Survey of Barley Sodium Transporter HvHKT1;1 Variants and Their Functional Analysis

Barley (Hordeum vulgare L.) employs the Na+ transporter HvHKT1;1, which is an N+-selective transporter. This study characterized the full-length HvHKT1;1 (HvHKT1;1-FL) and three mRNA variants (HvHKT1;1-V1, -V2, and -V3), which encode polypeptides of 64.7, 54.0, 40.5, and 32.9 kDa, respectively. Tissue-specific expression profiling revealed that HvHKT1;1-FL is the most abundant transcript across leaf, sheath, and root tissues under normal conditions, with the highest expression in leaves. Under 150 mM NaCl stress, HvHKT1;1-FL and its variants showed a dynamic, time-dependent expression pattern, with peak leaf expression at 2 h, sheath expression at 12 h, and root expression at 2 h, suggesting their roles in early stress response. Functional analysis using two-electrode voltage-clamp measurements demonstrated that HvHKT1;1-FL is highly selective for Na+, with minimal conductance for K+, Li+, Rb+, or Cs+. It demonstrated high Na+ transport efficiency, characterized by higher Vmax and lower Km values, while the variants showed reduced Na+ currents, lower Vmax, and higher Km values, indicating decreased Na+ transport capacity. Reversal potential analyses further confirmed Na+ selectivity, with HvHKT1;1-FL displaying the strongest preference for Na+. Notably, while all variants retained Na+ selectivity, they showed reduced efficiency, as indicated by a more negative reversal potential in low Na+ conditions. These findings highlight the functional diversity among HvHKT1;1 variants, with HvHKT1;1-FL playing a dominant role in Na+ transport. The tissue-specific regulation of these variants under salinity stress underscores their importance in barley’s adaptive responses.