Characterization of Two HKT1;4 Transporters from Triticum monococcum to Elucidate the Determinants of the Wheat Salt Tolerance Nax1 QTL.

TmHKT1;4-A1 and TmHKT1;4-A2 are two Na+ transporter genes that have been identified as associated with the salt tolerance Nax1 locus found in a durum wheat (Triticum turgidum L. subsp. durum) line issued from a cross with T. monococcum. In the present study, we were interested in getting clues on the molecular mechanisms underpinning this salt tolerance quantitative trait locus (QTL). By analyzing the phylogenetic relationships between wheat and T. monococcum HKT1;4-type genes, we found that durum and bread wheat genomes possess a close homolog of TmHKT1;4-A1, but no functional close homolog of TmHKT1;4-A2. Furthermore, performing real-time reverse transcription-PCR experiments, we showed that TmHKT1;4-A1 and TmHKT1;4-A2 are similarly expressed in the leaves but that TmHKT1;4-A2 is more strongly expressed in the roots, which would enable it to contribute more to the prevention of Na+ transfer to the shoots upon salt stress. We also functionally characterized the TmHKT1;4-A1 and TmHKT1;4-A2 transporters by expressing them in Xenopus oocytes. The two transporters displayed close functional properties (high Na+/K+ selectivity, low affinity for Na+, stimulation by external K+ of Na+ transport), but differed in some quantitative parameters: Na+ affinity was 3-fold lower and the maximal inward conductance was 3-fold higher in TmHKT1;4-A2 than in TmHKT1;4-A1. The conductance of TmHKT1;4-A2 at high Na+ concentration (>10 mM) was also shown to be higher than that of the two durum wheat HKT1;4-type transporters so far characterized. Altogether, these data support the hypothesis that TmHKT1;4-A2 is responsible for the Nax1 trait and provide new insight into the understanding of this QTL.

Comparative functional analysis of two wheat Na(+)/H (+) antiporter SOS1 promoters in Arabidopsis thaliana under various stress conditions.

The bread wheat TaSOS1 has been previously shown to be induced by salt stress treatment. To further investigate the regulation of the TaSOS1 gene, the two genomic fragments Pr SOS1-AB and Pr SOS1-D have been isolated and sequenced. Pr SOS1-AB and Pr SOS1-D are the promoter regions of SOS1 alleles, which are localised on genomes A and/or B, and on genome D, respectively. Sequence analysis of these two promoters revealed the presence of cis-regulatory elements which could be required for abiotic stress and abscisic acid (ABA) responsiveness. Histochemical assays of stably transformed Arabidopsis T3 plants showed that Pr SOS1-AB and Pr SOS1-D are active in this heterologous system, and their activities were almost the same at early developmental stages (4-, 8- and 12-day-old transgenic Arabidopsis seedlings). Nevertheless, ß-glucuronidase (GUS) activity was detected only in plants carrying the Pr SOS1-AB -gusA construct grown for 20 or 30 days. Furthermore, in these plants, the application of abiotic stress produced an accumulation in gusA transcripts. Taken together, these results show that, in this heterologous dicot system and under normal growth conditions, Pr SOS1-AB and Pr SOS1-D are age-dependent and organ-specific promoters. However, in the presence of different stress conditions, the activities of these two promoters became different and only Pr SOS1-AB is an abiotic stress-inducible promoter at different developmental stages. Thus, Pr SOS1-AB can be used for the development of abiotic stress-tolerant transgenic plants.

Functional characterization in Xenopus oocytes of Na+ transport systems from durum wheat reveals diversity among two HKT1;4 transporters.

Plant tolerance to salinity constraint involves complex and integrated functions including control of Na(+) uptake, translocation, and compartmentalization. Several members of the high-affinity K(+) transporter (HKT) family, which comprises plasma-membrane transporters permeable to K(+) and Na(+) or to Na(+) only, have been shown to play major roles in plant Na(+) and K(+) homeostasis. Among them, HKT1;4 has been identified as corresponding to a quantitative trait locus (QTL) of salt tolerance in wheat but was not functionally characterized. Here, we isolated two HKT1;4-type cDNAs from a salt-tolerant durum wheat (Triticum turgidum L. subsp. durum) cultivar, Om Rabia3, and investigated the functional properties of the encoded transporters using a two-electrode voltage-clamp technique, after expression in Xenopus oocytes. Both transporters displayed high selectivity for Na(+), their permeability to other monovalent cations (K(+), Li(+), Cs(+), and Rb(+)) being ten times lower than that to Na(+). Both TdHKT1;4-1 and TdHKT1;4-2 transported Na(+) with low affinity, although the half-saturation of the conductance was observed at a Na(+) concentration four times lower in TdHKT1;4-1 than in TdHKT1;4-2. External K(+) did not inhibit Na(+) transport through these transporters. Quinine slightly inhibited TdHKT1;4-2 but not TdHKT1;4-1. Overall, these data identified TdHKT1;4 transporters as new Na(+)-selective transporters within the HKT family, displaying their own functional features. Furthermore, they showed that important differences in affinity exist among durum wheat HKT1;4 transporters. This suggests that the salt tolerance QTL involving HKT1;4 may be at least in part explained by functional variability among wheat HKT1;4-type transporters.