The Hydrophilic C-terminus of Yeast Plasma-membrane Na+/H+ Antiporters Impacts Their Ability to Transport K

Abstract

Yeast plasma-membrane Na⁺/H⁺ antiporters (Nha/Sod) ensure the optimal intracellular level of alkali-metal cations and protons in cells. They are predicted to consist of 13 transmembrane segments (TMSs) and a large hydrophilic C-terminal cytoplasmic part with seven conserved domains. The substrate specificity, specifically the ability to recognize and transport K⁺ cations in addition to Na⁺ and Li⁺, differs among homologs. In this work, we reveal that the composition of the C-terminus impacts the ability of antiporters to transport particular cations. In the osmotolerant yeast Zygosaccharomyces rouxii, the Sod2-22 antiporter only efficiently exports Na⁺ and Li⁺, but not K⁺. The introduction of a negative charge or removal of a positive charge in one of the C-terminal conserved regions (C3) enabled ZrSod2-22 to transport K⁺. The same mutations rescued the low level of activity and purely Li⁺ specificity of ZrSod2-22 with the A179T mutation in TMS6, suggesting a possible interaction between this TMS and the C-terminus. The truncation or replacement of the C-terminal part of ZrSod2-22 with the C-terminus of a K⁺-transporting Nha/Sod antiporter (Saccharomyces cerevisiae Nha1 or Z. rouxii Nha1) also resulted in an antiporter with the capacity to export K⁺. In addition, in ScNha1, the replacement of three positively charged arginine residues 539-541 in the C3 region with alanine caused its inability to provide cells with tolerance to Li⁺. All our results demonstrate that the physiological functions of yeast Nha/Sod antiporters, either in salt tolerance or in K⁺ homeostasis, depend on the composition of their C-terminal parts.

Keywords: C-terminus; cation/H(+) antiport; fungi; potassium homeostasis; salt tolerance.