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Updated: Aug 11, 2026
Metabolic Labeling and Membrane Fractionation for Comparative Proteomic Analysis of Arabidopsis thaliana Suspension Cell Cultures
Published on: September 28, 2013
A mass spectrometric approach to identify arbuscular mycorrhiza-related proteins in root plasma membrane fractions
Benoît Valot1, Luc Negroni, Michel Zivy
1UMR 1088 INRA/CNRS 5184/UB Plante-Microbe-Environnement, Dijon, France.
Abstract:
One of the most important morphological changes occurring in arbuscular mycorrhizal (AM) roots takes place when the plant plasma membrane (PM) invaginates around the fungal arbuscular structures resulting in the periarbuscular membrane formation. To investigate whether AM symbiosis-specific proteins accumulate at this stage, two complementary MS approaches targeting the root PM from the model legume Medicago truncatula were designed. Membrane extracts were first enriched in PM using a discontinuous sucrose gradient method. The resulting PM fractions were further analysed with (i) an automated 2-D LC-MS/MS using a strong cation exchange and RP chromatography, and (ii) SDS-PAGE combined with a systematic LC-MS/MS analysis. Seventy-eight proteins, including hydrophobic ones, were reproducibly identified in the PM fraction from non-inoculated roots, representing the first survey of the M. truncatula root PM proteome. Comparison between non-inoculated and Glomus intraradices-inoculated roots revealed two proteins that differed in the mycorrhizal root PM fraction. They corresponded to an H(+)-ATPase (Mtha1) and a predicted glycosylphosphatidylinositol-anchored blue copper-binding protein (MtBcp1), both potentially located on the periarbuscular membrane. The exact role of MtBcp1 in AM symbiosis remains to be investigated.
Insights
Researchers identified two key proteins, an H(+)-ATPase and a blue copper-binding protein, accumulating in Medicago truncatula root plasma membranes during arbuscular mycorrhizal (AM) symbiosis. This finding sheds light on the molecular mechanisms of this crucial plant-fungal interaction.
Area of Science:
- Plant Biology
- Mycology
- Biochemistry
Background:
- Arbuscular mycorrhizal (AM) symbiosis is a vital plant-fungal interaction.
- Plant plasma membrane (PM) invagination forms the periarbuscular membrane (PAM) during AM symbiosis.
- Understanding proteins involved in PAM formation is crucial for AM symbiosis research.
Purpose of the Study:
- To investigate the proteome of Medicago truncatula root PM.
- To identify proteins that accumulate specifically in the PM during AM symbiosis.
Main Methods:
- PM enrichment using discontinuous sucrose gradient.
- Proteomic analysis using 2-D LC-MS/MS and SDS-PAGE.
- Comparative analysis of non-inoculated and AM-symbiotic roots.
Main Results:
- First comprehensive survey of the Medicago truncatula root PM proteome, identifying 78 proteins.
- Identified two proteins, H(+)-ATPase (Mtha1) and a blue copper-binding protein (MtBcp1), that accumulate in the mycorrhizal root PM.
- Both identified proteins are potentially located on the periarbuscular membrane.
Conclusions:
- The study provides insights into the proteomic changes in root PM during AM symbiosis.
- Mtha1 and MtBcp1 are potential key players in the periarbuscular membrane during AM symbiosis.
- Further research is needed to elucidate the specific role of MtBcp1 in AM symbiosis.
