In the model yeast Saccharomyces cerevisiae, Trk1 is the main K+ importer. It is involved in many important physiological processes, such as the maintenance of ion homeostasis, cell volume, intracellular pH, and plasma-membrane potential. The ScTrk1 protein can be of great interest to industry, as it was shown that changes in its activity influence ethanol production and tolerance in S. cerevisiae and also cell performance in the presence of organic acids or high ammonium under low K+ conditions. Nonconventional yeast species are attracting attention due to their unique properties and as a potential source of genes that encode proteins with unusual characteristics. In this work, we aimed to study and compare Trk proteins from Debaryomyces hansenii, Hortaea werneckii, Kluyveromyces marxianus, and Yarrowia lipolytica, four biotechnologically relevant yeasts that tolerate various extreme environments. Heterologous expression in S. cerevisiae cells lacking the endogenous Trk importers revealed differences in the studied Trk proteins' abilities to support the growth of cells under various cultivation conditions such as low K+ or the presence of toxic cations, to reduce plasma-membrane potential or to take up Rb+ . Examination of the potential of Trks to support the stress resistance of S. cerevisiae wild-type strains showed that Y. lipolytica Trk1 is a promising tool for improving cell tolerance to both low K+ and high salt and that the overproduction of S. cerevisiae's own Trk1 was the most efficient at improving the growth of cells in the presence of highly toxic Li+ ions.

Department of Agricultural Chemistry Edaphology and Microbiology University of Córdoba Córdoba Spain

Laboratory of Membrane Transport Institute of Physiology of the Czech Academy of Sciences Prague 4 Czech Republic

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Aggarwal, M., & Mondal, A. K. (2006). Role of N-terminal hydrophobic region in modulating the subcellular localization and enzyme activity of the bisphosphate nucleotidase from Debaryomyces hansenii. Eukaryotic Cell, 5(2), 262-271.

Ariño, J., Ramos, J., & Sychrova, H. (2019). Monovalent cation transporters at the plasma membrane in yeasts. Yeast, 36(4), 177-193.

Ariño, J., Ramos, J., & Sychrová, H. (2010). Alkali metal cation transport and homeostasis in yeasts. Microbiology and Molecular Biology Reviews, 74(1), 95-120.

Benito, B., Garciadeblas, B., & Rodriguez-Navarro, A. (2012). HAK transporters from Physcomitrella patens and Yarrowia lipolytica mediate sodium uptake. Plant & Cell Physiology, 53(6), 1117-1123.

Bertl, A., Ramos, J., Ludwig, J., Lichtenberg-Fraté, H., Reid, J., Bihler, H., Calero, F., Martínez, P., & Ljungdahl, P. O. (2003). Characterization of potassium transport in wild-type and isogenic yeast strains carrying all combinations of trk1, trk2 and tok1 null mutations. Molecular Microbiology, 47(3), 767-780.

Borovikova, D., Herynkova, P., Rapoport, A., & Sychrova, H. (2014). Potassium uptake system Trk2 is crucial for yeast cell viability during anhydrobiosis. FEMS Microbiology Letters, 350(1), 28-33.

Chao, H., Yen, Y., & Ku, M. S. (2009). Characterization of a salt-induced DhAHP, a gene coding for alkyl hydroperoxide reductase, from the extremely halophilic yeast Debaryomyces hansenii. BMC Microbiology, 9, 182.

Dibalova-Culakova, H., Alonso-del-Real, J., Querol, A., & Sychrova, H. (2018). Expression of heterologous transporters in Saccharomyces kudriavzevii: A strategy for improving yeast salt tolerance and fermentation performance. International Journal of Food Microbiology, 268, 27-34.

Dujon, B., Sherman, D., Fischer, G., Durrens, P., Casaregola, S., Lafontaine, I., De Montigny, J., Marck, C., Neuvéglise, C., Talla, E., Goffard, N., Frangeul, L., Aigle, M., Anthouard, V., Babour, A., Barbe, V., Barnay, S., Blanchin, S., Beckerich, J. M., … Souciet, J. L. (2004). Genome evolution in yeasts. Nature, 430(6995), 35-44.

