Populus Euphratica PM Na⁺/H⁺antiport System Mediates Salt-induced H₂O₂Signaling In Arabidopsis

The response of plants to salt stress is through the root cell plasma membrane (PM), meanwhile the salt-induced damage also occurs in the plasma membrane (PM). The plant PM Na+/H+antiport plays a crucial role in controlling K+/Na+homeostasis under salt stress.Our previous microarray analysis indicated that Populus euphratica retained a higher abundance of PM H+-ATPase and PeSOS1transcript versus a salt-sensitive poplar. It has shown that the PM Na+/H+antiport system mediates the plant response to salinity. Moreover, the salt-induced H2O2is of great importance in the control of ionic homeostasis.To clarify the roles of the PM Na+/H+antiport system in salt sensing and adaptation, we isolated the PM H+-ATPase gene PeHA1and Na+/H+antiporter PeSOS1from P. euphratica and introduced it into Arabidopsis thaliana. We investigated the salt-induced alterations of seed germination, root length, biomass, content of K+, Na+, and Ca2+, root fluxes of K+, Na+, and H+, PM ATP hydrolytic activity, proton pumping, and Na+/H+antiport activity in WT and transgenic plants. Results show that transgenic plants were more salt tolerant than WT in term of growth and physiological responses. Compared to wildtype, PeHA1-or PeSOS1-transgenic Arabidopsis had a greater germination rate, root length, and biomass under NaCl stress (50-150mM). Ectopic expression of PeHA1or PeSOSl remarkably enhanced the capacity to control the homeostasis of ions and reactive oxygen species in salinized Arabidopsis. NMT flux data from salinized roots showed that transgenic plants exhibited a more pronounced Na+/H+antiport and less reduction of K+influx versus wildtype. Enhanced PM ATP hydrolytic activity, proton pumping, and Na+/H+antiport in PeHA1-transgenic plants, were consistent to those observed in vivo, i.e., H+extrusion, external acidification, and Na+efflux. In transgenic Arabidopsis roots, H2O2production was higher under control conditions and increased more rapidly than wildtype when plants were subjected to NaCl treatment. Interestingly, transgenic plants were unable to control K+/Na+homeostasis when salt-induced H2O2production was inhibited by diphenylene iodonium, an inhibitor of NADPH oxidase. These observations suggest that PeHA1or PeSOS1accelerates salt tolerance partially through rapid H2O2production upon salt treatment, which triggers adjustments in K+/Na+homeostasis and antioxidant defense in Arabidopsis. It is noting that activities of the antioxidant enzymes ascorbate peroxidase and catalase were typically higher in transgenic seedlings irrespective of salt concentration. This is presumably that the salt-induced H2O2up-regulated the antioxidant system in the Arabidopsis.Accordingly, we conclude that the PM Na+/H+antiport system may function as an ionic stress receptor for the induction of H2O2signaling pathway under salinity conditions. The stimulation of H2O2contributes to K+/Na+homeostasis control through Ca2+-dependent signaling pathway. Moreover, the salt-induced H2O2upregulates the antioxidant defense system over prolonged salinity stress, and thus, the salt-induced ionic and oxidative damage is consequently alleviated.