Proteomic identification of glyceraldehyde 3-phosphate dehydrogenase as an inhibitory target of hydrogen peroxide in Arabidopsis

Hydrogen peroxide (H 2O 2) is now recognised as a key signalling molecule in eukaryotes. In plants, H 2O 2 is involved in regulating stomatal closure, gravitropic responses, gene expression and programmed cell death. Although several kinases, such as oxidative signal-inducible 1 (OXI1) kinase and mitogen-activated protein kinases are known to be activated by exogenous H 2O 2, little is known about the proteins that directly react with H 2O 2. Here, we utilised a proteomic approach, using iodoacetamide-based fluorescence tagging of proteins in conjunction with mass spectrometric analysis, to identify several proteins that might be potential targets of H 2O 2 in the cytosolic fraction of Arabidopsis thaliana, the most prominent of which was cytosolic glyceraldehyde 3-phosphate dehydrogenase (cGAPDH; EC 1.2.1.12). cGAPDH from Arabidopsis is inactivated by H 2O 2 in vitro, and this inhibition is reversible by the subsequent addition of reductants such as reduced glutathione (GSH). It has been suggested recently that Arabidopsis GAPDH has roles outside of its catalysis as part of glycolysis, while in other systems this includes that of mediating reactive oxygen species (ROS) signalling. Here, we suggest that cGAPDH in Arabidopsis might also have such a role in mediating ROS signalling in plants.