumulation capacity in yeast and reveals for the first time the importance of DNA damage repair in yeast ethanol tolerance.

Vyšlo v časopise: Comparative Polygenic Analysis of Maximal Ethanol Accumulation Capacity and Tolerance to High Ethanol Levels of Cell Proliferation in Yeast. PLoS Genet 9(6): e32767. doi:10.1371/journal.pgen.1003548
Kategorie: Research Article
prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.pgen.1003548

Souhrn

Zdroje

1. CaseyGP, IngledewWM (1986) Ethanol tolerance in yeasts. Crit Rev Microbiol 13: 219–280.

2. D'AmoreT, StewartGG (1987) Ethanol tolerance of yeast. Enzyme and Microbial Technology 9: 322–330.

3. RozpedowskaE, HellborgL, IshchukOP, OrhanF, GalafassiS, et al. (2011) Parallel evolution of the make-accumulate-consume strategy in Saccharomyces and Dekkera yeasts. Nat Commun 2: 302.

4. DingJ, HuangX, ZhangL, ZhaoN, YangD, et al. (2009) Tolerance and stress response to ethanol in the yeast Saccharomyces cerevisiae. Appl Microbiol Biotechnol 85: 253–263.

5. SwinnenS, SchaerlaekensK, PaisT, ClaesenJ, HubmannG, et al. (2012) Identification of novel causative genes determining the complex trait of high ethanol tolerance in yeast using pooled-segregant whole-genome sequence analysis. Genome Res 22: 975–984.

6. LitiG, LouisEJ (2012) Advances in quantitative trait analysis in yeast. PLoS Genet 8: e1002912.

7. SwinnenS, TheveleinJM, NevoigtE (2012) Genetic mapping of quantitative phenotypic traits in Saccharomyces cerevisiae. FEMS Yeast Res 12: 215–227.

8. EhrenreichIM, TorabiN, JiaY, KentJ, MartisS, et al. (2010) Dissection of genetically complex traits with extremely large pools of yeast segregants. Nature 464: 1039–1042.

9. PartsL, CubillosFA, WarringerJ, JainK, SalinasF, et al. (2011) Revealing the genetic structure of a trait by sequencing a population under selection. Genome Res 21: 1131–1138.

10. SteinmetzLM, SinhaH, RichardsDR, SpiegelmanJI, OefnerPJ, et al. (2002) Dissecting the architecture of a quantitative trait locus in yeast. Nature 416: 326–330.

11. WinzelerEA, RichardsDR, ConwayAR, GoldsteinAL, KalmanS, et al. (1998) Direct allelic variation scanning of the yeast genome. Science 281: 1194–1197.

12. DeutschbauerAM, DavisRW (2005) Quantitative trait loci mapped to single-nucleotide resolution in yeast. Nat Genet 37: 1333–1340.

13. BremRB, YvertG, ClintonR, KruglyakL (2002) Genetic dissection of transcriptional regulation in budding yeast. Science 296: 752–755.

14. MarulloP, AigleM, BelyM, Masneuf-PomaredeI, DurrensP, et al. (2007) Single QTL mapping and nucleotide-level resolution of a physiologic trait in wine Saccharomyces cerevisiae strains. FEMS Yeast Res 7: 941–952.

15. NogamiS, OhyaY, YvertG (2007) Genetic complexity and quantitative trait loci mapping of yeast morphological traits. PLoS Genet 3: e31.

16. PerlsteinEO, RuderferDM, RobertsDC, SchreiberSL, KruglyakL (2007) Genetic basis of individual differences in the response to small-molecule drugs in yeast. Nat Genet 39: 496–502.

17. PuligundiaP, SmogrovicovaD, ObulamVSR, KoS (2011) Very high gravity (VHG) ethanolic brewing and fermentation: a research update. J Ind Microbiol Biotechnol 38: 1133–1144.

18. AlbersE, LarssonC (2009) A comparison of stress tolerance in YPD and industrial lignocellulose-based medium among industrial and laboratory yeast strains. J Ind Microbiol Biotechnol 36: 1085–1091.

19. BassoTO, DarioMG, TonsoA, StambukBU, GombertAK (2010) Insufficient uracil supply in fully aerobic chemostat cultures of Saccharomyces cerevisiae leads to respiro-fermentative metabolism and double nutrient-limitation. Biotechnol Lett 32: 973–977.

20. WatanabeM, WatanabeD, AkaoT, ShimoiH (2009) Overexpression of MSN2 in a sake yeast strain promotes ethanol tolerance and increases ethanol production in sake brewing. J Biosci Bioeng 107: 516–518.

21. Kodama K (1993) Sake-brewing yeast. In: Rose AH, Harrison JS, editors. The yeasts. London, United Kingdom: Academic Press. pp. 129–168.

22. ClaesenJ, ClementL, ShkedyZ, Foulquié-MorenoMR, BurzykowskiT (2013) Simultaneous mapping of multiple gene loci with pooled segregants. PLoS ONE 8(2): e55133 doi:10.1371/journal.pone.0055133

23. DohmJC, LottazC, BorodinaT, HimmelbauerH (2008) Substantial biases in ultra-short read data sets from high-throughput DNA sequencing. Nucleic Acids Res 36: e105.

