Submitted to: Plant Physiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/8/1999
Publication Date: N/A
Citation: Interpretive Summary: Nitrogen (N) is a major nutrient required by plants. In legume species, like alfalfa, N is acquired through symbiotic N2 fixation. This process occurs in small wart-like structures on roots, termed nodules. A soil bacterium, Rhizobium, lives within the nodule and provides the plant with N fertilizer derived from symbiotic N2 fixation. The plant gives the Rhizobium sugars from photosynthesis to provide energy. Before the plant can utilize the N derived from symbiotic N2 fixation, that N must be converted to a useful form. This conversion is brought about by plant enzymes (protein catalysts). To keep N assimilation in nodules working at maximum speed the plant must direct enzymes to different cells and unique locations within the cell. In this report we identify the cells and locations within the cells for an important enzyme in N assimilation in root nodules. This enzyme, glutamate synthase, is involved in the initial steps in the conversion of fixed N into a useful form. Glutamate synthase was found to be primarily accumulating in the cells where active N2 fixation is occurring and it is located in very small particles, amyloplasts, within the cell. The amount of glutamate synthase is reduced or absent in root nodules that do not fix N2. These findings are important because strategies aimed at improving N2 fixation and N assimilation require the exact location of enzymes in the process be known. In addition, the data showing that glutamate synthase is in amyloplasts indicate that N assimilation is closely linked to plant carbon metabolism, since amyloplasts are the known location of starch formation. Such data show plant biologists that efforts to improve or change N assimilation must take into account accompanying changes in carbon metabolism.
Technical Abstract: In root nodules of alfalfa, (Medicago sativa L.), N2 is reduced to NH4+ in the bacteroid by the nitrogenase enzyme and then released into the plant cytosol. The NH4+ is then assimilated by the combined action of glutamine synthetase (GS) and NADH-dependent glutamate synthase (NADH-GOGAT; EC 188.8.131.52) into glutamine and glutamate. The alfalfa nodule NADH-GOGAT protein has a 101 amino acid presequence, but the subcellular location of the protein is unknown. Using immunocytochemical localization, we determined first that the NADH-GOGAT protein is found throughout the infected cell region of both 19- and 33-day-old nodules. Second, in alfalfa root nodules NADH-GOGAT is localized predominantly to the amyloplast of infected cells. This finding, together with earlier localization and fractionation studies, indicates that in alfalfa the infected cells are the main location for the initial assimilation of fixed N2.