Figure 2.

Functional characterization of NPF5.11, NPF5.12 and NPF5.16 in oocytes. (a–e) Currents elicited in oocytes injected with H₂O (a), CHL1 cRNA (b), NPF5.11 cRNA (c), NPF5.12 cRNA (d) or NPF5.16 cRNA (e). Oocytes were voltage clamped at −60 mV and representative inward currents elicited by 10 mM NO₃ ⁻ at pH 5.5 were recorded. (f–h) Nitrate uptake activity in oocytes injected with H₂O, NPF5.11 cRNA, NPF5.12 cRNA, NPF5.16 cRNA, NRT1.8 cRNA or CHL1 cRNA. Oocytes were incubated with 10 mM ¹⁵NO₃ ⁻ at pH 5.5 (f), 0.25 mM ¹⁵NO₃ ⁻ at pH 5.5 (g) or 10 mM ¹⁵NO₃ ⁻ at pH 7.4 (h) for 12 h. Values are means ± SD (n = 8–12). Asterisks indicate difference at P < 0.01 (**) compared with the H₂O-injected oocytes by Student’s t-test. (i–k) Uptake kinetics of NPF5.11 (i), NPF5.12 (j) and NPF5.16 (k). Oocytes injected with NPF5.11 cRNA (i), NPF5.12 cRNA (j) or NPF5.16 cRNA (k) were incubated with indicated concentrations of ¹⁵NO₃ ⁻ at pH 5.5 for 1.5 h, and the ¹⁵N contents were determined. Values are means ± SD (n = 6–12). The Km was calculated by fitting to the Michaelis-Menten equation using a nonlinear least squares method in the SigmaPlot program. The Km was 2.57 mM, 4.84 mM, or 2.91 mM for NPF5.11, NPF5.12 or NPF5.16, respectively.