24. AkaoT, YashiroI, HosoyamaA, KitagakiH, HorikawaH, et al. (2011) Whole-genome sequencing of sake yeast Saccharomyces cerevisiae Kyokai no. 7. DNA Res 18: 423–434.

25. ScheetP, StephensM (2006) A fast and flexible statistical model for large-scale population genotype data: applications to inferring missing genotypes and haplotypic phase. Am J Hum Genet 78: 629–644.

26. CullenPJ, SpragueGFJr (2002) The Glc7p-interacting protein Bud14p attenuates polarized growth, pheromone response, and filamentous growth in Saccharomyces cerevisiae. Eukaryot Cell 1: 884–894.

27. MyasnikovAN, SasnauskasKV, JanulaitisAA, SmirnovMN (1991) The Saccharomyces cerevisiae ADE1 gene: structure, overexpression and possible regulation by general amino acid control. Gene 109: 143–147.

28. MouraDJ, CastilhosB, ImmichBF, CanedoAD, HenriquesJA, et al. (2010) Kin3 protein, a NIMA-related kinase of Saccharomyces cerevisiae, is involved in DNA adduct damage response. Cell Cycle 9: 2220–2229.

29. BardinAJ, BoselliMG, AmonA (2003) Mitotic exit regulation through distinct domains within the protein kinase Cdc15. Mol Cell Biol 23: 5018–5030.

30. RistowH, SeyfarthA, LochmannER (1995) Chromosomal damages by ethanol and acetaldehyde in Saccharomyces cerevisiae as studied by pulsed field gel electrophoresis. Mutat Res 326: 165–170.

31. KitagakiH, ArakiY, FunatoK, ShimoiH (2007) Ethanol-induced death in yeast exhibits features of apoptosis mediated by mitochondrial fission pathway. FEBS Lett 581: 2935–2942.

32. IbeasJI, JimenezJ (1997) Mitochondrial DNA loss caused by ethanol in Saccharomyces flor yeasts. Appl Environ Microbiol 63: 7–12.

33. KidoR, SatoI, TsudaS (2006) Detection of in vivo DNA damage induced by ethanol in multiple organs of pregnant mice using the alkaline single cell gel electrophoresis (Comet) assay. J Vet Med Sci 68: 41–47.

34. BrooksPJ (1997) DNA damage, DNA repair, and alcohol toxicity–a review. Alcohol Clin Exp Res 21: 1073–1082.

35. UmezuK, AmayaT, YoshimotoA, TomitaK (1971) Purification and properties of orotidine-5′-phosphate pyrophosphorylase and orotidine-5′-phosphate decarboxylase from baker's yeast. J Biochem 70: 249–262.

36. MillerBG, HassellAM, WolfendenR, MilburnMV, ShortSA (2000) Anatomy of a proficient enzyme: the structure of orotidine 5′-monophosphate decarboxylase in the presence and absence of a potential transition state analog. Proc Natl Acad Sci U S A 97: 2011–2016.

37. EddyAA, HopkinsP (1998) Proton stoichiometry of the overexpressed uracil symport of the yeast Saccharomyces cerevisiae. Biochem J 336(Pt 1): 125–130.

38. VollandC, Urban-GrimalD, GeraudG, Haguenauer-TsapisR (1994) Endocytosis and degradation of the yeast uracil permease under adverse conditions. J Biol Chem 269: 9833–9841.

39. FerrerasJM, IglesiasR, GirbesT (1989) Effect of the chronic ethanol action on the activity of the general amino-acid permease from Saccharomyces cerevisiae var. ellipsoideus. Biochim Biophys Acta 979: 375–377.

40. AndoA, TanakaF, MurataY, TakagiH, ShimaJ (2006) Identification and classification of genes required for tolerance to high-sucrose stress revealed by genome-wide screening of Saccharomyces cerevisiae. FEMS Yeast Res 6: 249–267.

41. HohmannS (2002) Osmotic stress signaling and osmoadaptation in yeasts. Microbiol Mol Biol Rev 66: 300–372.

42. BoydAR, GunasekeraTS, AttfieldPV, SimicK, VincentSF, et al. (2003) A flow-cytometric method for determination of yeast viability and cell number in a brewery. FEMS Yeast Res 3: 11–16.

43. ShermanF, HicksJ (1991) Micromanipulation and dissection of asci. Methods Enzymol 194: 21–37.

44. HuxleyC, GreenED, DunhamI (1990) Rapid assessment of S. cerevisiae mating type by PCR. Trends Genet 6: 236.

45. Johnston JR, editor (1994) Molecular genetics of yeast: a practical approach. New York: Oxford University Press.

46. BenjaminiY, YekutieliD (2005) Quantitative trait Loci analysis using the false discovery rate. Genetics 171: 783–790.

47. GietzRD, SchiestlRH, WillemsAR, WoodsRA (1995) Studies on the transformation of intact yeast cells by the LiAc/SS-DNA/PEG procedure. Yeast 11: 355–360.

48. HoffmanCS, WinstonF (1987) A ten-minute DNA preparation from yeast efficiently releases autonomous plasmids for transformation of Escherichia coli. Gene 57: 267–272.

49. ShepherdA, PiperPW (2010) The Fps1p aquaglyceroporin facilitates the use of small aliphatic amides as a nitrogen source by amidase-expressing yeasts. FEMS Yeast Res 10: 527–534.