Immunolocalization of glutathione biosynthesis enzymes in Arabidopsis thaliana

In plants, glutathione serves as a versatile redox buffer and cellular protective compound against a range of biotic and abiotic stresses. Glutathione production involves glutamate-cysteine ligase (GCL), the redox-regulated limiting enzyme of the pathway, and glutathione synthetase (GS). Because the sub-cellular and sub-organellar localization of these enzymes will have an impact on metabolism, here we examine the localization of GCL and GS in the leaves of Arabidopsis thaliana. Immuno-electron microscopy of leaf cells indicates localization of GCL primarily to the chloroplast with GS found in both the chloroplast and cytosol. Detailed examination of the localization of both enzymes within chloroplasts was performed using fractionation followed by immunoblot analysis and indicates that GCL and GS are found in the stroma. The localization of these enzymes to the stroma of chloroplasts has implications for the redox-regulation of GCL and plant glutathione biosynthesis.