Pectins esterification in the apoplast of aluminum-treated pea root nodules

Aiming to elucidate the possible involvement of pectins in aluminum-mediated growth inhibition the distribution of pectins in the apoplast of root nodules was investigated. Experiments were performed on the pea (Pisum sativum L.) root nodules treated with aluminum (50 μM AlCl3, for 2 or 24 h). For histochemical acidic pectin localization we used ruthenium red staining. Immunolabeling techniques with monoclonal antibodies specific to high methyl-esterified pectin (JIM7), low methyl-esterified pectin (JIM5) and calcium cross-linked pectin (2F4) were used to re-examine the pattern of pectin esterification and distribution. After immunolabeling the samples were observed using a fluorescent and transmission electron microscope.Ruthenium red staining showed that acid pectin content increased in the apoplast of Al-treated nodules and immunolocalization of pectin epitopes revealed that the fraction of de-esterified pectins increased significantly under Al stress. JIM5 and 2F4 epitopes were located on the inner surface of the primary cell wall with higher intensity at cell corners lining the intercellular spaces and at infection threads (ITs) walls. By contrast, JIM 7 labels all walls uniformly throughout the nodule. In the presence of Al, the increase of JIM5 and 2F4 labeling in thick plant and IT walls, together with a decrease of JIM7 labeling was observed.These results indicate a specific role for pectin de-esterification in the process of wall thickening and growth inhibition. In particular, Al-dependent increase in pectin content and their low methyl esterification degree correlate with wall thickness and higher rigidity, and in this way, may affect IT and nodules growth.