Mapping of HKT1;5 Gene in Barley Using GWAS Approach and Its Implication in Salt Tolerance Mechanism

Khaled M Hazzouri, Basel Khraiwesh, Khaled M A Amiri, Duke Pauli, Tom Blake, Mohammad Shahid, Sangeeta K Mullath, David Nelson, Alain L Mansour, Kourosh Salehi-Ashtiani, Michael Purugganan, Khaled Masmoudi

Sodium (Na+) accumulation in the cytosol will result in ion homeostasis imbalance and toxicity of transpiring leaves. Studies of salinity tolerance in the diploid wheat ancestorTriticum monococcumshowed thatHKT1;5-like gene was a major gene in the QTL for salt tolerance, namedNax2. In the present study, we were interested in investigating the molecular mechanisms underpinning the role of theHKT1;5gene in salt tolerance in barley (Hordeum vulgare). A USDA mini-core collection of 2,671 barley lines, part of a field trial was screened for salinity tolerance, and a Genome Wide Association Study (GWAS) was performed. Our results showed important SNPs that are correlated with salt tolerance that mapped to a region whereHKT1;5ion transporter located on chromosome four. Furthermore, sodium (Na+) and potassium (K+) content analysis revealed that tolerant lines accumulate more sodium in roots and leaf sheaths, than in the sensitive ones. In contrast, sodium concentration was reduced in leaf blades of the tolerant lines under salt stress. In the absence of NaCl, the concentration of Na+and K+were the same in the roots, leaf sheaths and leaf blades between the tolerant and the sensitive lines. In order to study the molecular mechanism behind that, alleles of theHKT1;5gene from five tolerant and five sensitive barley lines were cloned and sequenced. Sequence analysis did not show the presence of any polymorphism that distinguishes between the tolerant and sensitive alleles. Our real-time RT-PCR experiments, showed that the expression ofHKT1;5gene in roots of the tolerant line was significantly induced after challenging the plants with salt stress. In contrast, in leaf sheaths the expression was decreased after salt treatment. In sensitive lines, there was no difference in the expression ofHKT1;5gene in leaf sheath under control and saline conditions, while a slight increase in the expression was observed in roots after salt treatment. These results provide stronger evidence thatHKT1;5gene in barley play a key role in withdrawing Na+from the xylem and therefore reducing its transport to leaves. Given all that, these data support the hypothesis thatHKT1;5gene is responsible for Na+unloading to the xylem and controlling its distribution in the shoots, which provide new insight into the understanding of this QTL for salinity tolerance in barley.