Dataset Information

Identification of A. thaliana 2-Cys peroxiredoxins interacting proteins

ABSTRACT: Peroxiredoxins are ubiquitous thiol-dependent peroxidases for which chaperone and signaling roles have been reported in various types of organisms in the last years. In plants, the peroxidase function of the two typical plastidial 2-Cys peroxiredoxins (2-Cys PRX A and B) has been highlighted while the other functions, particularly in ROS-dependent signal transduction pathways, are still elusive due notably to the lack of knowledge on interacting partners. Using an ex vivo approach based on co-immunoprecipitation of leaf extracts from Arabidopsis thaliana wild-type and mutant plants lacking 2-Cys PRX expression followed by mass spectrometry-based proteomics, 158 proteins were found associated to 2-Cys PRXs, notably already known partners like thioredoxin-related electron donors (Chloroplastic Drought-induced Stress Protein of 32 kDa, Atypical Cysteine Histidine rich Thioredoxin 2) and enzymes involved in chlorophyll synthesis (Protochlorophyllide OxidoReductase B) or carbon metabolisms (Fructose-1,6-BisPhosphatase), validating the relevance of this approach.

INSTRUMENT(S): LTQ Orbitrap Velos

ORGANISM(S): Arabidopsis Thaliana (mouse-ear Cress)

TISSUE(S): Leaf

SUBMITTER: Yohann Couté

LAB HEAD: Christophe Bruley

PROVIDER: PXD003923 | Pride | 2016-10-11

REPOSITORIES: Pride

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Publications

Characterization of the Arabidopsis thaliana 2-Cys peroxiredoxin interactome.

Cerveau Delphine D Kraut Alexandra A Stotz Henrik U HU Mueller Martin J MJ Couté Yohann Y Rey Pascal P

Plant science : an international journal of experimental plant biology 20160710

Peroxiredoxins are ubiquitous thiol-dependent peroxidases for which chaperone and signaling roles have been reported in various types of organisms in recent years. In plants, the peroxidase function of the two typical plastidial 2-Cys peroxiredoxins (2-Cys PRX A and B) has been highlighted while the other functions, particularly in ROS-dependent signaling pathways, are still elusive notably due to the lack of knowledge of interacting partners. Using an ex vivo approach based on co-immunoprecipit ...[more]

PMID: 27717466

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Project description:Cysteine is a rare and conserved amino acid involved in most cellular functions. The thiol group of cysteine can be subjected to diverse oxidative modifications that regulate most physio-pathological states. Here, a Cysteine-specific Phosphonate Adaptable Tag (CysPAT) was synthesized to selectively label cysteine-containing peptides (Cys peptides) followed by enrichment with titanium dioxide (TiO2) and mass spectrometric analysis. The CysPAT strategy was developed using a synthetic peptide, a standard protein and subsequently the strategy was applied to Hela cells lysate, achieving high specificity and enrichment efficiency. In particular,From 500µg of Hela cell lysate, the method led to the identification of 9229 unique Cys peptides starting from 500µg of Hela cell lysate . the CysPAT labelled Cys peptides showed increased hydrophobicity. Finally, the method was further developed to simultaneously enrich Cys peptides and phosphorylated peptidesThe method was further developed to simultaneously enrich Cys peptides and phosphorylated peptides from SILAC labelled Hela cells subjected to 5 min epidermal growth factor (EGF) stimulation. In total, of 22972 unique reversibly modified Cys peptides (3886 proteins) and 18549 unique phosphopeptides (2257 proteins) were simultaneously identified by Orbitrap Fusion. Significant regulation was observed in both phosphorylation and reversible Cys modification of proteins interacted with EGF receptor (EGFR)-binding proteins and proteins involved in EGFR signaling pathway. EGF stimulation can activate the phosphorylation of EGFR and downstream signaling molecules, such as mitogen-activated protein kinases (MAPK1 and MAPK3), meanwhile, it also leads to the activation of multiple protein tyrosine phosphatases (PTPs) by reduction of the catalytic Cys site in the conserved putative phosphatase HC(X)5R motif, Llater the activated PTPs would dephosphorylate and inactivate the molecules activated by EGF stimulation. Overall, the CysPAT strategy is a promising tool for studying redox proteomics and the simultaneous enrichment strategy offers an excellent solution for characterization of cross-talk between phosphorylation and redox induced cysteine modifications.

Dataset Information

Identification of A. thaliana 2-Cys peroxiredoxins interacting proteins

Publications

Characterization of the Arabidopsis thaliana 2-Cys peroxiredoxin interactome.

Similar Datasets

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