Identification of a novel gene (Hsdr4) involved in water-stress tolerance in wild barley

Drought is one of the most severe stresses limiting plant growth and yield. Genes involved in water stress tolerance of wild barley (Hordeum spontaneoum), the progenitor of cultivated barley, were investigated using genotypes contrasting in their response to water stress. Gene expression profiles of water-stress tolerant vs. water-stress sensitive wild barley genotypes, under severe dehydration stress applied at the seedling stage, were compared using cDNA-AFLP analysis. Of the 1100 transcript-derived fragments (TDFs) amplified about 70 displayed differential expression between control and stress conditions. Eleven of them showed clear difference (up- or down-regulation) between tolerant and susceptible genotypes. These TDFs were isolated, sequenced and tested by RT-PCR. The differential expression of seven TDFs was confirmed by RT-PCR, and TDF-4 was selected as a promising candidate gene for water-stress tolerance. The corresponding gene, designated Hsdr4 (Hordeum spontaneum dehydration-responsive), was sequenced and the transcribed and flanking regions were determined. The deduced amino acid sequence has similarity to the rice Rho-GTPase-activating protein-like with a Sec14 p-like lipid-binding domain. Analysis of Hsdr4 promoter region that was isolated by screening a barley BAC library, revealed a new putative miniature inverted-repeat transposable element (MITE), and several potential stress-related binding sites for transcription factors (MYC, MYB, LTRE, and GT-1), suggesting a role of the Hsdr4 gene in plant tolerance to dehydration stress. Furthermore, the Hsdr4 gene was mapped using wild barley mapping population to the long arm of chromosome 3H between markers EBmac541 and EBmag705, within a region that previously was shown to affect osmotic adaptation in barley.