Proteomic Analysis of Early-Responsive Redox-Sensitive Proteins in <i>Arabidopsis</i>

Regulation of protein function through oxidative modification has emerged as an important molecular mechanism modulating various biological processes. Here, we report a proteomic study of redox-sensitive proteins in <i>Arabidopsis</i> cells subjected to H₂O₂ treatment. Four gel-based approaches were employed, leading to the identification of four partially overlapping sets of proteins whose thiols underwent oxidative modification in the H₂O₂-treated cells. Using a method based on differential labeling of thiols followed by immunoprecipitation and Western blotting, five of the six selected putative redox-sensitive proteins were confirmed to undergo oxidative modification following the oxidant treatment in <i>Arabidopsis</i> leaves. Another method, which is based on differential labeling of thiols coupled with protein electrophoretic mobility shift assay, was adopted to reveal that one of the H₂O₂-sensitive proteins, a homologue of cytokine-induced apoptosis inhibitor 1 (AtCIAPIN1), also underwent oxidative modification in <i>Arabidopsis</i> leaves after treatments with salicylic acid or the peptide elicitor flg22, two inducers of defense signaling. The redox-sensitive proteins identified from the proteomic study are involved in various biological processes such as metabolism, the antioxidant system, protein biosynthesis and processing, and cytoskeleton organization. The identification of novel redox-sensitive proteins will be helpful toward understanding of cellular components or pathways previously unknown to be redox-regulated