In this study, the cell-wall-enriched subproteomes at three different heights of alfalfa stems were compared. Since these three heights correspond to different states in stem development, a view on the dynamics of the cell wall proteome during cell maturation is obtained. This study of cell wall protein-enriched fractions forms the basis for a description of the development process of the cell wall and the linking cell wall localized proteins with the evolution of cell wall composition and structure. The sequential extraction of cell wall proteins with CaCl2, EGTA, and LiCl-complemented buffers was combined with a gel-based proteome approach and multivariate analysis. Although the highest similarities were observed between the apical and intermediate stem regions, the proteome patterns are characteristic for each region. Proteins that bind carbohydrates and have proteolytic activity, as well as enzymes involved in glycan remobilization, accumulate in the basal stem region. Beta-amylase and ferritin likewise accumulate more in the basal stem segment. Therefore, remobilization of nutrients appears to be an important process in the oldest stem segment. The intermediate and apical regions are sites of cell wall polymer remodeling, as suggested by the high abundance of proteins involved in the remodeling of the cell wall, such as xyloglucan endoglucosylase, beta-galactosidase, or the BURP-domain containing polygalacturonase non-catalytic subunit. However, the most striking change between the different stem parts is the strong accumulation of a DUF642-conserved domain containing protein in the apical region of the stem, which suggests a particular role of this protein during the early development of stem tissues.
Method for characterizing extracellular proteins from the cell wall proteome of the copper tolerant fungus Phialophora malorum
