NITRATE FLUXES IN SOYBEAN SEEDLING ROOTS AND THEIR RESPONSE TO AMINO-ACIDS - AN APPROACH USING N-15

This study was undertaken to determine which of the two NO; fluxes (in flux or efflux) across plasma membranes of root cells is the target of those amino acids which have been shown to inhibit net NO3- uptake (M uller & Touraine 1992, Journal of Experimental Botany 43, 617-623), Pa rallel experiments were performed to measure either the time course of (NO3-)-N-15 release from roots of soybean seedlings previously labell ed with this isotope into non-labelled solution, or the time course of N-15 accumulation from labelled (NO3-)-N-15 solution in non-labelled seedlings, Focusing on the fate of (NO3-)-N-15 in the cytoplasmic comp artment, a model is developed to describe the time courses of the accu mulation and release of tracer across the plasma membranes of root cel ls, Both time courses can be described by the sum of an exponential pl us a linear term, In our material, the linear part of the accumulation time course is obscured by the NO3- fluxes exiting the cytoplasm, and the curve thus appears to be quasilinear over several minutes, Howeve r, we show that the use of the net tracer accumulation rate during thi s time period as an estimate of NO3- influx does not provide accurate estimates of influx and efflux, By contrast, (NO3-)-N-15 efflux analys is permits calculation of the unidirectional fluxes across plasma memb ranes of root cells and the kinetic parameters of the cytoplasmic NO3- pool, Under our experimental conditions, efflux accounted for 30 to 5 0% of influx, and the cytoplasmic NO3- content was found to be in the 70-400 nmol g(-1) fw range, Using this methodology, the effect of amin o acid accumulation on unidirectional fluxes of nitrate was then exami ned, Pretreatments of the seedlings with an amino acid which has been shown to inhibit net NO3- uptake led to concomitant decreases in net a ccumulation rates of (NO3-)-N-15 and of reduced N-15 in roots and tota l N-15 in cotyledons, NO3- influx was markedly inhibited by these trea tments, while NO3- efflux remained essentially unaffected, or even dec reased, It is concluded that the target of the regulation of NO3- upta ke by phloem-translocated amino acids is the influx system.