Skip to main content
Log in

FLO gene-dependent phenotypes in industrial wine yeast strains

  • Applied Genetics and Molecular Biotechnology
  • Published:
Applied Microbiology and Biotechnology Aims and scope Submit manuscript

Abstract

Most commercial yeast strains are nonflocculent. However, controlled flocculation phenotypes could provide significant benefits to many fermentation-based industries. In nonflocculent laboratory strains, it has been demonstrated that it is possible to adjust flocculation and adhesion phenotypes to desired specifications by altering expression of the otherwise silent but dominant flocculation (FLO) genes. However, FLO genes are characterized by high allele heterogeneity and are subjected to epigenetic regulation. Extrapolation of data obtained in laboratory strains to industrial strains may therefore not always be applicable. Here, we assess the adhesion phenotypes that are associated with the expression of a chromosomal copy of the FLO1, FLO5, or FLO11 open reading frame in two nonflocculent commercial wine yeast strains, BM45 and VIN13. The chromosomal promoters of these genes were replaced with stationary phase-inducible promoters of the HSP30 and ADH2 genes. Under standard laboratory and wine making conditions, the strategy resulted in expected and stable expression patterns of these genes in both strains. However, the specific impact of the expression of individual FLO genes showed significant differences between the two wine strains and with corresponding phenotypes in laboratory strains. The data suggest that optimization of the flocculation pattern of individual commercial strains will have to be based on a strain-by-strain approach.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8
Fig. 9

Similar content being viewed by others

References

  • Ausubel FM, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, Struhl K (1995) Current protocols in molecular biology. Wiley, New York

    Google Scholar 

  • Bely M, Sablayrolles JM, Barre P (1990) Automatic detection of assimilable nitrogen deficiencies during alcoholic fermentation in enological conditions. J Ferment Bioeng 70:246–252

    Article  CAS  Google Scholar 

  • Bony M, Thines-Sempoux D, Barre P, Blondin B (1997) Localization and cell surface anchoring of the Saccharomyces cerevisiae flocculation protein Flo1p. J Bacteriol 179:4929–4936

    CAS  Google Scholar 

  • Caro LH, Tettelin H, Vossen JH, Ram AF, van den Ende H, Klis FM (1997) In silicio identification of glycosyl-phosphatidylinositol-anchored plasma-membrane and cell wall proteins of Saccharomyces cerevisiae. Yeast 13:1477–1489

    Article  CAS  Google Scholar 

  • Chambers P, Issaka A, Palecek SP (2004) Saccharomyces cerevisiae JEN1 promoter activity is inversely related to concentration of repressing sugar. Appl Environ Microbiol 70:8–17

    Article  CAS  Google Scholar 

  • Ciriacy M (1997) Alcohol dehydrogenases. In: Zimmermann FK, Entian K-D (eds) Yeast sugar metabolism: biochemistry, genetics, biotechnology and applications. Technomic, Lancaster, pp 213–223

    Google Scholar 

  • Cunha AF, Missawa SK, Gomes LH, Reis SF, Pereira GA (2006) Control by sugar of Saccharomyces cerevisiae flocculation for industrial ethanol production. FEMS Yeast Res 6:280–287

    Article  CAS  Google Scholar 

  • D’Hautcourt O, Smart KA (1999) The measurement of brewing yeast flocculation. J Am Soc Brew Chem 57:123–128

    Google Scholar 

  • De Groot PW, Hellingwerf KJ, Klis FM (2003) Genome-wide identification of fungal GPI proteins. Yeast 20:781–796

    Article  Google Scholar 

  • Du Preez JC, Mare JE, Albertyn J, Kilian SG (2001) Transcriptional repression of ADH2-regulated beta-xylanase production by ethanol in recombinant strains of Saccharomyces cerevisiae. FEMS Yeast Res 1:233–240

    Article  Google Scholar 

  • Fichtner L, Schulze F, Braus GH (2007) Differential Flo8p-dependent regulation of FLO1 and FLO11 for cell–cell and cell-substrate adherence of S. cerevisiae S288C. Mol Microbiol 66:1276–1289

    Article  CAS  Google Scholar 

  • Fidalgo M, Barrales RR, Ibeas JI, Jimenez J (2006) Adaptive evolution by mutations in the FLO11 gene. Proc Natl Acad Sci USA 103:11228–11233

    Article  CAS  Google Scholar 

  • Govender P, Domingo JL, Bester MC, Pretorius IS, Bauer FF (2008) Controlled expression of the dominant flocculation genes FLO1, FLO5, and FLO11 in Saccharomyces cerevisiae. Appl Environ Microbiol 74:6041–6052

