Quantitative Proteomics Reveals Dynamic Changes in the Plasma Membrane During Arabidopsis Immune Signaling

James Mitch Elmore,Gitta Coaker,Jun Liu,Brett Phinney,Barrett Smith

doi:10.1074/mcp.m111.014555

Abstract

The plant plasma membrane is a crucial mediator of the interaction between plants and microbes. Understanding how the plasma membrane proteome responds to diverse immune signaling events will lead to a greater understanding of plant immunity and uncover novel targets for crop improvement. Here we report the results from a large scale quantitative proteomics study of plasma membrane-enriched fractions upon activation of the Arabidopsis thaliana immune receptor RPS2. More than 2300 proteins were identified in total, with 1353 proteins reproducibly identified across multiple replications. Label-free spectral counting was employed to quantify the relative protein abundance between different treatment samples. Over 20% of up-regulated proteins have known roles in plant immune responses. Significantly changing proteins include those involved in calcium and lipid signaling, membrane transport, primary and secondary metabolism, protein phosphorylation, redox homeostasis, and vesicle trafficking. A subset of differentially regulated proteins was independently validated during bacterial infection. This study presents the largest quantitative proteomics data set of plant immunity to date and provides a framework for understanding global plasma membrane proteome dynamics during plant immune responses.

Highlights

Because many classes of plant pathogens remain outside the host cell membrane during infection, the plant plasma membrane (PM)1 controls numerous facets of plant-microbe interactions
1 The abbreviations used are: PM, plasma membrane; effector-triggered immunity (ETI), effectortriggered immunity; HR, hypersensitive response; pathogen-associated molecular pattern (PAMP), pathogenassociated molecular pattern; pattern-triggered immune (PTI), pattern-triggered immunity; spectral counts (SpC), spectral count(s); DAMP, damage-associated molecular pattern; Dex, dexamethasone; FDR, false discovery rate; GO, Gene Ontology; HRP, horseradish peroxidase; SNARE, soluble N-ethylmaleimide-sensitive factor attachment protein receptor; SAM, S-adenosylmethionine; and many downstream defense responses occur at the PM [1]
The bacterial effector AvrRpt2 is a protease that undergoes autoproteolysis, localizes to the PM, and cleaves the plant protein RIN4 twice at the sequence VPxFGxW, a molecular event that is recognized by the RPS2 resistance gene in the Col-0 background [5, 6, 45, 46]

Summary

Introduction

Because many classes of plant pathogens remain outside the host cell membrane during infection, the plant plasma membrane (PM)1 controls numerous facets of plant-microbe interactions. Leaf rosette tissue was harvested from Col-0 and GVG-AvrRpt2 plants 6 h post-Dex application, and the total microsomal fraction was immediately subjected to aqueous two-phase partitioning to enrich for plasma membrane vesicles [24]. The most highly enriched up-regulated processes include cellular responses to cadmium, primary and secondary metabolism, immune responses, abiotic stress responses, and proteasome-mediated protein catabolism (Fig. 3 and supplemental Table 2).

Results

Conclusion