Da-Quan Xu

Reversible association of ribulose-1, 5-bisphosphate carboxylase/oxygenase activase with the thylakoid membrane depends upon the ATP level and pH in rice without heat stress

Ribulose-1, 5-bisphosphate carboxylase/oxygenase (Rubisco) activase (RCA) in the thylakoid membrane (TM) has been shown to play a role in protection and regulation of photosynthesis under moderate heat stress. However, the physiological significance of RCA bound to the TM (TM–RCA) without heat stress remains unknown. In this study, it is first shown, using experiments in vivo, that the TM–RCA varies in rice leaves at different development stages, under different environmental conditions, and in a rice mutant. Furthermore, it is shown that the amount of TM–RCA always increased when the Rubisco activation state and the pH gradient across the TM (ΔpH) decreased. It was then demonstrated in vitro that the RCA bound dynamically to TM and the amount of TM–RCA increased during Rubisco activation. A high level of ATP and a high pH value promoted the dissociation of RCA from the TM. Both the RCA association with and dissociation from the TM showed conformational changes related to the ATP level or pH as indicated by the changes in fluorescence intensity of 1-anilinonaphthalene-8-sulphonic acid (ANS) binding to RCA. These results suggest that the reversible association of RCA with the TM is ATP and pH (or ΔpH) dependent; it might be involved in the RCA activation of Rubisco, in addition to the previously discovered role in the protection and regulation of photosynthesis under heat stress.

Light intensity-dependent reversible down-regulation and irreversible damage of PSII in soybean leaves

Abstract Light intensity dependence of reversible down-regulation and irreversible damage of PSII was studied by determination of chlorophyll fluorescence, PSII electron transport, D1 protein level, and oligomeric state of PSII complexes. After illumination with moderate light (700 μmol photons m−2 s−1, saturated for photosynthesis) for 3 h, all of the light-saturated PSII electron transport rate, D1 protein level and proportion of PSII dimer in total PSIIs in soybean leaves grown at 250–300 μmol m−2 s−1 had no significant change. Although PSII photochemical efficiency (Fv/Fm) declined significantly, it could recover completely after 4 h in the dark. After illumination with strong light (2000 μmol m−2 s−1) for 3 h, however, the significant decreases in all parameters above were observed, and Fv/Fm could not restore completely after 4 h in the dark. Moreover, the chlorophyll fluorescence parameter F685/F735 measured at 77 K dropped significantly in thylakoids from all soybean leaves illuminated at 700 (L2), 1200 (L4) and 2000 (L7) μmol m−2 s−1 for 3 h. After subsequent dark recovery for 3 h, F685/F735 in thylakoids from soybean leaves illuminated with L2 could recover to the level of dark control, but not in those from soybean leaves illuminated with L4 and L7. Those results indicate that the illumination with moderate light results in reversible down-regulation of some PSIIs, while illumination with strong light leads to damage of some PSIIs, namely, both reversible down-regulation and irreversible damage of PSII are light intensity-dependent.

Role of light-harvesting complex 2 dissociation in protecting the photosystem 2 reaction centres against photodamage in soybean leaves and thylakoids

The protective role of light-harvesting complex 2 (LHC2) dissociation from photosystem 2 (PS2) complex was explored by the 5′-p-fluorosulfonylbenzoyl adenosine (FSBA, an inhibitor of protein kinase) treatment at saturating irradiance (SI) in soybean leaves and thylakoids. The dissociation of some LHC2s from PS2 complex occurred after SI treatment, but FSBA treatment inhibited the dissociation as demonstrated by analysis of sucrose density gradient centrifugation of thylakoid preparation and low-temperature (77 K) chlorophyll (Chl) fluorescence. A significant increase in F0 and decrease in Fv/Fm occurred after SI, and the two parameters could largely recover to the levels of dark-adapted leaves after subsequent 3 h in the dark, but they could not recover in the FSBA-treated leaves at SI. Neither the electron transport activity of PS2 nor the D1 protein amount in vivo had significant change after SI without FSBA, whereas FSBA treatment at SI could result in significant decreases in both the PS2 electron transport activity and the D1 protein amount. When thylakoids instead of leaves were used, the PS2 electron transport activity and the D1 protein amount declined more after SI with FSBA than without FSBA. The phosphorylation level of PS2 core proteins increased, while the phosphorylation level of LHC2 proteins was reduced after SI. Also, the phosphorylation of PS2 core proteins could be greatly inhibited by the FSBA treatment at SI. Hence in soybean leaf the LHC2 dissociation is an effective strategy protecting PS2 reaction centres against over-excitation and photodamage by reducing the amount of photons transferred to the centres under SI, and the phosphorylation of PS2 core proteins plays an important role in the dissociation.

Light-harvesting regulation from leaf to molecule with the emphasis on rapid changes in antenna size

In the sunlight-fluctuating environment, plants often encounter both light-deficiency and light-excess cases. Therefore, regulation of light harvesting is absolutely essential for photosynthesis in order to maximize light utilization at low light and avoid photodamage of the photosynthetic apparatus at high light. Plants have developed a series of strategies of light-harvesting regulation during evolution. These strategies include rapid responses such as leaf movement and chloroplast movement, state transitions, and reversible dissociation of some light-harvesting complex of the photosystem II (LHCIIs) from PSII core complexes, and slow acclimation strategies such as changes in the protein abundance of light-harvesting antenna and modifications of leaf morphology, structure, and compositions. This review discusses successively these strategies and focuses on the rapid change in antenna size, namely reversible dissociation of some peripheral light-harvesting antennas (LHCIIs) from PSII core complex. It is involved in protective role and species dependence of the dissociation, differences between the dissociation and state transitions, relationship between the dissociation and thylakoid protein phosphorylation, and possible mechanism for thermal dissipation by the dissociated LHCIIs.

