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 arid 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.