Physiological Function Of Nitric Oxide In The Suspension Cultures From Two Ecotypes Of Reed (Phragmites Communis Trin.) Under Drought Stress

Suspension cultures from two ecotypes of reed (phragmites communis Trin.) plant (dune reed and swamp reed), which show different sensitivity to osmotic stress, were used to study plant tolerance to osmotic stress. The protective effects of nitric oxide against oxidative damage and its role during the ABA induced proline accumulation in the suspensions from two ecotypes of reed were investigated. The main results are summarized as follows:Under osmotic stress mediated by polyethylene glycol (PEG-6000), suspension culture of SR showed higher ion leakage, and more oxidative damage to the membrane lipids and proteins were observed compared with the relatively tolerant DR. Treatment with 100μM sodium nitroprusside (SNP), a donor of nitric oxide (NO), markedly alleviated the oxidative damage to the suspension cultures from dune reed and swamp reed caused by PEG. Compared with those of DR and SR suspension cultures under osmotic stress, 100μM SNP treatment significantly enhanced the activities of superoxide dismutase (SOD), catalase (CAT) and ascorbate peroxidase (APX), which may result in the decreased accumulation of H₂O₂ and O₂^- in DR and SR suspension cultures under osmotic stress. The results showed that osmotic stress caused the decrease of reduced glutathione (GSH) in SR suspension cultures, and as a consequence, the contents of oxidized glutathione (GSSG) increased. However, treatment with 100μM SNP dramatically increased GSH content but had no effects on the content of GSSG.. The ratio of GSH/GSSG in DR and SR suspension cultures was elevated in the presence of exogenous NO donor when subjected to osmotic stress, which might be due to the increased activities of glutathione reductase (GR) exerted. Moreover, NO production increased significantly in DR while remained unchanged in SR under osmotic stress. Depletion of endogenous NO with PTIO induced aggravated oxidative damage concomitant with significant decrease in the activities of SOD, CAT and APX in DR suspension culture. Further investigations showed that the NO induced drought tolerance of DR and SR suspension cultures might be due to the activation of plasma membrane H⁺-ATPase and PPase activities. The involvement of Ca²⁺ as intermediate component in NO induced activities of plasma membrane H⁺-ATPase and PPase in DR and SR suspension cultures was also proposed based on the inhibitor experiments under osmotic stress.Proline has been found to accumulate under the adverse environmental conditions, which may be essential for stress tolerance. The content of proline in DR and SR suspension cultures under osmotic stress were increased in the presence of 50 to 100μM SNP treatment, but decreased in the presence of 500μM SNP. The NO mediated proline accumulation could be reversed by the application of PTIO, which indicated the specific role of NO in proline accumulation under osmotic stress. ABA had been shown to play important roles in the tolerance of plants to drought, including the inducement of proline. The results displayed that might function downstream the ABA induced proline accumulation in DR and SR suspension cultures under osmotic stress. The activities ofΔ¹ -pyrroline-5-carboxylate synthetase (P5CS), a key enzyme in proline synthesis, was evevated in the presence of NO and ABA. Further experiments with CaSO₄ and EGTA suggested that Ca²⁺ was involved in mediating the NO signaling pathway leading to proline accumulation under osmotic stress.