The aim of the present study was to test the hypothesis that the N content or the composition of the phloem sap that supplies nodulated roots may play a role in the feedback regulation of nitrogenase activity by increasing nodule resistance to O, diffusion. High nitrate levels in soils are known to inhibit both nodule formation and nitrogenase activity in legume nodules (Streeter, 1988). The inhibitory effects of nitrate on specific nitrogenase activity have been classified into those that occur within the first 2 or 3 d (Vessey et al., 1988b;Minchin et al., 1989; Escuredo et al., 1996) and those that result from longer term exposure to nitrate (Minchin et al., 1989; Escuredo et al., 1996). There is widespread evidence that the initial effect of nitrate exposure causes the respiration rate and nitrogenase activity of legume nodules to become severely O, limited, as a result of an increase in the nodule resistance to O, diffusion and a decrease in the concentration of O, in the bacteria-infected cells (Schuller et al., 1988;Vessey et al., 1988a;Minchin et al., 1989;Layzell et al., 1990;Vessey and Waterer, 1992; Hunt and Layzell, 1993; Denison and Harter, 1995; Escuredo et al., 1996). In nitrate-inhibited nodules nitrogenase activity and nodule respiration rates can recover partially or sometimes to rates approximating a pre-decline status by increases in externa1 p0, to levels greater than that in air (Minchin et al., 1986(Minchin et al., , 1989Schuller et al., 1988;Vessey et al., 1988a; Faurie and Soussana, 1993; Kaiser et al., 1994; Escuredo et al., 1996).Various hypotheses have been proposed to explain the initial inhibitory effects of nitrate and the role that O, plays in the phenomena. For example, within the first few days of exposure, nitrate accumulates in the nodule cortex where it has been proposed to cause water to move out of cortical cells and into the gas-filled intercellular spaces, thus increasing the resistance of the nodule to O, diffusion (Sprent et al., 1987;Minchin et al., 1989;Serraj et al., 1995). Nitrate could also enter the central zone of nodules, where it may be converted to nitric oxide, which could bind to leghemoglobin to form nitrosylleghemoglobin, a heme protein that is unable to facilitate the diffusion of O, to the bacteroids (Kanayama and Yamamoto, 1990a, 1990b Kanayama et al., 1990). However, the latter hypothesis has been recently challenged by a nodule oximetry study (Denison and Harter, 1995) that showed no decrease in the functional leghemoglobin concentration in nodules that are exposed to nitrate.Other studies have shown that nitrate reduces photosynthate partitioning to nodules (Vessey et al., 1988a, 198813; Faurie and Soussana, 1993), and because treatments that disrupt the phloem sap supply to nodules (e.g. nodule excision, plant defoliation, stem girdling, or stem chilling) also induce severe O, limitation within the infected cells (Minchin et al., 1985;Walsh et al., 1987;Schuller et al., 1988; Vessey et al., 1988a, 198813; Faurie and Soussana, 1993; Hartwig ...

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In soybeans (Glycine max L. Merr.), high levels of soil nitrate inhibit N2 fixation, and nitrate‐tolerant symbioses have been identified within a chemically mutagenized line of cv. Bragg denoted nts382 and within the line K466, a genotype representative of a number of Korean soybean cultivars. The genotypes nts382 and K466 were examined to see if they could be used as a model system for studying the mechanism responsible for the short‐term (i.e. 3‐day) inhibition of specific nitrogenase activity, especially the mechanism behind the greater O2 limitation of nodule metabolism that is characteristic of nitrate inhibition of N2 fixation in soybean. In nts382, total nitrogenase activity (TNA = H2 production in Ar:O2) was inhibited to a lesser degree (48% of control) relative to Bragg (30% of control), and the nitrate‐treated symbioses showed less of an O2 limitation of nodule metabolism in nts382 than in Bragg. However, the relative proportion of O2 limitation to the total nitrate inhibition was similar (40 and 41%) in nts382 and Bragg, respectively. Therefore, the nts382 symbioses may be useful in elucidating the general mechanism for down‐regulation of nitrogenase activity in soybean, but would not be a useful model system for studying the control of O2‐limited metabolism following nitrate exposure. The effects of nitrate on TNA and on the degree of O2 limitation of nodule metabolism were the same in K466 and a reference cultivar Maple Arrow. Consequently, the tolerance of K466 to nitrate reported previously was attributed to the ability of this symbiosis to maintain nodule biomass in the presence of nitrate, not to any ability to maintain specific nitrogenase activity in the presence of nitrate.