    Article  CAS  Google Scholar 

  • Guo B, Styles CA, Feng Q, Fink GR (2000) A Saccharomyces gene family involved in invasive growth, cell–cell adhesion, and mating. Proc Natl Acad Sci USA 97:12158–12163

    Article  CAS  Google Scholar 

  • Hamada K, Fukuchi S, Arisawa M, Baba M, Kitada K (1998) Screening for glycosylphosphatidylinositol (GPI)-dependent cell wall proteins in Saccharomyces cerevisiae. Mol Gen Genet 258:53–59

    Article  CAS  Google Scholar 

  • Hinchcliffe E, Box WG, Walton EF, Appleby M (1985) The influences of cell wall hydrophobicity on the top fermenting properties of brewing yeast. Proc Eur Brew Congr 20:323–330

    Google Scholar 

  • Ishigami M, Nakagawa Y, Hayakawa M, Iimura Y (2004) FLO11 is essential for flor formation caused by the C-terminal deletion of NRG1 in Saccharomyces cerevisiae. FEMS Microbiol Lett 237:425–430

    CAS  Google Scholar 

  • James TC, Campbell SG, Bond UM (2002) Comparative analysis of global gene expression in lager and laboratory yeast strains grown in wort. Proc IEEE 90:1887–1899

    Article  CAS  Google Scholar 

  • Kealey JT, Liu L, Santi DV, Betlach MC, Barr PJ (1998) Production of a polyketide natural product in nonpolyketide-producing prokaryotic and eukaryotic hosts. Proc Natl Acad Sci USA 95:505–509

    Article  CAS  Google Scholar 

  • Kjeldsen T (2000) Yeast secretory expression of insulin precursors. Appl Microbiol Biotechnol 54:277–286

    Article  CAS  Google Scholar 

  • Kobayashi O, Hayashi N, Kuroki R, Sone H (1998) Region of FLO1 proteins responsible for sugar recognition. J Bacteriol 180:6503–6510

    CAS  Google Scholar 

  • Lee KM, DaSilva NA (2005) Evaluation of the Saccharomyces cerevisiae ADH2 promoter for protein synthesis. Yeast 22:431–440

    Article  CAS  Google Scholar 

  • Lipke PN, Ovalle R (1998) Cell wall architecture in yeast: new structure and new challenges. J Bacteriol 180:3735–3740

    CAS  Google Scholar 

  • Manthey GM, Navarro MS, Bailis AM (2004) DNA fragment transplacement in Saccharomyces cerevisiae: some genetic considerations. Methods Mol Biol 262:157–172

    CAS  Google Scholar 

  • Maury J, Asadollahi MA, Moller K, Clark A, Nielsen J (2005) Microbial isoprenoid production: an example of green chemistry through metabolic engineering. Adv Biochem Eng Biotechnol 100:19–51

    CAS  Google Scholar 

  • Ness F, Lavallée F, Dubourdieu D, Aigle M, Dulau L (1993) Identification of yeast strains using the polymerase chain reaction. J Sci Food Agric 62:89–94

    Article  CAS  Google Scholar 

  • Nevoigt E, Kohnke J, Fischer CR, Alper H, Stahl U, Stephanopoulos G (2006) Engineering of promoter replacement cassettes for fine-tuning of gene expression in Saccharomyces cerevisiae. Appl Environ Microbiol 72:5266–5273

    Article  CAS  Google Scholar 

  • Noronha SB, Kaslow DC, Shiloach J (1998) Transition phase in the production of recombinant proteins in yeast under the ADH2 promoter: an important step for reproducible manufacturing of a malaria transmission blocking vaccine candidate. J Ind Microbiol 20:192–199

    Article  CAS  Google Scholar 

  • Pretorius IS (2000) Tailoring wine yeast for the new millennium: novel approaches to the ancient art of winemaking. Yeast 16:675–729

    Article  CAS  Google Scholar 

  • Pretorius IS, Bauer FF (2002) Meeting the consumer challenge through genetically customized wine-yeast strains. Trends Biotechnol 20:426–432

    Article  CAS  Google Scholar 

  • Rossouw D, Naes T, Bauer FF (2008) Linking gene regulation and the exo-metabolome: a comparative transcriptomics approach to identify genes that impact on the production of volatile aroma compounds in yeast. BMC Genomics 9:530

    Article  Google Scholar 

  • Saitoh S, Ishida N, Onishi T, Tokuhiro K, Nagamori E, Kitamoto K, Takahashi H (2005) Genetically engineered wine yeast produces a high concentration of L-lactic acid of extremely high optical purity. Appl Environ Microbiol 71:2789–2792

