Action spectrum of the redox state of the plastoquinone pool defines its function in plant acclimation.

The plastoquinone (PQ) pool mediates electron flow and regulates photoacclimation in plants. Here we report the action spectrum of the redox state of the PQ pool in Arabidopsis thaliana, showing that 470-500, 560 or 650-660 nm light favors Photosystem II (PSII) and reduces the PQ pool, whereas 420-440, 520 or 690 nm light favors Photosystem I (PSI) and oxidizes PQ. These data were used to construct a model predicting the redox state of PQ from the spectrum of any polychromatic light source. Moderate reduction of the PQ pool induced transition to light state 2, whereas state 1 required highly oxidized PQ. In low-intensity PSI light, PQ was more oxidized than in darkness and became gradually reduced with light intensity, while weak PSII light strongly reduced PQ. Natural sunlight was found to favor PSI, which enables plants to use the redox state of the PQ pool as a measure of light intensity.

Measurement of the redox state of the plastoquinone pool in cyanobacteria.

Here, we developed a method for measuring the in vivo redox state of the plastoquinone (PQ) pool in the cyanobacteria Synechocystis sp. PCC 6803. Cells were illuminated on a glass fiber filter, PQ was extracted with ethyl acetate and determined with HPLC. Control samples with fully oxidized and reduced photoactive PQ pool were prepared by far-red and high light treatments, respectively, or by blocking the photosynthetic electron transfer chemically before or after PQ in moderate light. The photoactive pool comprised 50% of total PQ. We find that the PQ pool of cyanobacteria behaves under light treatments qualitatively similarly as in plant chloroplasts, is less reduced during growth under high than under ambient CO2 and remains partly reduced in darkness.

Plastoquinol as a singlet oxygen scavenger in photosystem II.

It has been found that in Chlamydomonas reinhardtii cells, under high-light stress, the level of reduced plastoquinone considerably increases while in the presence of pyrazolate, an inhibitor of plastoquinone and tocopherol biosynthesis, the content of reduced plastoquinone quickly decreases, similarly to alpha-tocopherol. In relation to chlorophyll, after 18 h of growth under low light with the inhibitor, the content of alpha-tocopherol was 22.2 mol/1000 mol chlorophyll and that of total plastoquinone (oxidized and reduced) was 19 mol/1000 mol chlorophyll, while after 2 h of high-light stress the corresponding amounts dropped to 6.4 and 6.2 mol/1000 mol chlorophyll for alpha-tocopherol and total plastoquinone, respectively. The degradation of both prenyllipids was partially reversed by diphenylamine, a singlet oxygen scavenger. It was concluded that plastoquinol, as well as alpha-tocopherol is decomposed under high-light stress as a result of a scavenging reaction of singlet oxygen generated in photosystem II. The levels of both alpha-tocopherol and of the reduced plastoquinone are not affected significantly in the absence of the inhibitor due to a high turnover rate of both prenyllipids, i.e., their degradation is compensated by fast biosynthesis. The calculated turnover rates under high-light conditions were twofold higher for total plastoquinone (0.23 nmol/h/ml of cell culture) than for alpha-tocopherol (0.11 nmol/h/ml). We have also found that the level of alpha-tocopherolquinone, an oxidation product of alpha-tocopherol, increases as the alpha-tocopherol is consumed. The same correlation was also observed for gamma-tocopherol and its quinone form. Moreover, in the presence of pyrazolate under low-light growth conditions, the synthesis of plastoquinone-C, a hydroxylated plastoquinone derivative, was stimulated in contrast to plastoquinone, indicating for the first time a functional role for plastoquinone-C. The presented data also suggest that the two plastoquinones may have different biosynthetic pathways in C. reinhardtii.

The electrical and spectroscopic properties of planar asymmetrical membranes incorporating chlorophyll a and plastoquinone-9. Influence of surface charges on electron transfer.

We studied the influence of surface charges on the efficiencies of electron transfer between a donor molecule, chlorophyll a (Chla), and an acceptor molecule, plastoquinone-9 (PQ-9), asymmetrically incorporated into a phospholipid matrix built from phosphatidylethanolamine, phosphatidylserine, and dimethyldistearylammonium bromide. Membrane conductance and capacitance measurements, as well as fluorescence emission experiments, were performed on bilayers containing positive or negative surface charges. The conductance of the bilayers showed important increases upon illumination of the Chla, this effect being observed only when both the donor and the acceptor molecules were present within the bilayer. This suggested that an electron transfer between Chla and PQ-9 occurred. The same kind of behaviour was observed with the membrane capacitance, but the amplitude of the effect was smaller. The results showed that an electric field gradient favorably oriented to promote electron transfer from Chla to PQ-9 maximized the electron transfer between these two molecules. However, both the membrane resistance and capacitance were permanently modified when illumination was stopped. On the other hand, the fluorescence results showed that for the range of surface charges covered, the position of the maximum of absorption was found unchanged around 675 nm and the intensity of fluorescence was almost constant, of the order of 7 x 10(6)-8 x 10(6) photons.s-1. This suggested that Chla was embedded as microdomains within the bilayer. The results presented here were also compared with what is known on the organization of the donor and acceptor molecules within the reaction centres of photosynthetic bacteria.

The mechanism of UV-A radiation-induced inhibition of photosystem II electron transport studied by EPR and chlorophyll fluorescence.

The UV-A (320-400 nm) component of sunlight is a significant damaging factor of plant photosynthesis, which targets the photosystem II complex. Here we performed a detailed characterization of UV-A-induced damage in photosystem II membrane particles using EPR spectroscopy and chlorophyll fluorescence measurements. UV-A irradiation results in the rapid inhibition of oxygen evolution accompanied by the loss of the multiline EPR signal from the S(2) state of the water-oxidizing complex. Gradual decrease of EPR signals arising from the Q(A)(-)Fe(2+) acceptor complex, Tyr-D degrees, and the ferricyanide-induced oxidation of the non-heme Fe(2+) to Fe(3+) is also observed, but at a significantly slower rate than the inhibition of oxygen evolution and of the multiline signal. The amplitude of Signal II(fast), arising from Tyr-Z degrees in the absence of fast electron donation from the Mn cluster, was gradually increased during the course of UV-A treatment. However, the amount of functional Tyr-Z decreased to a similar extent as Tyr-D as shown by the loss of amplitude of Signal II(fast) that could be measured in the UV-A-treated particles after Tris washing. UV-A irradiation also affects the relaxation of flash-induced variable chlorophyll fluorescence. The amplitudes of the fast (600 micros) and slow (2 s) decaying components, assigned to reoxidation of Q(A)(-) by Q(B) and by recombination of (Q(A)Q(B))(-) with donor side components, respectively, decrease in favor of the 15-20 ms component, which reflects PQ binding to the Q(B) site. In the presence of DCMU, the fluorescence relaxation is dominated by a 1 s component due to recombination of Q(A)(-) with the S(2) state. After UV-A radiation, this is partially replaced by a much faster component (30-70 ms) arising from recombination of Q(A)(-) with a stabilized intermediate PSII donor, most likely Tyr-Z degrees. It is concluded that the primary damage site of UV-A irradiation is the catalytic manganese cluster of the water-oxidizing complex, where electron transfer to Tyr-Z degrees and P(680)(+) becomes inhibited. Modification and/or inactivation of the redox-active tyrosines and the Q(A)Fe(2+) acceptor complex are subsequent events. This damaging mechanism is very similar to that induced by the shorter wavelength UV-B (280-320) radiation, but different from that induced by the longer wavelength photosynthetically active light (400-700 nm).