Planta (1984) 162:32%333 Planta 9 Springer-Verlag 1984 Nitrogen nutrition and the development of biochemical functions associated with nitrogen fixation and ammonia assimilation of nodules on cowpea seedlings C.A. Atkins, B.J. Shelp, P.J. Storer and J.S. Pate Department of Botany, University of Western Australia, Nedlands, WA 6009, Australia Abstract. During early development (up to 18 d after sowing) of nodules of an "effective" cowpea symbiosis (Vigna unguiculata (L.) Walp cv. Vita 3:Rhizobium strain CB756), rapidly increasing ni- trogenase (EC 1.7.99.2) activity and leghaemoglo- bin content were accompanied by rapid increases in activities of glutamine synthetase (EC 6.3.1.2), glutamate synthase (EC 2.6.1.53), enzymes of de- novo purine synthesis (forming inosine monophos- phate) xanthine oxidoreductase (EC 1.2.3.2), urate oxidase (EC 1.7.3.3), phosphoenolpyruvate car- boxylase (EC 4.1.1.31) and led to increased export of ureides (allantoin and allantoic acid) to the shoot of the host plant in the xylem. Culturing plants with the nodulated root systems maintained in the absence of N 2 (in 80 At:20 02, v/v) had little effect on the rates of induction and increase in nitrogenase activity and leghaemoglobin content but, in the absence of N 2 fixation and consequent ammonia production by bacteroids, there was no stimulation of activity of enzymes of ammonia as- similation or of the synthesis of purines or ureides. Addition of NO2 (0.1-0.2 mM) relieved host-plant nitrogen deficiency caused by the At: O2 treatment but failed to increase levels of enzymes of N metab- olism in either the bacteroid or the plant-cell frac- tions of the nodule. Premature senescence in Ar:O2-grown nodules occurred at 18-20 d after sowing, and resulted in reduced levels of nitrogen- ase activity and leghaemoglobin but increased the activity of hydroxybutyrate oxidoreductase (EC 1.1.1.30). Key words: Ammonia - Nitrogen fixation - Nod- ule - Senescence (root nodules) - Ureide - Vigna. Introduction Many factors are likely to be involved in regulating the development and maturation of an effective Rhizobium-legume host symbiosis (for a recent re- view, see Sutton 1983). Once successful infection has resulted in the establishment of a pre-emergent nodule structure (for review, see Newcomb 1981) within the legume root, transformation into a func- tional nodule is extremely rapid and involves a complex set of morphological and biochemical changes, especially in relation to the transition of meristematic plant cells to infected cells. These events are usually associated with extremely rapid growth by the nodule and a dramatic increase in nitrogenase activity (Atkins et al. 1980b). Studies of a wide range of legume nodules have shown that concurrent with the above changes there is an equally rapid rise in the activity of en- zymes associated with ammonia assimilation in the host cells and it has been supposed (Robertson et al. 1975a, b; see review by Atkins 1982) that induction of these enzymes results from the onset of ammonia excretion by the bacteroids. In this paper, we attempt to determine directly the extent to which the nitrogenous products of nitrogenase activity might influence the differentiation of in- fected plant tissues and nodule functioning, by ex- amining patterns of enzymic activity in nodules of cowpea formed in the presence of N 2 or in an atmosphere of Ar: 0 2 in which ammonia forma- tion by nitrogenase is prevented. In a companion paper (Atkins et al. 1984) the general effects of Ar:O 2 on cowpea seedling development have been described, and it has been shown how nodule growth and structure are influenced by this gas- eous treatment in the presence or absence of NO 3. Materials and methods Plant material. Cowpea (Vigna unguiculata (L.) Walp cv. Vita 3) seedlings nodulated with Rhizobium strain CB756 were grown in N-free or in NO3--supplemented liquid culture with a rooting atmosphere of air or Ar:O2 (80:20, v/v) as described in Atkins et al. (1984).