Regulation of the plant-type 5 '-adenylyl sulfate reductase by oxidative stress

5'-Adenylyl sulfate (APS) reductase (EC 1.8.4.9) catalyzes a key reaction i n the plant sulfate assimilation pathway leading to the synthesis of cystei ne and the antioxidant glutathione. In Arabidopsis thaliana APS reductase i s encoded by a family of three genes. In vitro biochemical studies revealed that the enzyme product derived from one of them (APR1) is activated by ox idation, probably through the formation of a disulfide bond. The APR1 enzym e is 45-fold more active when expressed in a trxB strain of Escherichia col i than in a trxB(+) wild type. The enzyme is inactivated in vitro by treatm ent with disulfide reductants and is reactivated with thiol oxidants. Redox titrations show that the regulation site has a midpoint potential of -330 mV at pH 8.5 and involves a two-electron redox reaction. Exposure of a vari ety of plants to ozone induces a rapid increase in APS reductase activity t hat correlates with the oxidation of the glutathione pool and is followed b y an increase in free cysteine and total glutathione. During the response t o ozone. the level of immunodetectable APS reductase enzyme does not increa se. Treatment of A. thaliana seedlings with oxidized glutathione or paraqua t induces APS reductase activity even when transcription or translation is blocked with inhibitors. The results suggest that a posttranslational mecha nism controls APS reductase. A model is proposed whereby redox regulation o f APS reductase provides a rapidly responding, self-regulating mechanism to control the glutathione synthesis necessary to combat oxidative stress.