Apoplastic superoxide level in wheat protoplast under photooxidative stress is regulated by chloroplast redox signals: Effects on the antioxidant system

Apoplastic superoxide radical level in wheat mesophyll protoplasts was evaluated under different light intensities. Conditions that enhanced chloroplastic reactive oxygen species (ROS) production, such as high light intensity, paraquat treatments, or choloplastic Mn-superoxide dismutase (Mn-SOD) overexpression, increased the level of apoplastic superoxide. By contrast, the addition of diphenyl iodonium, a suicide NADPH oxidase inhibitor; DCMU, an herbicide that blocks plastoquinone reduction; and EGTA, a Ca2+ chelator, decreased the superoxide level in the apoplast. Intracellular ROS production was rapidly stimulated by high light intensity and paraquat, and decreased in the presence of DCMU. The association between apoplastic superoxide level and total activities of the antioxidant enzymes superoxide dismutase (SOD) (E.C.:1.15.1.1), glutathione reductase (GR) (E.C.:1.6.4.2), ascobate peroxidase (APX) (E.C.:1.11.1.11), and catalase (CAT) (E.C.:1.11.1.6) was also evaluated. Changes in apoplastic superoxide level were positively correlated with activities of antioxidant enzymes mainly located in the chloroplasts, but not with CAT activity, which is located in peroxisomes and mitochondria. Regulation of apoplastic superoxide level by chloroplastic ROS and plastoquinone redox state is discussed, as well as its role in modulating antioxidant defense responses under photooxidative stress.