Pathways of ammonium assimilation

Prior to the demonstration that GOGAT occurs in higher plants (Lea and Miflin, 1974), GDH was considered to be the primary enzyme of ammonium assimilation in both plants and micro-organisms. There is now considerable evidence that in vascular plants ammonium assimilation occurs almost exclusively via the GS-GOGAT cycle (see Miflin and Lea, 1980 for review). Incubating plants with methionine sulphoxi-mine—a specific inhibitor of GS—almost invariably results in a complete inhibition of nitrogen assimilation in both root and shoot tissue. Feeding plants with 15N-labelled nitrate or ammonium and following the enrichment of amino acid pools by gas chromatography/mass spectrometry (GC/MS) or by nuclear magnetic resonance spectroscopy shows a rapid labelling of amido nitrogen of the glutamine pool that is indicative of the entry of inorganic nitrogen to organic molecules primarily via GS activity. These results exclude GDH from playing a role in ammonium assimilation by higher plants.

The relative importance of the GDH pathway and GS-GOGAT cycle for ammonium assimilation in mycorrhizal fungi is, however, not clear. It may be noted that a large group of fungi lack GOGAT activities. Furthermore, 15N labelling studies have produced convincing evidence for the assimilation of ammonium via the GDH pathway in the fungus

Candida utilis (Folkes and Sims, 1974). There are, however, conflicting reports about the presence of GOG AT in mycorrhizal fungi. Recently, Vezina et al. (1989) reported an active NADH-dependent GOG AT in Laccaria bicolor. However, the levels of GOG AT activity measured by Vezina et al. (1989) appear extremely low compared with the activity of GS and other enzymes of nitrogen assimilation present in the same fungal extract. Using a combination of 15N labelling of Cenococcum geophilum and the inhibition of its GS activity by methionine sulphoxi-mine, the work of Martin et al. (1988) has excluded a role of GOG AT in the synthesis of glutamate by this ectomycorrhizal ascomycete. The incorporation of nitrogen into amino acids in this fungus probably occurs via both the reductive amination reaction of GDH producing glutamate and the amination of glutamate by GS producing glutamine. A concurrent role of GS and NADPH-linked GDH pathways was also evident in our study of L. bicolor (Ahmad et al., 1990) which showed that the activity of these enzymes reached maximum levels in rapidly growing mycelia and declined rapidly during the onset of the stationary growth phase. A highly active NADPH-GDH has also been reported in Hebeloma spp. (Dell et al., 1989). These basidiomycetes and possibly other mycorrhizal fungi with similar enzymic characteristics provide useful material for elucidating the relative importance of the two potential pathways in the assimilation of nitrogen by fungal symbionts. It will be rewarding to adopt the isotopic procedures described by Martin and his co-workers (Martin, 1985; Martin et al., 1986) for C. geophilum in monitoring 15N labelling patterns of other fungal symbionts. The use of the GS inhibitor, methionine sulphoximine, has afforded information regarding the contribution of the GS pathway in the nitrogen assimilation of higher plants and green algae (Fentem et al., 1983; Ahmad and Hellebust, 1985, 1986), and one expects that it can be applied equally successfully in mycorrhizal studies. Selection of GS and GDH fungal mutants has surprisingly not been undertaken so far in mycorrhizal research. Intraspecific variability of the NADPH-GDH from Hebeloma has been studied by Wagner et al. (1989).

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