Characterization and regulation of ammonium transport systems in Citrus plants

We have investigated both the kinetics and regulation of (NH4+)-N-15 influx in roots of 3-month-old hydroponically grown Citrus (Citrus sinensis L. Os beck x Poncirus trifoliata Blanco) seedlings. The (NH4+)-N-15 influx is sat urable below an external ammonium concentration of 1 mM, indicating the act ion of a high-affinity transport system (HATS). The HATS is under feedback repression by the N status of the plant, being downregulated in plants adeq uately supplied with N during growth, and up-regulated by N-starvation. Whe n assayed between 1 and 50 mM [(NH4+)-N-15](0), the (NH4+)-N-15 influx show ed a linear response typical of a low-affinity transport system (LATS). The activity of the LATS increased in plants supplied with NH4+ as compared wi th plants grown on an N-free medium. Transfer of the plants to N-free solut ion resulted in a marked decrease in the LATS-mediated (NH4+)-N-15 influx. Accordingly, resupply of NH4+ after N-starvation triggered a dramatic stimu lation of the activity of the LATS. These data provide evidence that in Cit rus plants, the LATS or at least one of its components is inducible by NH4. Even when up-regulated, both the HATS and the LATS displayed a limited ca pacity, as compared with that usually found in herbaceous species. The use of various metabolic uncouplers or inhibitors indicated that (NH4+)-N-15 in flux mediated by the HATS is strongly dependent on energy metabolism and H transmembrane electrochemical gradient. By contrast, the LATS is not affec ted by protonophores or inhibitors of the H+-ATPase, suggesting that its ac tivity is mostly driven by the NH4+/NH3 transmembrane gradient. In agreemen t with these hypotheses, the HATS-mediated (NH4+)-N-15 influx was strongly inhibited when the solution pH was raised from 4 to 7, whereas influx media ted by the LATS was slightly stimulated.