title: Acetic acid inhibits nutrient uptake in Saccharomyces cerevisiae: auxotrophy confounds the use of yeast deletion libraries for strain improvement.
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.
authors.affiliation: Department of Food Science and Technology, Oregon State University, Corvallis, OR, USA.
authors.forename: Jun
authors.initials: J
authors.lastname: Ding
authors.forename: Jan
authors.initials: J
authors.lastname: Bierma
authors.forename: Mark R
authors.initials: MR
authors.lastname: Smith
authors.forename: Eric
authors.initials: E
authors.lastname: Poliner
authors.forename: Carole
authors.initials: C
authors.lastname: Wolfe
authors.forename: Alex N
authors.initials: AN
authors.lastname: Hadduck
authors.forename: Severino
authors.initials: S
authors.lastname: Zara
authors.forename: Mallori
authors.initials: M
authors.lastname: Jirikovic
authors.forename: Kari
authors.initials: K
authors.lastname: van Zee
authors.forename: Michael H
authors.initials: MH
authors.lastname: Penner
authors.forename: Jana
authors.initials: J
authors.lastname: Patton-Vogt
authors.forename: Alan T
authors.initials: AT
authors.lastname: Bakalinsky
mesh_terms: D019342:Acetic Acid / Q000633:toxicity*; D000596:Amino Acids / Q000378:metabolism; D005285:Fermentation; D017353:Gene Deletion; D015966:Gene Expression Regulation, Fungal / Q000187:drug effects*; D005947:Glucose / Q000378:metabolism; D026901:Membrane Transport Proteins / Q000378:metabolism*; D010710:Phosphates / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000187:drug effects* / Q000378:metabolism*; D014498:Uracil / Q000378:metabolism
chemical_list: D000596:Amino Acids; D026901:Membrane Transport Proteins; D010710:Phosphates; D014498:Uracil; D005947:Glucose; D019342:Acetic Acid