The roles of photorespiration and alternative electron acceptors in the responses of photosynthesis to elevated temperatures in cowpea

Abstract

We explored the effects, on photosynthesis in cowpea (Vigna unguiculata) seedlings, of high temperature and light-environmental stresses that often co-occur under field conditions and can have greater impact on photosynthesis than either by itself. We observed contrasting responses in the light and carbon assimilatory reactions, whereby in high temperature, the light reactions were stimulated while CO₂ assimilation was substantially reduced. There were two striking observations. Firstly, the primary quinone acceptor (Q_A ), a measure of the regulatory balance of the light reactions, became more oxidized with increasing temperature, suggesting increased electron sink capacity, despite the reduced CO₂ fixation. Secondly, a strong, O₂ -dependent inactivation of assimilation capacity, consistent with down-regulation of rubisco under these conditions. The dependence of these effects on CO₂ , O₂ and light led us to conclude that both photorespiration and an alternative electron acceptor supported increased electron flow, and thus provided photoprotection under these conditions. Further experiments showed that the increased electron flow was maintained by rapid rates of PSII repair, particularly at combined high light and temperature. Overall, the results suggest that photodamage to the light reactions can be avoided under high light and temperatures by increasing electron sink strength, even when assimilation is strongly suppressed.

Keywords: PSII efficiency; Vigna unguiculata; heat stress; high temperature; high-light; net CO2 assimilation.

Figure 1:. Temperature response of photosynthesis under different light intensities and in leaves of different maturity.

A) photosynthetic efficiency (ɸ_II), B) light-induced thylakoid proton motive force (pmf) estimated by the ECSt parameter, C) ATP synthase activity (g_H+), D) non-photochemical quenching (NPQ), E) redox state of primary quinone acceptor (q_L), F) v_H+/LEF and G) Relative change in the v_H+/LEF ratio. Data were normalized to values at 30 °C from Figure 1F. Leaf maturity is indicated by the line type: dashed line, mature leaves (ML, unifoliate leaves from 12 -day old seedlings); solid line, young leaves (YL, unifoliate leaves from 4-day old seedlings). The different colored lines represent measurements at low (300 μmol m⁻² s⁻¹) and high (1000 μmol m⁻² s⁻¹ and 1500 μmol m⁻² s⁻¹) light intensities as indicated in (A). Grey area represents recovery for at least 1 h. Data are from cowpea genotype ‘Yacine’. The temperatures reported are the air temperature in the chamber, which was changed every 2h, although, measurements were made after at least 1 h under each temperature. Data are means of 8–14 replicates from 2–3 independent experiments ±1 S.E.

Figure 2:. Effects of lincomycin on maximum efficiency of PSII (FV/FM) under HL (1500 μmol m⁻² s⁻¹) or combined HL and HT stress.

Starting from 1 h after lincomycin treatment, the kinetics of relative FV/FM are shown for A) GT+HL and B) HT+HL treatments. Values are expressed relative to the initial F_V/F_M values for each day. Leaves were syringe infiltrated with 0.2 g/L lincomycin or deionized water. Dashed lines with hollow markers represent lincomycin treatment and solid lines with filled markers represent mock (water) infiltrated leaves. Line color indicates the temperature and light intensity treatment. Red = HT+HL (high temperature + high-light), black=GT+HL (growth temperature + high-light). For the GT+HL treatment, the temperature was kept constant (30 °C) throughout the 6 h period whereas the temperature for the HT+HL are indicated in the colored boxes above the data points. Data are from young cowpea leaves and the two genotypes (Yacine and 58–77) are used to show the differences in HL sensitivity which is abolished in HT+HL. The means of 4 replicates ±1 S.E are shown.

Figure 3:. Temperature responses of CO2 assimilation-related processes in young leaves of cowpea seedlings under different light intensities.

A) Net CO2 assimilation (A), B) Stomatal conductance (g_s) and C) Intercellular CO₂ concentration (C_i). The different colored lines represent measurements at low (300 μmol m⁻² s⁻¹) and high (1000 μmol m⁻² s⁻¹ and 1500 μmol m⁻² s⁻¹) light intensities as indicated in the legend in Figure 3B. Grey area represents recovery for at least 1 h. Data are from young leaves (YL) of cowpea genotype ‘Yacine’. The temperatures reported are the air temperature in the chamber, which was changed every 2h, although, measurements were made after at least 1 h under each temperature. Data are means ±1 S.E of at least 8 biological replicates from 2–3 independent experiments.

