Cytosolic Ascorbate Peroxidase 1 Is a Central Component of the Reactive Oxygen Gene Network of Arabidopsis

TLDR

Reactive oxygen species are central to plant metabolism and defense, with chloroplasts being major producers and scavengers, yet the role of cytosolic ROS‑scavenging mechanisms remains largely unknown. The study aims to identify proteins oxidized in APX1‑deficient Arabidopsis and characterize the signaling that follows under moderate light stress. Using a dominant‑negative approach and RbohD knockout plants, the authors show that heat shock transcription factors sense H2O2 stress and that RbohD may amplify ROS signaling during light stress. Loss of cytosolic APX1 collapses the chloroplastic H2O2‑scavenging system, increases H2O2 and protein oxidation, and demonstrates that cytosolic APX1 protects chloroplasts during light stress through cross‑compartment protection of thylakoid and stromal/mitochondrial APXs.

Abstract

Reactive oxygen species (ROS), such as O2- and H2O2, play a key role in plant metabolism, cellular signaling, and defense. In leaf cells, the chloroplast is considered to be a focal point of ROS metabolism. It is a major producer of O2- and H2O2 during photosynthesis, and it contains a large array of ROS-scavenging mechanisms that have been extensively studied. By contrast, the function of the cytosolic ROS-scavenging mechanisms of leaf cells is largely unknown. In this study, we demonstrate that in the absence of the cytosolic H2O2-scavenging enzyme ascorbate peroxidase 1 (APX1), the entire chloroplastic H2O2-scavenging system of Arabidopsis thaliana collapses, H2O2 levels increase, and protein oxidation occurs. We further identify specific proteins oxidized in APX1-deficient plants and characterize the signaling events that ensue in knockout-Apx1 plants in response to a moderate level of light stress. Using a dominant-negative approach, we demonstrate that heat shock transcription factors play a central role in the early sensing of H2O2 stress in plants. Using knockout plants for the NADPH oxidase D protein (knockout-RbohD), we demonstrate that RbohD might be required for ROS signal amplification during light stress. Our study points to a key role for the cytosol in protecting the chloroplast during light stress and provides evidence for cross-compartment protection of thylakoid and stromal/mitochondrial APXs by cytosolic APX1.