Antisense reductions in the PsbO protein of photosystem II leads to decreased quantum yield but similar maximal photosynthetic rates - PubMed (original) (raw)
Antisense reductions in the PsbO protein of photosystem II leads to decreased quantum yield but similar maximal photosynthetic rates
Simon A Dwyer et al. J Exp Bot. 2012 Aug.
Abstract
Photosystem (PS) II is the multisubunit complex which uses light energy to split water, providing the reducing equivalents needed for photosynthesis. The complex is susceptible to damage from environmental stresses such as excess excitation energy and high temperature. This research investigated the in vivo photosynthetic consequences of impairments to PSII in Arabidopsis thaliana (ecotype Columbia) expressing an antisense construct to the PsbO proteins of PSII. Transgenic lines were obtained with between 25 and 60% of wild-type (WT) total PsbO protein content, with the PsbO1 isoform being more strongly reduced than PsbO2. These changes coincided with a decrease in functional PSII content. Low PsbO (less than 50% WT) plants grew more slowly and had lower chlorophyll content per leaf area. There was no change in content per unit area of cytochrome b6f, ATP synthase, or Rubisco, whereas PSI decreased in proportion to the reduction in chlorophyll content. The irradiance response of photosynthetic oxygen evolution showed that low PsbO plants had a reduced quantum yield, but matched the oxygen evolution rates of WT plants at saturating irradiance. It is suggested that these plants had a smaller pool of PSII centres, which are inefficiently connected to antenna pigments resulting in reduced photochemical efficiency.
Figures
Fig. 1.
(A) Relationship between total leaf PsbO protein content and functional (oxygen-evolving) PSII centres for a number of independent T2 antisense PsbO lines (r 2 = 0.79, P < 0.0001). PsbO protein content was determined on a leaf area basis through Western blotting relative to a dilution series of a WT standard extraction. Functional PSII was determined from oxygen yield per single turnover flash (see Materials and Methods). (B) Resolution of the two PsbO isoforms from isolated thylakoids of a WT and two low PsbO transgenic lines. Gels were loaded on a chlorophyll basis (note from Table 1 that the chlorophyll content per leaf area in low PsbO plants is around half that of WT). The PsbO line 8 sample was run on the same gel but not on the adjacent lane. (C) The proportion of the total PsbO band intensity accounted for by the PsbO1 isoform. Shown are the averages and standard errors for three biological replicates of WT (mean PSII content 1.11 µmol m–2) and low PsbO (lines #8 and #15, mean PSII content 0.47 µmol m–2). ***, significant difference (P = 0.0001).
Fig. 2.
Changes in the dark adapted maximum quantum yield from chlorophyll fluorescence (FV/FM) on the basis of PsbO protein content determined by Western blot (A) and on the basis of functional PSII content determined by the oxygen yield from single turnover flashes (B). The two data sets partially overlap but not all plants that had PsbO content measured were subsequently measured for functional PSII centres, and vice versa.
Fig. 3.
Examples of phenotypic differences between WT (A, C, E, G) and low PsbO (B, D, F, H) plants: (A, B) growth differences, (C, D) transverse leaf sections (magnification ×200), (E–H) chloroplasts from palisade cells. Refer to Table 1 for quantifications from microscopy images. Bars, 200 µm (C, D), 2 µm (E, F), 0.4 µm (G, H) (this figure is available in colour at JXB online).
Fig. 4.
Relationship between PsbO leaf protein content and major proteins representing the key steps in photosynthesis. PsbD (PSII), PsaD (PSI), Rieske FeS (cytochrome b6f), and AtpC (ATP synthase) were determined by Western blotting. Total Rubisco catalytic sites was determined on a different set of plants by14C-CABP binding. The ∆A max parameter is proportional to total photo-oxidizable P700 content per area. It is included to give physiological support to the change in PSI content indicated by PsaD. Solid lines indicate a statistically significant relationship: for PsbD the regression equation is y = 1.01_x_; for PsaD,y = 0.58 + 21.28; for ∆Amax,y = 0.02_x_ + 1.94; for Rubisco, y_= 0.04_x + 13.64. Note that the slope of the regression line for PsaD is approximately half of that for PsbD.
Fig. 5.
Blue native polyacrylamide gel electrophoresis of isolated thylakoid membranes from a WT and a low PsbO plant. Gels were loaded on an equivalent chlorophyll basis (4 µg). The image shows the original Blue native gel and the same gel after further staining with Coomassie blue. Molecular weights are indicated with a High Molecular Weight Calibration Kit for native electrophoresis (GE Healthcare). Band identifications are taken from Järvi_et al._ (2011) (this figure is available in colour at_JXB_ online).
Fig. 6.
Gross oxygen evolution per leaf area measured at approximately 1000 µbar CO2 for WT plants (>0.85 µmol m–2 functional PSII centres, n = 5) and antisense mutants with low PSII functional centres (<0.6 µmol m–2, n = 8) PSII functional centres. Gross oxygen evolution rates were calculated relative to the signal drift in the dark, which was taken to be mitochondrial respiration. Solid lines are a fit of the average points to the model given in Equation 1. Model parameters for WT plants are E Omax = 19.25, ठ = 0.93, ΦO = 0.40. For low PsbO plants, E Omax =19.56, ठ = 0.85, ΦO = 0.21.
Fig. 7.
Parameters from light response curves as a function of PSII functional centres. (A) The quantum yield for oxygen evolution, determined from the initial slope of light response curves. (B) The maximum rate of gross oxygen evolution, averaged from the three highest irradiances. There is a significant correlation between PSII functional centres and quantum yield (r 2 = 0.67, P = 0.0001).
Fig. 8.
Maximum oxygen evolution rates per leaf PSII content as a function of the PSII to cytochrome b6f ratio, for Arabidopsis, spinach, and pea. Open symbols are values calculated from other studies which have measured maximum electron transport-limited photosynthetic rate at 25 °C, total PSII centres and cytochrome b6f content: spinach grown under various nitrogen nutrition (Evans and Terashima, 1987), irradiance (Terashima and Evans, 1988), and temperature (Yamori et al., 2008); and pea grown under various irradiance (Evans, 1987b). For atrazine binding, measurements were converted to the equivalent functional PSII measurements obtained through flash yield, by dividing by 1.14 (Chow and Anderson, 1987; Chow et al., 1989). For Yamori et al.(2008) PSII centres were estimated from the chlorophyll content and cytochrome f content taken from Yamori et al. (2005). The line is a fit to the _Arabidopsis_data only (r 2 = 0.93).
Fig. 9.
Fluorescence parameters measured concurrently with oxygen evolution rates. (A) The quantum yield of PSII, ΦPSII, given by 
(Genty et al., 1989). (B′) The quantum yield of non-photochemical quenching, ΦNPQ, given by 
(Hendrickson et al., 2004). (C) The quantum yield of fluorescence and constitutive thermal energy dissipation, Φf,D, given by 
(Hendrickson et al., 2004). (D) The proportion of ‘open’ PSII centres, qL, given by 
(Kramer et al., 2004b), where F'O was calculated according to Oxborough and Baker (1997b). (E) An estimation of the proportion of absorbed irradiance used for water splitting at PSII, X, calculated according to Equation 3. The dashed line at 0.5 shows the expected value if PSII and PSI centres were using an equal proportion of the absorbed light.
References
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- Anderson JM. 1992. Cytochrome b6f complex: dynamic molecular organization, function and acclimation Photosynthesis Research 34 341–357 -PubMed
- Anderson JM, Chow WS, Goodchild DJ. 1988. Thylakoid membrane organisation in sun/shade acclimation Functional Plant Biology 15 11–26
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