To investigate the short-term (30-240 min) interactions among nitrogenase activity, NH 4 ؉ assimilation, and plant glycolysis, we measured the concentrations of selected C and N metabolites in alfalfa (Medicago sativa L.) root nodules after detopping and during continuous exposure of the nodulated roots to Ar:O 2 (80:20, v/v). Both treatments caused an increase in the ratios of glucose-6-phosphate to fructose-1,6-bisphosphate, fructose-6-phosphate to fructose-1,6-bisphosphate, phosphoenolpyruvate (PEP) to pyruvate, and PEP to malate. This suggested that glycolytic flux was inhibited at the steps catalyzed by phosphofructokinase, pyruvate kinase, and PEP carboxylase. In the Ar:O 2 -treated plants the apparent inhibition of glycolytic flux was reversible, whereas in the detopped plants it was not. In both groups of plants the apparent inhibition of glycolytic flux was delayed relative to the decline in nitrogenase activity. The decline in nitrogenase activity was followed by a dramatic increase in the nodular glutamate to glutamine ratio. In the detopped plants this was coincident with the apparent inhibition of glycolytic flux, whereas in the Ar:O 2 -treated plants it preceded the apparent inhibition of glycolytic flux. We propose that the increase in the nodular glutamate to glutamine ratio, which occurs as a result of the decline in nitrogenase activity, may act as a signal to decrease plant glycolytic flux in legume root nodules.N fixation, the reduction of N 2 to NH 4 ϩ by (Brady)rhizobium bacteria, in legume root nodules is catalyzed by the bacterial enzyme nitrogenase. Nitrogenase is O 2 labile, but the root nodule provides an environment that protects it from excess O 2 . Nitrogenase activity in legume root nodules is rapidly inhibited by treatments that affect plant C and N metabolism, such as exposure of the roots to C 2 H 2 , replacement of N 2 in the rhizosphere with Ar, NO 3 Ϫ fertilization, defoliation, detopping, phloem girdling, and drought (Minchin et al
