Assessing the role of root plasma membrane and tonoplast Na⁺/H⁺ exchangers in salinity tolerance in wheat: in planta quantification methods

Abstract

This work investigates the role of cytosolic Na⁺ exclusion in roots as a means of salinity tolerance in wheat, and offers in planta methods for the functional assessment of major transporters contributing to this trait. An electrophysiological protocol was developed to quantify the activity of plasma membrane Na⁺ efflux systems in roots, using the microelectrode ion flux estimation (MIFE) technique. We show that active efflux of Na⁺ from wheat root epidermal cells is mediated by a SOS1-like homolog, energized by the plasma membrane H⁺-ATPase. SOS1-like efflux activity was highest in Kharchia 65, a salt-tolerant bread wheat cultivar. Kharchia 65 also had an enhanced ability to sequester large quantities of Na⁺ into the vacuoles of root cells, as revealed by confocal microscopy using Sodium Green. These findings were consistent with the highest level of expression of both SOS1 and NHX1 transcripts in plant roots in this variety. In the sensitive wheat varieties, a greater proportion of Na⁺ was located in the root cell cytosol. Overall, our findings suggest a critical role of cytosolic Na⁺ exclusion for salinity tolerance in wheat and offer convenient protocols to quantify the contribution of the major transporters conferring this trait, to screen plants for salinity tolerance.