Identification of Novel PAMP-Triggered Phosphorylation and Dephosphorylation Events in Arabidopsis thaliana by Quantitative Phosphoproteomic Analysis
Résumé
Signaling cascades rely strongly on protein kinase-mediated substrate phosphorylation. Currently a major challenge in signal transduction research is to obtain high confidence substrate phosphorylation sites and assign them to specific kinases. In response to bacterial flagellin, a pathogen-associated molecular pattern (PAMP), we searched for rapidly phosphorylated proteins in Arabidopsis thaliana by combining multistage activation (MSA) and electron transfer dissociation (ETD) fragmentation modes, which generate complementary spectra and identify phosphopeptide sites with increased reliability. Of a total of 825 phosphopeptides, we identified 58 to be differentially phosphorylated. These peptides harbor kinase motifs of mitogen-activated protein kinases (MAPKs) and calcium-dependent protein kinases (CDPKs), as well as yet unknown protein kinases. Importantly, 12 of the phosphopeptides show reduced phosphorylation upon flagellin treatment. Since protein abundance levels did not change, these results indicate that flagellin induces not only various protein kinases but also protein phosphatases, even though a scenario of inhibited kinase activity may also be possible.
Mots clés
protein dephosphorylation
priority journal
pathogen associated molecular pattern
nonhuman
molecular dynamics
liquid chromatography
immunoblotting
chemistry
metabolism
molecular genetics
protein kinase
unclassified drug
vegetable protein
phosphoprotein
Arabidopsis thaliana
article
enzyme activity
protein determination
protein motif
protein phosphorylation
Arabidopsis
phosphoprotein phosphatase
phosphopeptide
mitogen activated protein kinase
calcium protein kinase
physiology
Amino Acid Sequence
Chromatography
Liquid
Flagellin
Molecular Sequence Data
Phosphoproteins
Phosphorylation
Proteome
Proteomics
Tandem Mass Spectrometry