The Hrk1 kinase is a determinant of acetic acid tolerance in yeast by modulating H+ and K+ homeostasis

Abstract

Acetic acid-induced stress is a common challenge in natural environments and industrial bioprocesses, significantly affecting the growth and metabolic performance of Saccharomyces cerevisiae. The adaptive response and tolerance to this stress involves the activation of a complex network of molecular pathways. This study aims to delve deeper into these mechanisms in S. cerevisiae, particularly focusing on the role of the Hrk1 kinase. Hrk1 is a key determinant of acetic acid tolerance, belonging to the NPR/Hal family, whose members are implicated in the modulation of the activity of plasma membrane transporters that orchestrate nutrient uptake and ion homeostasis. The influence of Hrk1 on S. cerevisiae adaptation to acetic acid-induced stress was explored by employing a physiological approach based on previous phosphoproteomics analyses. The results from this study reflect the multifunctional roles of Hrk1 in maintaining proton and potassium homeostasis during different phases of acetic acid-stressed cultivation. Hrk1 is shown to play a role in the activation of plasma membrane H+-ATPase, maintaining pH homeostasis, and in the modulation of plasma membrane potential under acetic acid stressed cultivation. Potassium (K+) supplementation of the growth medium, particularly when provided at limiting concentrations, led to a notable improvement in acetic acid stress tolerance of the hrk1∆ strain. Moreover, abrogation of this kinase expression is shown to confer a physiological advantage to growth under K+ limitation also in the absence of acetic acid stress. The involvement of the alkali metal cation/H+ exchanger Nha1, another proposed molecular target of Hrk1, in improving yeast growth under K+ limitation or acetic acid stress, is proposed.

The role of ion homeostasis in adaptation and tolerance to acetic acid stress in yeasts

“…The activation of Pma1 is among the cellular responses when cells are exposed to acetic acid stress (Carmelo et al 1997 , Stratford et al 2013a , Antunes et al 2023 ). Under this stress, Pma1 actively pumps protons out of the cell, helping to restore pHi up to more physiological values and preventing further acidification and potential harm to the cell (Carmelo et al 1997 ).…”

“…Specific yeast protein kinases play an instrumental role in the activation of Pma1 through phosphorylation. The protein kinase Hrk1 is a major acetic acid stress tolerance determinant in yeast (Antunes et al 2018 , 2023 , Mira et al 2010b , Bernardo et al 2017 , Guerreiro et al 2017 , Xu et al 2019a ), belonging to the NPR/Hal family (Antunes and Sá-Correia 2022 ). Under acetic acid stress conditions, Hrk1 contributes to the phosphorylation of Pma1 residues Ser911 and Thr912 (Guerreiro et al 2017 ), whereas in response to glucose metabolism the kinases Ptk1 and Ptk2 act both in a largely redundant manner in mediating the phosphorylation of Ser911 and Thr912, with Ptk2 promoting the double phosphorylation more efficiently than Ptk1 (Guarini et al 2024 ).…”

“…Yeast cells must have the ability to quickly and effectively respond to sudden environmental challenges to ensure their fitness and survival, a trait extensively explored in the yeast model and cell factory Saccharomyces cerevisiae (Estruch 2000 , López-Maury et al 2008 , Saini et al 2018 ). Central to yeast’s stress response is the maintenance of ionic homeostasis, where dissimilarities in ion concentrations across cellular membranes establish crucial gradients for proper cellular function (Orij et al 2011 , Ke et al 2013 , Yenush 2016 , Antunes et al 2023 ). These ionic gradients, which define the transmembrane electrochemical potential, are fundamental for maintaining cellular physiology, solute translocation, and energy homeostasis, preventing toxicity from intracellular surplus ion concentrations (Canadell and Ariño 2016 ).…”

“…The regulation of intracellular pH (pHi) is a paradigmatic example of the critical nature of ion homeostasis in yeast response to acetic acid-induced toxicity, highly dependent on the medium pH (i.e. H + concentration) (Orij et al 2011 , Stratford et al 2013b , Ullah et al 2013b , Antunes et al 2023 ). Low pH values in the medium enhance the effectiveness of microbial growth inhibition by acetic acid (Pampulha and Loureiro-Dias 1989 , Mira et al 2010b , Palma et al 2018 ).…”

“…This effectively counteracts the futile cycle created by the active efflux of acetate and protons if followed by the re-entry of the liposoluble acid form (Van Der Rest et al 1995 , Ullah et al 2013a , Ribeiro et al 2021 ). Additionally, the presence and nature of nutrients (carbon and nitrogen sources, amino acids, and vitamins) and minerals (such as K + , Fe 2+/3+ , and Zn 2+ ) also contributes to yeast adaptation to acetic acid-induced stress (Macpherson et al 2005 , Mira et al 2010b , Wan et al 2015 , Zhang et al 2017 , Xu et al 2019b , Castillo-Plata et al 2022 , Chen et al 2022 , Antunes et al 2023 ).…”