The purification of plant cell walls is challenging because they constitute an open compartment which is not limited by a membrane like the cell organelles. Different strategies have been established to limit the contamination by proteins of other compartments in cell wall proteomics studies. Non-destructive methods rely on washing intact cells with various types of solutions without disrupting the plasma membrane in order to elute cell wall proteins. In contrast, destructive protocols involve the purification of cell walls prior to the extraction of proteins with salt solutions. In both cases, proteins known to be intracellular have been identified by mass spectrometry in cell wall proteomes. The aim of this chapter is to provide tools to assess the subcellular localization of the proteins identified in cell wall proteomics studies, including: (1) bioinformatic predictions, (2) immunocytolocalization of proteins of interest on tissue sections and (3) in muro observation of proteins of interest fused to reporter fluorescent proteins by confocal microscopy. Finally, a qualitative assessment of the work can be performed and the strategy used to prepare the samples can be optimized if necessary.
Keywords: Bioinformatics; Cell wall protein; Fluorescent reporter protein; Immunocytolocalization; Microscopy; Plant; Proteomics; Subcellular localization.