Dehydration inhibits petal expansion resulting in abnormal flower opening and results in quality loss during the marketing of cut flowers. We constructed a suppression subtractive hybridization library from rose ( Rosa hybrida ) flowers containing 3,513 unique expressed sequence tags and analyzed their expression profiles during cycles of dehydration. We found that 54 genes were up-regulated by the first dehydration, restored or even down-regulated by rehydration, and once again up-regulated by the second dehydration. Among them, we identified a putative NAC family transcription factor ( RhNAC2 ). With transactivation activity of its carboxyl-terminal domain in yeast ( Saccharomyces cerevisiae ) cell and Arabidopsis ( Arabidopsis thaliana ) protoplast, RhNAC2 belongs to the NAC transcription factor clade related to plant development in Arabidopsis. A putative expansin gene named RhEXPA4 was also dramatically up-regulated by dehydration. Silencing RhNAC2 or RhEXPA4 in rose petals by virus-induced gene silencing significantly decreased the recovery of intact petals and petal discs during rehydration. Overexpression of RhNAC2 or RhEXPA4 in Arabidopsis conferred strong drought tolerance in the transgenic plants. RhEXPA4 expression was repressed in RhNAC2- silenced rose petals, and the amino-terminal binding domain of RhNAC2 bound to the RhEXPA4 promoter. Twenty cell wall-related genes, including seven expansin family members, were up-regulated in Arabidopsis plants overexpressing RhNAC2 . These data indicate that RhNAC2 and RhEXPA4 are involved in the regulation of dehydration tolerance during the expansion of rose petals and that RhEXPA4 expression may be regulated by RhNAC2. Glossary WDS water deficit stress GO Gene Ontology SSH suppression subtractive hybridization RT reverse transcription UTR untranslated region SD synthetic dextrose VIGS virus-induced gene silencing ABA abscisic acid EMSA electrophoresis mobility shift assay GST glutathione S -transferase MS Murashige and Skoog TAIR The Arabidopsis Information Resource cDNA complementary DNA uniEST unique EST TRV tobacco rattle virus
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