Figure 4:. Effect of temperature on velocity of rubisco for carboxylation (v_c) and oxygenation (v_o) and electron transport rate in cowpea.

A–B) Temperature response of v_c and v_o in leaves of different maturity and under light intensities of A) 300 μmol photons m⁻² s⁻¹ (low light — LL) and B) 1000 μmol photons m⁻² s⁻¹ (high-light — HL). C–D). The temperature response and light intensity dependence of electron transport rate measured by chlorophyll fluorescence (ETR) or calculated from gas exchange (ETR_GE). The colors represent the different parameters as indicated in the y-axes of the leading (far left) figures. Closed symbols = young leaves (YL), open symbols = mature leaves (ML). LL Data are means of 6–8 replicates with error bars being ± 1 S.E. Assumptions used in calculating v_c and v_o are outlined in the Materials and Methods.

Figure 5:. Comparison of responses of net CO₂ assimilation (A) and ETR/4 to internal CO₂ concentration (C_i) in different light intensities, O₂ levels and temperature in young cowpea leaves.

Response of A and ETR/4 against C_i under low light (300 μmol photons m⁻² s⁻¹) at A–B) 30 °C and C–D) 45 °C respectively. Corresponding measurements at high light (1000 μmol photons m⁻² s⁻¹) are shown in E–F and G–H respectively. The circled points represent the first measurements made under ambient CO2 and have been replotted in I&J for clarity. I–L) Effect of O₂ concentration on responses of net CO₂ assimilation (A) and PSII efficiency (ɸ_II) to temperature under different light intensities in ambient CO₂, during initial measurements (I&K) and after exposure to changing CO₂ concentrations (J&L). Data are means ± 1 S.E of 3–10 biological replicates from 1–3 independent experiments. ns, *, **, *** indicate not significant (p>0.05), and significant at p<0.05, p<0.01 and p<0.001 respectively.

Figure 6:. Comparison of responses of net CO₂ assimilation (A) and ETR/4 to internal CO2 concentration (C_i) in different light intensities, O₂ levels and temperature in mature cowpea leaves (ML).

Response of A and ETR/4 against C_i under low light (LL — 300 μmol photons m⁻² s⁻¹) at A–B) 30 °C and C–D) 45 °C respectively. Corresponding measurements at high light (LL — 1000 μmol photons m⁻² s⁻¹) are shown in E–F and G–H respectively. The circled points represent the first measurements made under ambient CO₂ and have been replotted in I&J for clarity. I–L) Effect of O₂ concentration on responses of net CO₂ assimilation (A) and PSII efficiency (ɸ_II) to temperature under different light intensities in ambient CO₂, during initial measurements (I&K) and after exposure to changing CO2 concentrations (J&L). Data are means ± 1 S.E of 3–8 biological replicates from 1–3 independent experiments. ns, *, **, *** indicate not significant (p>0.05), and significant at p<0.05, p<0.01 and p<0.001 respectively.

Figure 7:. HT treated plants exhibit less photo-inhibitory quenching under prolonged dynamic and high light conditions.

Changes in A) ɸ_II, B) Non-photochemical quenching (NPQ_(T)), C) Photo-inhibitory quenching (q_I(T)) and D) the rapidly reversible (photoprotective) quenching (q_E(T)) during a 4-day experiment with corresponding temperature and light intensity (PAR=photosynthetically active radiation — μmol photons m⁻² s⁻¹) changes indicated in the top panel of Figure 7A. The color of the lines corresponds to the colors of the temperature. Dashed line = 58–77, solid lines=Yacine. The suffix after the genotype names indicates the temperature treatment. Data are means of 5–8 replicates from 1–2 independent experiments. Measurements were made at 20 min intervals on days 1 and 4 and at 1 h intervals on days 2 and 3 using the CCD (charge coupled device) cameras in the DEPI (Dynamic Environment Photosynthesis Imager) chamber (1).