NITRATE FLUXES IN SQUASH SEEDLINGS MEASURED WITH N-13

Influx of N-13--labelled NO3- was studied in roots of intact squash seedlings (Cucurbita max. Duch.) previously grown in nutrient solution without (uninduced) or with NO3- (induced). At the uninduced constitutive level NO3- influx was saturable (V(max) = 6.2-mu-mol (g FW)-1.h-1; K(m) = 37.0-mu-M), but it increased again, probably passively, at high external NO3- concentrations. Initial NO3- efflux started during induction of the transport system when the influx rate increased and conceivably the NO3- concentration in the cytoplasm was raised. Likewise, this tendency was observed after a period of NO3- starvation. The kinetic parameters of NO3- influx of the substrate-induced high capacity transport system accounted for a V(max) of 14.0-mu-mol (g FW)-1 h-1 and a K(m) of 15-mu-M. Both the uninduced and the induced uptake reacted in a similar pattern to the treatment of inhibitors affecting the protein synthesis (cycloheximide, fluorophenylalanine or puromycin) suggesting a single membrane protein population carrying NO3- across the plasmalemma. A rapid onset of a strong inhibition of the NO3- influx by the treatment with 1 mM phenyglyoxal was revealed. The influx recovered, but only slowly and was not much enhanced after 5 h of recovery at increasing NO3-concentrations (100 to 8000-mu-M) in the ambient solution. Compared with barley, the relative tolerance of squash NO3- influx to the presence of up to 5 MM NH4+ in the solution was confirmed. Only after pretreatment with 5 mM NH4+ was the influx depressed, presumably because of feedback signals resulting from metabolite accumulation from the NH4+ assimilation.