title: Identification of redox-sensitive cysteines in the Arabidopsis proteome using OxiTRAQ, a quantitative redox proteomics method.
abstract: Cellular redox status plays a key role in mediating various physiological and developmental processes often through modulating activities of redox-sensitive proteins. Various stresses trigger over-production of reactive oxygen/nitrogen species which lead to oxidative modifications of redox-sensitive proteins. Identification and characterization of redox-sensitive proteins are important steps toward understanding molecular mechanisms of stress responses. Here, we report a high-throughput quantitative proteomic approach termed OxiTRAQ for identifying proteins whose thiols undergo reversible oxidative modifications in Arabidopsis cells subjected to oxidative stress. In this approach, a biotinylated thiol-reactive reagent is used for differential labeling of reduced and oxidized thiols. The biotin-tagged peptides are affinity purified, labeled with iTRAQ reagents, and analyzed using a paralleled HCD-CID fragmentation mode in an LTQ-Orbitrap. With this approach, we identified 195 cysteine-containing peptides from 179 proteins whose thiols underwent oxidative modifications in Arabidopsis cells following the treatment with hydrogen peroxide. A majority of those redox-sensitive proteins, including several transcription factors, were not identified by previous redox proteomics studies. This approach allows identification of the specific redox-regulated cysteine residues, and offers an effective tool for elucidation of redox proteomes.
authors.affiliation: Department of Biology, Hong Kong Baptist University, Hong Kong, P. R. China.
authors.forename: Pei
authors.initials: P
authors.lastname: Liu
authors.forename: Huoming
authors.initials: H
authors.lastname: Zhang
authors.forename: Hai
authors.initials: H
authors.lastname: Wang
authors.forename: Yiji
authors.initials: Y
authors.lastname: Xia
mesh_terms: D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000737:chemistry / Q000378:metabolism*; D003545:Cysteine / Q000032:analysis* / Q000378:metabolism; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D010940:Plant Proteins / Q000737:chemistry / Q000378:metabolism*; D020543:Proteome / Q000737:chemistry / Q000378:metabolism*; D040901:Proteomics / Q000379:methods; D013438:Sulfhydryl Compounds / Q000032:analysis* / Q000378:metabolism
chemical_list: D010940:Plant Proteins; D020543:Proteome; D013438:Sulfhydryl Compounds; D003545:Cysteine
keywords: Arabidopsis thaliana; OxiTRAQ; Oxidative stress; Plant proteomics; Redox-sensitive protein; Redoxome; iTRAQ