Fig. 3.

The PIF4 response to nitrate requires active phyB while the growth response does not. Seedlings transferred from 0.5 mM to 5.0 mM nitrate 1 h after the beginning of the photoperiod of day 4 are compared to the controls that remained on 0.5 mM nitrate. (A) Hypocotyl growth rate in seedlings grown under simulated shade (transferred to shade the day before nitrate treatments). (B) Hypocotyl growth rate in seedlings of the wild type, phyB, and cry1 mutants. (C) Nuclear fluorescence driven by the pPIF4:PIF4-GFP transgene, measured by confocal microscopy 6 h after the nitrate upshift in seedlings simultaneously transferred to simulated shade. (D) Kinetics of the bioluminescence driven by pPIF4:PIF4-LUC during the photoperiod of day 4 in seedlings that were simultaneously transferred to simulated shade. (E) Bioluminescence driven by pPIF4:PIF4-LUC 1.5 h after the beginning of the white light photoperiod (WL) and in seedlings measured simultaneously after receiving a pulse of far-red light (FR) at the subjective time of initiation of the photoperiod and followed by darkness. (F) Rate of hypocotyl growth in seedlings treated exactly as in E. (G) Number of small and large phyB nuclear bodies in hypocotyl cells. Data are means ± SE and individual values (A–C, E, and F) or means ± SE of at least 70 seedlings (four plates with control and nitrate seedlings on the same plate, D). Asterisks indicate the significance of the effect of nitrate in Student’s t tests (A, C, and D), the main effect of nitrate in two-way ANOVA that showed no significant interaction (B and G, in E data were log transformed for the analysis to obtain homogeneity of variance) or the effect of nitrate in Bonferroni tests following significant interaction (int, E): *P < 0.05; **P < 0.01; ***P < 0.001; ns, not significant.