Posttranslational Modification of 2-CysPrxs by Reduced Glutathione Interplays with CYP20-3-Dependent OPDA Signaling Pathways

Abstract

Glutathione, a tripeptide (GSH; a 307 Da γ-L-glutamyl-L-cysteinyl-glycine peptide) is the most abundantly present, thiol in plants and animals, and considered as a master antioxidant. It prevents damage to cells caused by reducing reactive oxygen species (ROS) and other peroxides. It is capable of donating electron (H+, e-) to ROS and is itself oxidized to glutathione disulphide (GSSG). Reduced GSH, beside GSSG, can directly crosslink with the sulfhydryl group of protein cysteine residues (e.g. 2-cysteine peroxiredoxins, 2CPs), hereafter referred to as ‘GSH-glutathionylation’. Arabidopsis contains 2 isoforms of 2CPs, called A and B, that are 96.5 % identical in amino acid sequences. There are only 7 amino acid differences between them and, because of this similarity, most studies have considered them as functionally redundant enzymes. However, we observed that GSH, as a posttranslational modifier, is able to form mixed disulfide bond with, and differentially regulates 2CPs; monomerizing and enhancing peroxidase activity of 2CPA, whereas decamerizing and showing chaperone activity of 2CPB. Plants constantly encounter a wide range of environmental stresses, and thus have developed efficient defense mechanisms to overcome them. (+)-12-oxo-phytodienoic acid (OPDA) is an important biologically active jasmonates, able to trigger and cause signaling pathway on its own and control the expression of selective genes, as well as activate plant defense and growth processes. Here we assessed if (1) OPDA signaling enhances an electron transfer (govern energy allocation) from thioredoxins (TRXs, photosystem) to cyclophilin 20-3 (CYP20-3, OPDA receptor) in plastid sulfur assimilation (defense activations), and (2) OPDA signaling stimulates CYP20-3-mediated reduction of 2CPs. Our in vitro and ex vivo protein analyses illuminated that CYP20-3 relays an OPDA signal during stress responsive regulation of cellular redox homeostasis. OPDA signaling stimulates an electron transfer from TRX to CYP20-3. Once activated, CYP20-3 target-reduces (deglutathionylates) and inactivates 2CPAGS, suppressing the peroxide detoxification in photosynthesis.