Figure 5. Impact of FNR location on photosynthetic electron transport in Arabidopsis following dark adaptation or light acclimation.

(A) ECS measurements after a 20 s high light pulse on dark adapted leaves (grey background) and light acclimated leaves (5 min 150 µE m⁻² s⁻¹, actinic light). The relaxation kinetics of ECS were measured at 520–546 nm after a 20 s pulse of actinic light at 1100 µmol photons m⁻² s⁻¹ (LEF + CEF stimulating, white boxes) or far red light (CEF stimulating, red boxes). Averages are shown for between five and seven independent measurements ± s.d. (B) Relative amount of CEF as a function of total electron flow calculated from the data in (A). Figure 5—figure supplement 1 shows a comparable experiment preformed with DCMU rather than far red light to drive PSI activity. (C) The response to high light in dark adapted leaves (grey background) and light acclimated leaves (5 min 150 µE m⁻² s⁻¹, actinic light, white background) was measured by detecting chlorophyll fluorescence and P700 oxidation with saturating pulses following 20 s of actinic light at 1100 µmol photons m⁻² s⁻¹. Measurements are averages ± s.e. of 6–12 replicates of the following genotypes: black, wt; white, fnr1; orange, fnr1-ZmFNR1; cyan, fnr1-ZmFNR2; purple, fnr1-ZmFNR3. Photosystem I parameters (quantum efficiency, ФI; donor limitation, Y(ND); acceptor limitation, Y(NA)) and photosystem II parameters (quantum efficiency, ФII; non-photochemical quenching, NPQ; photochemical quenching, qL). p-value significance in Tukey post hoc analysis of ANOVA is indicated as 0 ‘***’, 0.001 ‘**’ 0.01 ‘*’ 0.05 ‘.’. Unless indicated by bars, stars indicate variation from all other genotypes. Representative P700 and ECS traces are shown in Figure 5—figure supplement 2.

Figure 5—figure supplement 1. DCMU inhibition of PSII for measurement of cyclic electron flow in Arabidopsis.

Electrochromic band shift (ECS) assay to compare generation of ΔpH without (stimulation of both photosystems) or with 20 µM DCMU (inhibition of photosystem II). (A) ECS measurements on leaves from wild-type, fnr1, and fnr1 plants expressing either ZmFNR1, ZmFNR2, or ZmFNR3. The relaxation kinetics of ECS were measured at 520–546 nm after a 20 s pulse of actinic light at 1100 µE m⁻² s⁻¹ in leaf discs that had been infiltrated with 0.4 M manitol (white boxes) or 200 µM DCMU in 0.4 M manitol (black boxes). Leaves were either dark adapted and then subject to the illuminating pulse (left) or light acclimated at 150 µE m-2 s⁻¹ actinic for 5 min (right) before measurement of ECS. Averages are shown for three independent measurements with standard deviation. (B) Relative amount of CEF as a function of total electron flow calculated from the data in (A) following dark adaptation (left) and light acclimation (right).

Figure 5—figure supplement 2. Raw data from ECS and PSI responses to high light treatment.

(A) Representative normalised ECS traces following high light exposure to either red light (filled circles) or far red light (empty circles) after dark adaptation (left) and light acclimation at 150 µmol photons m⁻² s⁻¹ actinic light (right). Bars above graphs indicate the end of high light treatment and transition to dark relaxation. Typical traces from six to seven measurements on independent plants are shown. Genotypes from top to bottom: black, wt; grey, fnr1; orange, fnr1-ZmFNR1; cyan, fnr1-ZmFNR2; purple, fnr1-ZmFNR3. (B) P700 traces on high light exposure following dark adaptation (left) and light acclimation at 150 µmol photons m⁻² s⁻¹ actinic light (right). Black arrow indicates start of illumination at 1100 µmol photons m⁻² s⁻¹, and yellow arrows indicate application of saturating pulses. Data presented in Figure 5B is only calculated from pulses applied 20 s after the start of the high light treatment. Typical traces from 6 to 12 measurements on independent plants are shown. Genotypes coloured as for part (A). Values of P_m and their statistical analysis are provided in Supplementary file 2f.