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Keywords:

  • Sesbania rostrata;
  • chloroplasts;
  • glycoprotein;
  • immunogold-labellmg;
  • nitrogen fixation;
  • oxygen;
  • photosystem II;
  • stem nodules

ABSTRACT

The tropical wetland legume Sesbania rostrata Brem. produces nitrogen-fixing stem nodules which are green and contain chlorophyll, the chloroplasts being concentrated in a hand in the inner and mid-cortex close to the nitrogen-fixing cells. The photosystem II thylakoid membrane proteins D1, D2 and PsbO, which are essential for photo-synthetic O2 evolution, were shown by immunoblotting to be present in extracts of leaves and stem nodules. Immunogold labelling confirmed their presence on stem nodule thylakoids and showed that labelling was most intense in well-developed chloroplasts in the mid-cortex and least intense in the smaller, less-abundant chloroplasts adjacent to the nitrogen-fixing cells. Concentrations of the oxygen-carrying protein leghaemoglobin (Lb) did not differ between stem and S. rostrata root nodules, and Lb was localized in bacteroid-containing cells, including those immediately adjacent to the cortex, in both nodule types. Moreover, nitrogenase component 2 was localized in bacteroids within the outermost layers of infected cells, suggesting that a low pO2 was maintained, despite the nearby chloroplasts. Nodule extracts examined by ELISA and immunoblots, using the monoclonal antibody MAC265, showed greatly enhanced expression of a 139 kDa glycoprotein in stem compared to root nodules. Immunogold labelling showed that material containing the MAC265 antigen occluded intercellular spaces, and was present in cell walls, throughout the cortex of stem nodules (particularly in the chloroplasl-rich inner and mid-cortex), but was considerably less evident in root nodules.