UBC Theses and Dissertations
Ammonium uptake by rice roots Wang, Miao Yuan
¹³NH₄+ uptake was studied using 3-week-old rice plants ( Oryza sativa L. cv. M202), grown hydroponically in modified Johnson’s nutrient solution containing 2, 100 or 1000 μM NH₄+ (referred to hereafter as G2, G100 or G1000 plants, respectively). At steady-state, the influx and efflux of ¹³NH₄+ was increased as NH₄+ provision during growth was increased. The half-life of cytoplasmic ¹³NH₄+ exchange was calculated to be 8 min while the half-life for cell wall exchange was 1 min. Cytoplasmic [NH₄+] of G2, G100 and G1000 roots was estimated to be 3.72, 20.55, and 38.08 mM respectively. However about 72% to 92% of total root NH₄+ was located in the vacuole. During a 30 minute period G100 plants metabolized 19% of the newly absorbed ¹³NH₄+ and the remainder was partitioned among the cytoplasm (41%), vacuole (20%) and efflux (20%). Of the metabolized ¹³N, roughly one half was translocated to the shoots. In short-term, perturbation experiments, below 1 mM external concentration ([NH₄+]₀), ¹³NH₄+ influx of G2, G100 and G1000 roots was saturable and operated by means of a high affinity transport system (HATS). The Vmax values for this transport system were negatively correlated and Km values were positively correlated with NH 4provision during growth and root [NH₄+]. Between 1 and 40 mM [NH₄+]₀,¹³NH₄+ influx showed a linear response to external concentration due to a low affinity transport system (LATS). The ¹³NH₄+ influxes by the HATS, and to a lesser extent the LATS, are energy-dependent processes. Selected metabolic inhibitors reduced influx of the HATS by 50 to 80%, but of the LATS by only 31 to 51%. EstimatedQ10 values for HATS were greater than 2.4 at root temperatures from 5 to 10°C and constant at ~ 1.5 between 5 to 30°C for the LATS. Influx of ¹³NH₄+ by the HATS was insensitive to external pH in the range from 4.5 to 9.0, but influx by the LATS declined significantly beyond pH 6.0. The transmembrane electrical potential differences (Δψ) of epidermal and cortical cells of intact roots were in the range from -120 to -140 millivolts (mV) in the absence of NH₄+ in bathing solution and were -116 mV and -89 mV for G2 and G100 plants in 2 and 100 μM NH₄+ solutions, respectively. Introducing NH₄+ to the bathing medium caused a rapid depolarization which exhibited a biphasic response to external [ NH₄+ ]. Plots of membrane depolarization versus ¹³NH₄+ influx were also biphasic, indicating distinct coupling processes for the two transport systems, with a break-point between the two concentration ranges around 1 mM NH₄+. Depolarization of Δψ due to NH₄+ uptake was eliminated by a protonophore (carboxylcyanide-m-chlorophenylhydrazone), inhibitors of ATP synthesis (sodium cyanide plus salicylhydroxamic acid), or an ATPase inhibitor (diethyistilbestrol). ¹³NH₄+ influx was regulated by internal ammonium and its primary metabolites, amides and amino acids. When internal amide or amino acids concentrations were increased, the influx of ¹³NH₄+ was reduced. However, treating rice roots with L-Methionine DL-Sulfoximine (MSX) reduced the levels of ammonium assimilates but did not increase ¹³NH₄+ influx probably because internal [NH₄+] was increased. Short-term nitrogen depletion stimulated ¹³NH₄+ influx, but long-term N depletion caused NH₄+ influx to be reduced probably due to N limitation of carrier synthesis. A cascade regulation system is proposed to explain the multi-level regulation of NH₄+ influx. The interaction between ammonium and potassium showed that when N is adequate, K promoted NH₄+ uptake and utilization. Likewise, proper N nutrition promoted K+ uptake but the presence of NH₄+ in uptake solution strongly inhibited the K+ (⁸⁶Rb+) uptake at the transport step. The results indicated that NH₄+ and K+ may share the same channel but are regulated by different feedback signals.
Item Citations and Data