    Article  CAS  Google Scholar 

  • Sambrook KJ, Fritsch EF, Maniatis T (1989) Molecular cloning. A laboratory manual. Cold Spring Harbor Laboratory, Cold Spring Harbor

    Google Scholar 

  • Smukalla S, Caldara M, Pochet N, Beauvais A, Guadagnini S, Yan C, Vinces MD, Jansen A, Prevost MC, Latgé JP, Fink GR, Foster KR, Verstrepen KJ (2008) FLO1 is a variable green beard gene that drives biofilm-like cooperation in budding yeast. Cell 135:726–737

    Article  CAS  Google Scholar 

  • Stratford M (1989) Evidence for two mechanisms of flocculation in Saccharomyces cerevisiae. Yeast 5:S441–S445

    Article  CAS  Google Scholar 

  • Stratford M, Assinder S (1991) Yeast flocculation: Flo1 and NewFlo phenotypes and receptor structure. Yeast 7:559–574

    Article  CAS  Google Scholar 

  • Teunissen AW, Steensma HY (1995) Review: the dominant flocculation genes of Saccharomyces cerevisiae constitute a new subtelomeric gene family. Yeast 11:1001–1013

    Article  CAS  Google Scholar 

  • Thomson JM, Gaucher EA, Burgan MF, De Kee DW, Li T, Aris JP, Benner SA (2005) Resurrecting ancestral alcohol dehydrogenases from yeast. Nat Genet 37:630–635

    Article  CAS  Google Scholar 

  • Van Mulders SE, Christianen E, Saerens SM, Daenen L, Verbelen PJ, Willaert R, Verstrepen KJ, Delvaux FR (2009) Phenotypic diversity of Flo protein family-mediated adhesion in Saccharomyces cerevisiae. FEMS Yeast Res 9:178–190

    Article  Google Scholar 

  • Varela C, Cardenas J, Melo F, Agosin E (2005) Quantitative analysis of wine yeast gene expression profiles under winemaking conditions. Yeast 22:369–383

    Article  CAS  Google Scholar 

  • Verstrepen KJ, Klis FM (2006) Flocculation, adhesion and biofilm formation in yeasts. Mol Microbiol 60:5–15

    Article  CAS  Google Scholar 

  • Verstrepen KJ, Michiels C, Derdelinckx G, Delvaux FR, Winderickx J, Thevelein JM, Bauer FF, Pretorius IS (2001) Late fermentation expression of FLO1 in Saccharomyces cerevisiae. J Am Soc Brew Chem 59:69–76

    CAS  Google Scholar 

  • Verstrepen KJ, Reynolds TB, Fink GR (2004) Origins of variation in the fungal cell surface. Nat Rev Microbiol 2:533–540

    Article  CAS  Google Scholar 

  • Verstrepen KJ, Jansen A, Lewitter F, Fink GR (2005) Intragenic tandem repeats generate functional variability. Nat Genet 37:986–990

    Article  CAS  Google Scholar 

  • Wang D, Wang Z, Liu N, He X, Zhang B (2008) Genetic modification of industrial yeast strains to obtain controllable NewFlo flocculation property and lower diacetyl production. Biotechnol Lett 30:2013–2018

    Article  CAS  Google Scholar 

  • Watari J, Takata Y, Ogawa M, Murakami J, Koshino S (1991) Breeding of flocculent industrial Saccharomyces cerevisiae strains by introducing the flocculation gene FLO1. Agric Biol Chem 55:1547–1552

    CAS  Google Scholar 

  • Wills C (1976) Production of yeast alcohol dehydrogenase isoenzymes by selection. Nature 261:26–29

    Article  CAS  Google Scholar 

  • Winston F, Dollard C, Ricupero-Hovasse SL (1995) Construction of a set of convenient Saccharomyces cerevisiae strains that are isogenic to S288C. Yeast 11:53–55

    Article  CAS  Google Scholar 

Download references

Acknowledgments

This work was financially supported by the National Research Foundation (NRF) and the South African Wine Industry (Winetech).

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Florian F. Bauer.

Electronic supplementary material

Below is the link to the electronic supplementary material.

S1

(DOC 475 kb)

S2

(DOC 467 kb)

S3

(DOC 37 kb)

S4

(DOC 39 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Govender, P., Bester, M. & Bauer, F.F. FLO gene-dependent phenotypes in industrial wine yeast strains. Appl Microbiol Biotechnol 86, 931–945 (2010). https://doi.org/10.1007/s00253-009-2381-1

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00253-009-2381-1

Keywords

Navigation