HKT1 Mediates Sodium Uniport in Roots. Pitfalls in the Expression of HKT1 in Yeast

Abstract

The function of HKT1 in roots is controversial. We tackled this controversy by studying Na⁺ uptake in barley (Hordeum vulgare) roots, cloning the HvHKT1 gene, and expressing the HvHKT1 cDNA in yeast (Saccharomyces cerevisiae) cells. High-affinity Na⁺ uptake was not detected in plants growing at high K⁺ but appeared soon after exposing the plants to a K⁺-free medium. It was a uniport, insensitive to external K⁺ at the beginning of K⁺ starvation and inhibitable by K⁺ several hours later. The expression of HvHKT1 in yeast was Na⁺ (or K⁺) uniport, Na⁺-K⁺ symport, or a mix of both, depending on the construct from which the transporter was expressed. The Na⁺ uniport function was insensitive to external K⁺ and mimicked the Na⁺ uptake carried out by the roots at the beginning of K⁺ starvation. The K⁺ uniport function only took place in yeast cells that were completely K⁺ starved and disappeared when internal K⁺ increased, which makes it unlikely that HvHKT1 mediates K⁺ uptake in roots. Mutation of the first in-frame AUG codon of HvHKT1 to CUC changed the uniport function into symport. The expression of the symport from either mutants or constructs keeping the first in-frame AUG took place only in K⁺-starved cells, while the uniport was expressed in all conditions. We discuss here that the symport occurs only in heterologous expression. It is most likely related to the K⁺ inhibitable Na⁺ uptake process of roots that heterologous systems fail to reproduce.