Relationships between external nitrate availability, nitrate uptake and expression of nitrate reductase in roots of barley grown in N-limited split-root cultures

Despite the large number of studies of nitrate metabolism in plants, it remains undetermined to what extent this key plant system is controlled by overall plant N nutrition on the one hand, and by the nitrate ion itself on the other hand. To investigate these questions, Vmax for nitrate uptake (high-affinity range), and nitrate reductase (NR) mRNA and activity, were measured in roots of N-limited barley (Hordeum vulgare L. cv. Golf) grown under conditions of constant relative addition of nitrate, with the seminal roots split between two culture compartments. The total amount of nitrate added per unit time (0.09 d-1) was distributed between the two root parts (subroots) in ratios of 100: 0, 98: 2, 95: 5, 90: 10, 80: 20, and 50:50. These nitrate-addition ratios resulted in nitrate fluxes ranging from 0 to 23 micromoles nitrate get DW root.h-1, while the external nitrate concentrations varied between 0 and 1.2 micromolar. The apparent Vmax for net nitrate uptake showed saturation-type responses to nitrate flux maintained during preceding growth. The flux resulting in half-maximal "induction" of nitrate uptake was approximately 4 micromoles nitrate g-1 DW root h-1, corresponding to an external nitrate concentration of 0.7 micromolar. The activity of NR and levels of NR mRNA did not saturate within the range of nitrate fluxes studied. None of the parameters studied saturated with respect to the steady-state external nitrate concentration. At the zero nitrate addition--the 0%-root--initial uptake activity as determined in short-term 15N-labelling experiments was insignificant, and NR activity and NR mRNA were not detectable. However nitrate uptake was rapidly induced, showing that the 0%-root had retained the capacity to respond to nitrate. These results suggest that local nitrate availability has a significant impact on the nitrate uptake and reducing systems of a split-root part when the total plant nitrate nutrition is held constant and limiting.