title: A generic approach for "shotgun" analysis of the soluble proteome of plant cell suspension cultures.
abstract: Cell suspension cultures from different plant species act as important model systems for studying cellular processes in plant biology and are often used as "green factories" for the production of valuable secondary metabolites and recombinant proteins. While mass spectrometry based proteome analysis techniques are ideally suited to study plant cell metabolism and other fundamental cellular processes from a birds eye perspective, they remain underused in plant studies. We describe a comprehensive sample preparation and multidimensional 'shotgun' proteomics strategy that can be generically applied to plant cell suspension cultures. This strategy was optimized and tested on an Arabidopsis thaliana ecotype Landsberg erecta culture. Furthermore, the implementation of strong cation exchange chromatography as a peptide fractionation step is elaborately tested. Its utility in mass spectrometry based proteome analysis is discussed. Using the presented analytical platform, over 13,000 unique peptides and 2640 proteins could be identified from a single plant cell suspension sample. Finally, the experimental setup is validated using Nicotiana tabacum cv. "Bright Yellow-2" (BY-2) plant cell suspension cultures, thereby demonstrating that the presented analytical platform can also be valuable tool in proteome analysis of non-genomic model systems.
authors.affiliation: Center for Proteomics (CFP), Groenenborgerlaan 171, B-2020 Antwerp, Belgium; Laboratory of Plant Growth and Development, Department of Biology, University of Antwerp, Groenenborgerlaan 171, Antwerp, Belgium. Electronic address: thomas.devijlder@uantwerpen.be.
authors.forename: Thomas
authors.initials: T
authors.lastname: De Vijlder
authors.affiliation: Center for Proteomics (CFP), Groenenborgerlaan 171, B-2020 Antwerp, Belgium; Vlaamse Instelling voor Technologisch Onderzoek (VITO), Boeretang 200, B-2400 Mol, Belgium; Interuniversity Institute for Biostatistics and Statistical Bioinformatics, Hasselt University, Agoralaan 1, B-3590 Diepenbeek, Belgium.
authors.forename: Dirk
authors.initials: D
authors.lastname: Valkenborg
authors.affiliation: Biomolecular & Analytical Mass Spectrometry, Department of Chemistry, University of Antwerp, Groenenborgerlaan 171, B-2020 Antwerp, Belgium.
authors.forename: Debbie
authors.initials: D
authors.lastname: Dewaele
authors.affiliation: Center for Proteomics (CFP), Groenenborgerlaan 171, B-2020 Antwerp, Belgium; Laboratory of Plant Growth and Development, Department of Biology, University of Antwerp, Groenenborgerlaan 171, Antwerp, Belgium.
authors.forename: Noor
authors.initials: N
authors.lastname: Remmerie
authors.affiliation: Biomedical Informatics Research Center Antwerp (Biomina), University of Antwerp/Antwerp University Hospital, Wilrijkstraat 10, B-2650 Edegem, Belgium; Advanced Database Research and Modelling, Department of Mathematics and Computer Science, University of Antwerp, Middelheimlaan 1, B-2020 Antwerp, Belgium.
authors.forename: Kris
authors.initials: K
authors.lastname: Laukens
authors.affiliation: Center for Proteomics (CFP), Groenenborgerlaan 171, B-2020 Antwerp, Belgium; Vlaamse Instelling voor Technologisch Onderzoek (VITO), Boeretang 200, B-2400 Mol, Belgium; Laboratory for Ecophysiology, Biochemistry and Toxicology, Department of Biology, University of Antwerp, Groenenborgerlaan 171, B-2020 Antwerp, Belgium.
authors.forename: Erwin
authors.initials: E
authors.lastname: Witters
mesh_terms: D017360:Arabidopsis / Q000737:chemistry / Q000378:metabolism*; D002478:Cells, Cultured; D013058:Mass Spectrometry / Q000379:methods*; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D059828:Plant Cells / Q000737:chemistry* / Q000378:metabolism; D010940:Plant Proteins / Q000737:chemistry* / Q000378:metabolism; D040901:Proteomics; D014026:Nicotiana / Q000737:chemistry / Q000378:metabolism*
chemical_list: D010455:Peptides; D010940:Plant Proteins
keywords: Arabidopsis thaliana; Mass spectrometry; Nicotiona tabacum cv “Bright Yellow-2”; Plant cell suspension cultures; Proteome analysis