Hydrogen peroxide produced by NADPH oxidases increases proline accumulation during salt or mannitol stress in Arabidopsis thaliana

Abstract

Summary: Many plants accumulate proline, a compatible osmolyte, in response to various environmental stresses such as water deficit and salinity. In some stress responses, plants generate hydrogen peroxide (H 2 O 2) that mediates numerous physiological and biochemical processes. The aim was to study the relationship between stress‐induced proline accumulation and H 2 O 2 production. Using pharmacological and reverse genetic approaches in Arabidopsis thaliana, we investigated the role of NADPH oxidases, Respiratory burst oxidase homologues (Rboh), in the induction of proline accumulation was investigated in response to stress induced by either 200 mM NaCl or 400 mM mannitol. Stress from NaCl or mannitol resulted in a transient increase in H 2 O 2 content accompanied by accumulation of proline. Dimethylthiourea, a scavenger of H 2 O 2, and diphenylene iodonium (DPI), an inhibitor of H 2 O 2 production by NADPH oxidase, were found to significantly inhibit proline accumulation in these stress conditions. DPI also reduced the expression level of Δ 1‐pyrroline‐5‐carboxylate synthetase, the key enzyme involved in the biosynthesis of proline. Similarly, less proline accumulated in knockout mutants lacking either AtRbohD or AtRbohF than in wild‐type plants in response to the same stresses. Our data demonstrate that AtRbohs (A. thaliana Rbohs) contribute to H 2 O 2 production in response to NaCl or mannitol stress to increase proline accumulation in this plant.