Dušková, M., Cmunt, D., Papoušková, K., Masaryk, J., & Sychrová, H. (2021). Minority potassium-uptake system Trk2 has a crucial role in yeast survival of glucose-induced cell death. Microbiology, 167(6), 001065.

Elicharova, H., Herynkova, P., Zimmermannova, O., & Sychrova, H. (2019). Potassium uptake systems of Candida krusei. Yeast, 36(7), 439-448.

Elicharová, H., Hušeková, B., & Sychrová, H. (2016). Three Candida albicans potassium uptake systems differ in their ability to provide Saccharomyces cerevisiae trk1trk2 mutants with necessary potassium. FEMS Yeast Research, 16(4), fow039.

Gaber, R. F., Styles, C. A., & Fink, G. R. (1988). TRK1 encodes a plasma membrane protein required for high-affinity potassium transport in Saccharomyces cerevisiae. Molecular and Cellular Biology, 8(7), 2848-2859.

Gnügge, R., & Rudolf, F. (2017). Saccharomyces cerevisiae Shuttle vectors. Yeast, 34(5), 205-221.

Gorjan, A., & Plemenitaš, A. (2006). Identification and characterization of ENA ATPases HwENA1 and HwENA2 from the halophilic black yeast Hortaea werneckii. FEMS Microbiology Letters, 265(1), 41-50.

Gunde-Cimerman, N., Zalar, P., Hoog, S., Plemenitaš, A., & Gunde-Cimerman, N. (2000). Hypersaline waters in salterns-Natural ecological niches for halophilic black yeasts. FEMS Microbiology Ecology, 32(3), 235-240.

Hanscho, M., Ruckerbauer, D. E., Chauhan, N., Hofbauer, H. F., Krahulec, S., Nidetzky, B., Kohlwein, S. D., Zanghellini, J., & Natter, K. (2012). Nutritional requirements of the BY series of Saccharomyces cerevisiae strains for optimum growth. FEMS Yeast Research, 12(7), 796-808.

Herrera, R., Salazar, A., Ramos-Moreno, L., Ruiz-Roldan, C., & Ramos, J. (2017). Vacuolar control of subcellular cation distribution is a key parameter in the adaptation of Debaryomyces hansenii to high salt concentrations. Fungal Genetics and Biology, 100, 52-60.

Hess, D. C., Lu, W., Rabinowitz, J. D., & Botstein, D. (2006). Ammonium toxicity and potassium limitation in yeast. PLoS Biology, 4(11), e351.

Holland, S. L., Reader, T., Dyer, P. S., & Avery, S. V. (2014). Phenotypic heterogeneity is a selected trait in natural yeast populations subject to environmental stress. Environmental Microbiology, 16(6), 1729-1740.

Illarionov, A., Lahtvee, P. J., & Kumar, R. (2021). Potassium and sodium salt stress characterization in the yeasts Saccharomyces cerevisiae, Kluyveromyces marxianus, and Rhodotorula toruloides. Applied and Environmental Microbiology, 87(13), e0310020.

Inokuma, K., Ishii, J., Hara, K. Y., Mochizuki, M., Hasunuma, T., & Kondo, A. (2015). Complete genome sequence of Kluyveromyces marxianus NBRC1777, a nonconventional thermotolerant yeast. Genome Announcements, 3(2), e00389-15.

Kale, D., Spurny, P., Shamayeva, K., Spurna, K., Kahoun, D., Ganser, D., Zayats, V., & Ludwig, J. (2019). The S. cerevisiae cation translocation protein Trk1 is functional without its “long hydrophilic loop” but LHL regulates cation translocation activity and selectivity. Biochimica et Biophysica Acta (BBA)-Biomembranes, 1861(8), 1476-1488.

Karim, A., Gerliani, N., & Aïder, M. (2020). Kluyveromyces marxianus: An emerging yeast cell factory for applications in food and biotechnology. International Journal of Food Microbiology, 333, 108818.

Kinclová, O., Potier, S., & Sychrová, H. (2001). The Zygosaccharomyces rouxii strain CBS732 contains only one copy of the HOG1 and the SOD2 genes. Journal of Biotechnology, 88(2), 151-158.

