Absolute protein quantification by LC/MS(E) for global analysis of salicylic acid-induced plant protein secretion responses

• Published in: Journal of proteome research 2009-01, Vol.8 (1), p.82-93
• Main Authors: Cheng, Fang-yi, Blackburn, Kevin, Lin, Yu-min, Goshe, Micheal B, Williamson, John D
• Format: Article
• Language: English
• Subjects: Arabidopsis - drug effects; Arabidopsis - metabolism; Chromatography, Liquid - methods; Computational Biology; Mass Spectrometry - methods; Plant Physiological Phenomena; Plant Proteins - metabolism; Plant Proteins - secretion; Proteins - secretion; Proteome; Proteomics - methods; Salicylic Acid - metabolism; Salicylic Acid - pharmacology; Time Factors
• ISSN: 1535-3893
• DOI: 10.1021/pr800649s

The plant cell wall is a dynamic cellular compartment consisting of a complex matrix of components that can change dramatically in response to environmental stresses. During pathogen attack, for instance, a wide spectrum of proteins that participate in various sequential processes involved in plant defense is secreted into the cell wall. In this study, a mass spectrometry, data-independent acquisition approach known as LC/MS (E) was used to assess temporal changes in the cell wall proteome in response to different levels of an endogenous inducer of plant disease defense responses, salicylic acid (SA). LC/MS (E) was used as a label-free method that enabled simultaneous protein identification and absolute femtomole quantification of each protein secreted into the extracellular matrix. A total of 74 secreted proteins were identified, 63 of which showed increased specific secretion in response to SA. A majority of this induced secretion occurred within 2 h of treatment, indicating that many proteins are involved in the early stages of plant defenses. We also identified a number of apparently nonclassically secreted proteins, suggesting that, as in many nonplant systems, Golgi/ER-independent mechanisms exist for plant protein secretion. These results provide new insight into plant apoplastic defense mechanisms and demonstrate that LC/MS (E) is a powerful tool for obtaining both relative and absolute proteome-scale quantification that can be applied to complex, time- and dose-dependent experimental designs.