Abstract
Proteome-wide discovery of in vivo metacaspase substrates can be obtained by positional proteomics approaches such as N-terminal COFRADIC, for example by comparing the N-terminal proteomes (or N-terminomes) of wild-type plants to transgenic plants not expressing a given metacaspase. In this chapter we describe a protocol for the preparation of plant tissue proteomes, including differential isotopic labelling allowing for a comparison of in vivo N-terminomes that serves as the starting point for N-terminal COFRADIC studies.
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Acknowledgments
L.T. acknowledges support from the VIB International PhD Program and the Netherlands Proteomics Centre, a program embedded in The Netherlands Genomics Initiative. P.V.D. is a Postdoctoral Fellow of the Research Foundation Flanders (FWO-Vlaanderen) and S.S. is indebted to the Special Research Fund of Ghent University for a postdoctoral fellowship. F.V.B acknowledges support from grants of the Ghent University Multidisciplinary Research Partnership “Ghent BioEconomy” 27 (project no. 01MRB510W) and of the Belgian Science Policy Office (project IAP7/29). F.V.B. and K.G. acknowledge support from the Research Foundation Flanders (FWO-Vlaanderen), research project G.0038.09.
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Tsiatsiani, L., Stael, S., Van Damme, P., Van Breusegem, F., Gevaert, K. (2014). Preparation of Arabidopsis thaliana Seedling Proteomes for Identifying Metacaspase Substrates by N-terminal COFRADIC. In: V. Bozhkov, P., Salvesen, G. (eds) Caspases,Paracaspases, and Metacaspases. Methods in Molecular Biology, vol 1133. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-0357-3_16
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DOI: https://doi.org/10.1007/978-1-4939-0357-3_16
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