Ko, C. H., Gaber, R. F., Ko, C. H., & Gaber, R. F. (1991). TRK1 and TRK2 encode structurally related K+ transporters in Saccharomyces cerevisiae. Molecular and Cellular Biology, 11(8), 4266-4273.

Kodedova, M., & Sychrova, H. (2015). Changes in the sterol composition of the plasma membrane affect membrane potential, salt tolerance and the activity of multidrug resistance pumps in Saccharomyces cerevisiae. PLoS One, 10(9), e0139306.

Kogej, T., Ramos, J., Plemenitaš, A., & Gunde-Cimerman, N. (2005). The halophilic fungus Hortaea werneckii and the halotolerant fungus Aureobasidium pullulans maintain low intracellular cation concentrations in hypersaline environments. Applied and Environmental Microbiology, 71(11), 6600-6605.

Lam, F. H., Ghaderi, A., Fink, G. R., & Stephanopoulos, G. (2014). Biofuels. Science, 346(6205), 71-75.

Lane, M. M., & Morrissey, J. P. (2010). Kluyveromyces marxianus: A yeast emerging from its sister's shadow. Fungal Biology Reviews, 24(1-2), 17-26.

Lenassi, M., Gostinčar, C., Jackman, S., Turk, M., Sadowski, I., Nislow, C., Jones, S., Birol, I., Cimerman, N. G., & Plemenitaš, A. (2013). Whole genome duplication and enrichment of metal cation transporters revealed by de novo genome sequencing of extremely halotolerant black yeast Hortaea werneckii. PLoS One, 8(1), e71328.

Magnan, C., Yu, J., Chang, I., Jahn, E., Kanomata, Y., Wu, J., Zeller, M., Oakes, M., Baldi, P., & Sandmeyer, S. (2016). Sequence assembly of Yarrowia lipolytica strain W29/CLIB89 shows transposable element diversity. PLoS One, 11(9), e0162363.

Mamaev, D., & Zvyagilskaya, R. (2021). Yarrowia lipolytica: a multitalented yeast species of ecological significance. FEMS Yeast Research, 21(2), foab008.

Marešová, L., & Sychrová, H. (2007). Applications of a microplate reader in yeast physiology research. Biotechniques, 43(5), 667-672.

Martínez, J. L., Sychrova, H., & Ramos, J. (2011). Monovalent cations regulate expression and activity of the Hak1 potassium transporter in Debaryomyces hansenii. Fungal Genetics and Biology, 48(2), 177-184.

Masaryk, J., & Sychrová, H. (2022). Yeast Trk1 potassium transporter gradually changes its affinity in response to both external and internal signals. Journal of Fungi, 8(5), 432.

Michel, B., Lozano, C., Rodríguez, M., Coria, R., Ramírez, J., & Peña, A. (2006). The yeast potassium transporter TRK2 is able to substitute for TRK1 in its biological function under low K and low pH conditions. Yeast, 23(8), 581-589.

Miller, K. K., & Alper, H. S. (2019). Yarrowia lipolytica: more than an oleaginous workhorse. Applied Microbiology and Biotechnology, 103(23-24), 9251-9262.

Navarrete, C., Estrada, M., & Martínez, J. L. (2022). Debaryomyces hansenii: an old acquaintance for a fresh start in the era of the green biotechnology. World Journal of Microbiology & Biotechnology, 38(6), 99.

Navarrete, C., Petrezsélyová, S., Barreto, L., Martínez, J. L., Zahrádka, J., Ariño, J., Sychrová, H., & Ramos, J. (2010). Lack of main K+ uptake systems in Saccharomyces cerevisiae cells affects yeast performance in both potassium-sufficient and potassium-limiting conditions: Potassium transport and yeast physiology. FEMS Yeast Research, 10(5), 508-517.

Papouskova, K., & Sychrova, H. (2006). Yarrowia lipolytica possesses two plasma membrane alkali metal cation/H+ antiporters with different functions in cell physiology. FEBS Letters, 580(8), 1971-1976.

Papoušková, K., & Sychrová, H. (2007). Production of Yarrowia lipolytica Nha2 Na+/H+ antiporter improves the salt tolerance of Saccharomyces cerevisiae. Folia Microbiologica, 52(6), 600-602.

