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Isolation of Intact Vacuoles and Proteomic Analysis of Tonoplast from Suspension-Cultured Cells of Arabidopsis thaliana
- Source :
- Plant and Cell Physiology. 45:672-683
- Publication Year :
- 2004
- Publisher :
- Oxford University Press (OUP), 2004.
-
Abstract
- A large number of proteins in the tonoplast, including pumps, carriers, ion channels and receptors support the various functions of the plant vacuole. To date, few proteins involved in these activities have been identified at the molecular level. In this study, proteomic analysis was used to identify new tonoplast proteins. A primary requirement of any organelle analysis by proteomics is that the purity of the isolated organelle needs to be high. Using suspension-cultured Arabidopsis cells (Arabidopsis Col-0 cell suspension), a method was developed for the isolation of intact highly purified vacuoles. No plasma membrane proteins were detected in Western blots of the isolated vacuole fraction, and only a few proteins from the Golgi and endoplasmic reticulum. The proteomic analysis of the purified tonoplast involved fractionation of the proteins by SDS-PAGE and analysis by LC-MS/MS. Using this approach, it was possible to identify 163 proteins. These included well-characterized tonoplast proteins such as V-type H+ -ATPases and V-type H+ -PPases, and others with functions reasonably expected to be related to the tonoplast. There were also a number of proteins for which a function has not yet been deduced.
- Subjects :
- Proteomics
Physiology
Arabidopsis
Cell Culture Techniques
Plant Science
Vacuole
Cell Fractionation
symbols.namesake
Gene Expression Regulation, Plant
Organelle
Arabidopsis thaliana
biology
Arabidopsis Proteins
Endoplasmic reticulum
Intracellular Membranes
Cell Biology
General Medicine
Golgi apparatus
biology.organism_classification
Cell biology
Inorganic Pyrophosphatase
Proton-Translocating ATPases
Membrane protein
Vacuoles
symbols
Carrier Proteins
Subcellular Fractions
Subjects
Details
- ISSN :
- 14719053 and 00320781
- Volume :
- 45
- Database :
- OpenAIRE
- Journal :
- Plant and Cell Physiology
- Accession number :
- edsair.doi.dedup.....a9b17ce4df883892aa568db9f7b6e6b9