Figure 1. Nitrate triggers unique Ca²⁺-CPK signalling.

a, Dual nitrate functions in plants. b, Arabidopsis seedling development without exogenous nitrogen up to 4 days. Scale bar, 1 mm. The images are representative of 10 seedlings. c, d, Nitrate specifically promotes extensive shoot and root developmental programs. Different nitrogen sources (1 mM) were applied at day 5. Error bars, ± s.e.m., n=10 seedlings. Scale bar, 1 cm. e, Time-lapse images of Ca²⁺ signalling in the nucleus and cytosol stimulated by nitrate in mesophyll protoplasts expressing GCaMP6. KNO_3, 10 mM. BF, bright field. NLS-mC, nuclear HY5-mCherry. Scale bar, 10 μm. The images are representative of 10 protoplasts. f, Traces of GCaMP6 signals stimulated by nitrate in mesophyll protoplasts. KNO_3, 10 mM. (F-F₀)/F₀, relative fluorescence intensity. Error bars (grey areas, 199 error bars), ± s.e.m., n=10 protoplasts. g, Time-lapse images of Ca²⁺ signalling in mesophyll cells of cotyledons in GCaMP6 transgenic plants. KNO_3, 10 mM. Scale bar, 50 μm. The images are representative of 10 cotyledons. h, Subgroup III CPKac and nitrate synergistically activate NIR-LUC in protoplast transient assays. UBQ10-GUS is a control. KNO_3, 0.5 mM, 2 h. Error bars, s.d., n=3 biological replicates. Dash line: KNO₃ induction without CPKac. CPKac-FLAG expression was determined by immunoblot analyses. Source data can be found in Supplementary Information. i, The relation tree of subgroup III CPKs.