Petrezsélyová, S., Ramos, J., & Sychrová, H. (2011). Trk2 transporter is a relevant player in K+ supply and plasma-membrane potential control in Saccharomyces cerevisiae. Folia Microbiologica, 56(1), 23-28.

Petrezselyova, S., Zahradka, J., & Sychrova, H. (2010). Saccharomyces cerevisiae BY4741 and W303-1A laboratory strains differ in salt tolerance. Fungal Biology, 114(2-3), 144-150.

Prista, C., González-Hernández, J. C., Ramos, J., & Loureiro-Dias, M. C. (2007). Cloning and characterization of two K+ transporters of Debaryomyces hansenii. Microbiology, 153(Pt9), 3034-3043.

Prista, C., Michán, C., Miranda, I. M., & Ramos, J. (2016). The halotolerant Debaryomyces hansenii, the Cinderella of non-conventional yeasts. Yeast, 33(10), 523-533.

Ramos, J., Ariño, J., & Sychrová, H. (2011). Alkali-metal-cation influx and efflux systems in nonconventional yeast species. FEMS Microbiology Letters, 317(1), 1-8.

Reisser, C., Dick, C., Kruglyak, L., Botstein, D., Schacherer, J., & Hess, D. C. (2013). Genetic basis of ammonium toxicity resistance in a sake strain of yeast: A mendelian case. G3: Genes|Genomes|Genetics, 3(4), 733-740.

Rodríguez-Navarro, A., & Ramos, J. (1984). Dual system for potassium transport in Saccharomyces cerevisiae. Journal of Bacteriology, 159(3), 940-945.

Rodrı́guez-Navarro, A. (2000). Potassium transport in fungi and plants. Biochimica et Biophysica Acta (BBA)-Reviews on Biomembranes, 1469(1), 1-30.

Rouzeau, C., Dagkesamanskaya, A., Langer, K., Bibette, J., Baudry, J., Pompon, D., & Anton-Leberre, V. (2018). Adaptive response of yeast cells to triggered toxicity of phosphoribulokinase. Research in Microbiology, 169(6), 335-342.

Stříbný, J., Kinclová-Zimmermannová, O., & Sychrová, H. (2012). Potassium supply and homeostasis in the osmotolerant non-conventional yeasts Zygosaccharomyces rouxii differ from Saccharomyces cerevisiae. Current Genetics, 58(5-6), 255-264.

Wallis, J. W., Chrebet, G., Brodsky, G., Rolfe, M., & Rothstein, R. (1989). A hyper-recombination mutation in S. cerevisiae identifies a novel eukaryotic topoisomerase. Cell, 58(2), 409-419.

Xu, X., Williams, T. C., Divne, C., Pretorius, I. S., & Paulsen, I. T. (2019). Evolutionary engineering in Saccharomyces cerevisiae reveals a TRK1-dependent potassium influx mechanism for propionic acid tolerance. Biotechnology for Biofuels, 12, 97.

Zayats, V., Stockner, T., Pandey, S. K., Wörz, K., Ettrich, R., & Ludwig, J. (2015). A refined atomic scale model of the Saccharomyces cerevisiae K+-translocation protein Trk1p combined with experimental evidence confirms the role of selectivity filter glycines and other key residues. Biochimica et Biophysica Acta (BBA)-Biomembranes, 1848(5), 1183-1195.

Zimmermannová, O., Felcmanová, K., Rosas-Santiago, P., Papoušková, K., Pantoja, O., & Sychrová, H. (2019). Erv14 cargo receptor participates in regulation of plasma-membrane potential, intracellular pH and potassium homeostasis via its interaction with K+-specific transporters Trk1 and Tok1. Biochimica et Biophysica Acta (BBA)-Molecular Cell Research, 1866(9), 1376-1388.

Zimmermannova, O., Salazar, A., Sychrova, H., & Ramos, J. (2015). Zygosaccharomyces rouxii Trk1 is an efficient potassium transporter providing yeast cells with high lithium tolerance. FEMS Yeast Research, 15(4), fov029.

The superior growth of Kluyveromyces marxianus at very low potassium concentrations is enabled by the high-affinity potassium transporter Hak1