Suppression of Arabidopsis vesicle-SNARE expression inhibited fusion of H2O2-containing vesicles with tonoplast and increased salt tolerance.

Abstract

Intracellular vesicle trafficking performs essential functions in eukaryotic cells, such as membrane trafficking and delivery of molecules to their destinations. A major endocytotic route in plants is vesicle trafficking to the vacuole that plays an important role in plant salt tolerance. The final step in this pathway is mediated by the AtVAMP7C family of vesicle soluble N-ethylmaleimide-sensitive factor attachment protein receptors (v-SNAREs) that carry out the vesicle fusion with the tonoplast. Exposure to high-salt conditions causes immediate ionic and osmotic stresses, followed by production of reactive oxygen species. Here, we show that the reactive oxygen species are produced intracellularly, in endosomes that were targeted to the central vacuole. Suppression of the AtVAMP7C genes expression by antisense AtVAMP711 gene or in mutants of this family inhibited fusion of H2O2-containing vesicles with the tonoplast, which resulted in formation of H2O2-containing megavesicles that remained in the cytoplasm. The antisense and mutant plants exhibited improved vacuolar functions, such as maintenance of DeltapH, reduced release of calcium from the vacuole, and greatly improved plant salt tolerance. The antisense plants exhibited increased calcium-dependent protein kinase activity upon salt stress. Improved vacuolar ATPase activity during oxidative stress also was observed in a yeast system, in a DeltaVamp7 knockout strain. Interestingly, a microarray-based analysis of the AtVAMP7C genes showed a strong down-regulation of most genes in wild-type roots during salt stress, suggesting an evolutionary molecular adaptation of the vacuolar trafficking.