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Originally published In Press as doi:10.1074/mcp.M600429-MCP200 on February 21, 2007.
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Molecular & Cellular Proteomics 6:1198-1214, 2007.
© 2007 by The American Society for Biochemistry and Molecular Biology, Inc.

Research

Quantitative Phosphoproteomics of Early Elicitor Signaling in Arabidopsis*,S

Joris J. Benschop{ddagger},§,¶, Shabaz Mohammed§,||, Martina O'Flaherty||, Albert J. R. Heck||, Monique Slijper||,** and Frank L. H. Menke{ddagger},{ddagger}{ddagger}

From the {ddagger} Molecular Genetics, Utrecht University, Padualaan 8, 3584CH Utrecht, The Netherlands and || Department of Biomolecular Mass Spectrometry, Bijvoet Center for Biomolecular Research and Utrecht Institute for Pharmaceutical Sciences, Utrecht University, Sorbonnelaan 16, 3584CA Utrecht, The Netherlands

Perception of general elicitors by plant cells initiates signal transduction cascades that are regulated by protein phosphorylation. The earliest signaling events occur within minutes and include ion fluxes across the plasma membrane, activation of MAPKs, and the formation of reactive oxygen species. The phosphorylation events that regulate these signaling cascades are largely unknown. Here we present a mass spectrometry-based quantitative phosphoproteomics approach that identified differentially phosphorylated sites in signaling and response proteins from Arabidopsis cells treated with either flg22 or xylanase. Our approach was sensitive enough to quantitate phosphorylation on low abundance signaling proteins such as calcium-dependent protein kinases and receptor-like kinase family members. With this approach we identified one or more differentially phosphorylated sites in 76 membrane-associated proteins including a number of defense-related proteins. Our data on phosphorylation indicate a high degree of complexity at the level of post-translational modification as exemplified by the complex modification patterns of respiratory burst oxidase protein D. Furthermore the data also suggest that protein translocation and vesicle traffic are important aspects of early signaling and defense in response to general elicitors. Our study presents the largest quantitative Arabidopsis phosphoproteomics data set to date and provides a new resource that can be used to gain novel insight into plant defense signal transduction and early defense response.

** To whom correspondence may be addressed. E-mail: m.slijper{at}pharm.uu.nl

{ddagger}{ddagger} To whom correspondence may be addressed. E-mail: f.l.h.menke{at}bio.uu.nl


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