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Acetic acid inhibits nutrient uptake in Saccharomyces cerevisiae: auxotrophy confounds the use of yeast deletion libraries for strain improvement

  • Applied microbial and cell physiology
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Abstract

Acetic acid inhibition of yeast fermentation has a negative impact in several industrial processes. As an initial step in the construction of a Saccharomyces cerevisiae strain with increased tolerance for acetic acid, mutations conferring resistance were identified by screening a library of deletion mutants in a multiply auxotrophic genetic background. Of the 23 identified mutations, 11 were then introduced into a prototrophic laboratory strain for further evaluation. Because none of the 11 mutations was found to increase resistance in the prototrophic strain, potential interference by the auxotrophic mutations themselves was investigated. Mutants carrying single auxotrophic mutations were constructed and found to be more sensitive to growth inhibition by acetic acid than an otherwise isogenic prototrophic strain. At a concentration of 80 mM acetic acid at pH 4.8, the initial uptake of uracil, leucine, lysine, histidine, tryptophan, phosphate, and glucose was lower in the prototrophic strain than in a non-acetic acid-treated control. These findings are consistent with two mechanisms by which nutrient uptake may be inhibited. Intracellular adenosine triphosphate (ATP) levels were severely decreased upon acetic acid treatment, which likely slowed ATP-dependent proton symport, the major form of transport in yeast for nutrients other than glucose. In addition, the expression of genes encoding some nutrient transporters was repressed by acetic acid, including HXT1 and HXT3 that encode glucose transporters that operate by facilitated diffusion. These results illustrate how commonly used genetic markers in yeast deletion libraries complicate the effort to isolate strains with increased acetic acid resistance.

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Acknowledgments

We thank Garrett Holzwarth, Yan Campbell, and Chunxiao Guo for the technical assistance and Adrian Gombart and the Oregon State University (OSU) Linus Pauling Institute for the use of the luminometer and RT-qPCR system. The OSU Environmental Health Sciences Center (grant number P30 ES000210; NIEHS, NIH) provided the yeast deletion library. This work was supported by grant no. 2010-65504-20345 from the United States Department of Agriculture National Institute of Food and Agriculture (USDA-NIFA) program.

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Authors and Affiliations

  1. Department of Food Science and Technology, Oregon State University, Corvallis, OR, USA

    Jun Ding, Jan Bierma, Mark R. Smith, Alex N. Hadduck, Mallori Jirikovic, Michael H. Penner & Alan T. Bakalinsky

  2. Department of Biochemistry and Biophysics, Oregon State University, Corvallis, OR, USA

    Jun Ding, Kari van Zee & Alan T. Bakalinsky

  3. Biological Sciences, Duquesne University, Pittsburgh, PA, USA

    Eric Poliner, Carole Wolfe & Jana Patton-Vogt

  4. Dipartimento di Agraria, University of Sassari, Sassari, Italy

    Severino Zara

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  1. Jun Ding
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Correspondence to Alan T. Bakalinsky.

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Ding, J., Bierma, J., Smith, M.R. et al. Acetic acid inhibits nutrient uptake in Saccharomyces cerevisiae: auxotrophy confounds the use of yeast deletion libraries for strain improvement. Appl Microbiol Biotechnol 97, 7405–7416 (2013). https://doi.org/10.1007/s00253-013-5071-y

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  • DOI: https://doi.org/10.1007/s00253-013-5071-y

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