Detopping causes production of intercellular space occlusions in both the cortex and infected region of soybean nodules

A structural analysis was conducted to determine whether glycoprotein-containing intercellular space occlusions are involved in medium-term regulation of O-2 diffusion in soybean (Glycine max) nodules. Alterations in O-2 diffusion were induced by a 3 h detopping treatment, and glycoprotein was immunolocalized with the monoclonal antibodies MAC236 and MAC265. Western blots of unstressed nodules revealed that these antibodies recognize antigens with two different molecular weights in soybean nodules. Tissue printing of halved nodules showed that both antigens were present in fresh nodules from control and 3 h detopped plants. The main localization appeared to be the inner cortex, but some immunolabelling also occurred in the infected region. ELISAs demonstrated a significant increase in total nodule concentration of intercellular glycoprotein following detopping, and cryosections of fresh nodules from this treatment also showed localization of antigens within the intercellular spaces of the infected region. The production of intercellular space occlusions in both the mid-cortex and infected regions after 3 h detopping was confirmed by light microscopy and silver-enhanced immunolabelling; cortical changes were quantified by image analysis techniques. Electron microscopy revealed that the occlusions within the infected region were less dense and less heavily labelled than those in the cortex. These results are discussed in relation to O-2 diffusion regulation in soybean nodules.