D1 protein phosphorylation/dephosphorylation alone has no effect on the electron transport activity of photosystem II in soybean leaves

Abstract In order to explore the relationship between the phosphorylation of D1 protein and the function of PSII reaction center, the changes in chlorophyll fluorescence parameters, PSII electron transport activity and D1 protein amount were observed after dephosphorylation of phosphorylated D1 proteins caused by FSBA (5′- p -fluorosulfonylbenzoyl adenosine, an inhibitor of protein kinase) treatment. Also, phosphorylated and non-phosphorylated D1 proteins (D1* and D1, respectively) were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western Blotting. The following results were obtained. (1) D1* could be up to 74% of the total D1 proteins in dark-adapted soybean leaves and could be completely dephosphorylated by FSBA (1 mM) treatment for 2 h. (2) Dephosphorylation of D1* resulted in neither substantial net loss of D1 proteins nor the significant changes in value of chlorophyll fluorescence parameters. (3) The electron transport activity of PS II (H 2 O→1,4-BQ) was not changed significantly after D1* was completely dephosphorylated. Based on these results, it is concluded that D1 protein phosphorylation/dephosphorylation alone has no significant effect on the function of PS II reaction center in soybean leaves.

Different Mechanisms for Photosystem 2 Reversible Down-Regulation in Pumpkin and Soybean Leaves Under Saturating Irradiance

After saturating irradiation for 3 h (SI), the original fluorescence F0 increased while the photosystem 2 (PS2) photochemical efficiency (Fv/Fm) declined significantly. These parameters could largely recover to the levels of dark-adapted leaves after 3 h of subsequent dark recovery. No net loss of the D1 proteins occurred after SI. Soybean and pumpkin leaves had different responses to SI. Low temperature fluorescence parameters, F685 and F685/F735, decreased significantly in soybean leaves but not in pumpkin leaves. Part of the light-harvesting complex LHC2 dissociated from PS2 complexes in soybean leaves but not in pumpkin leaves, as shown by sucrose density gradient centrifugation and SDS-PAGE. The photon-saturated PS2 electron transport activity declined significantly in pumpkin thylakoids but not in soybean thylakoids. In addition, a large amount of phosphorylated D1 proteins was found in dark-adapted soybean leaves but not in dark-adapted pumpkin leaves. Hence at excessive irradiance soybean and pumpkin have the same protective strategy against photo-damage, reversible down-regulation of PS2, but two different mechanisms, namely the reversible down-regulation is related to the dissociation of LHC2 in soybean leaves but not in pumpkin leaves.

No Down-Regulation of Photosynthesis in the Offspring of Rice Grown Under Free-Air CO2 Enrichment (FACE)

Rising CO2 increases photosynthesis in C3 plants owing to the increase of its substrate concentration and inhibition of photorespiration. After long-term exposure to elevated CO2, however, the stimulatory effect decreases gradually in many C3 plants so that the net photosynthetic rate (P n) is lower than that in plants grown in ambient air when measured at the same CO2 concentration. This phenomenon, the so called down-regulation of photosynthesis, is often reported in CO2-enriched current generation plants. It has not been known whether the down-regulation is preserved or eliminated in the offspring from seeds of plants grown at elevated CO2. In Chinese free-air CO2 enrichment experiments the leaf Pn was significantly lower in CO2-enriched rice but not in the CO2-enriched rice offspring grown in ambient air when measured at comparable CO2 concentrations, indicating that no down-regulation of photosynthesis occurred in the offspring of CO2-enriched rice.

Saturating Irradiance-Induced Photoinhibition without Monomerisation of Photosystem 2 Dimer in Soybean Leaves

The oligomeric state of photosystem 2 (PS2) complex in soybean leaves treated with saturating irradiance was studied by non-denaturing polyacrylamide gel electrophoresis (PAGE) and gel filtration chromatography. PS2 dimers resolved by non-denaturing PAGE accounted for about 75 % of total PS2 complex and there was no significant difference in the ratio of PS2 dimer to monomer between samples from saturating irradiance-treated and fully dark-adapted leaves. Furthermore, BBY particles were resolved into four chlorophyll-enriched fractions by gel filtration chromatography. From their molecular masses and protein components, these fractions were deduced to be PS2 dimer, PS2 monomer, oligomeric light-harvesting complex 2 (LHC2), and monomeric LHC2. Also, no change in the proportion of PS2 dimer in total PS2 was observed in the granal region of thylakoid membranes from soybean leaves after saturating irradiation. Hence the dimer is the predominant natural form of PS2 in vivo and no monomerisation of PS2 dimer occurs during saturating irradiance-induced photoinhibition in soybean leaves.

Phosphorylation/dephosphorylation of D1 protein alone has no effect on the electron transport activity of photosystem II in soybean leaves

D1 protein, as a core component of the photosystem II (PSII) reaction center complex, can be reversibly phosphorylated/dephosphorylated in higher plants. The phosphorylated D1 protein (D1*) and non-phosphorylated D1 protein (D1), having slightly different mobilities on denaturing polyacrylamide gels, can be resolved by using SDS-PAGE and Western Blotting [Callahan et al, J Biol Chem 1990, 265: 15357-15360]. The relationship between the phosphorylation of D1 protein and the electron transport activity of PSII was investigated by changing the phosphorylation level of D1 proteins with an inhibitor of protein kinase, 5?-p-fluorosulfonylbenzoyl adenosine (FSBA). The following results were obtained. (1) D1* could be up to 74% of total D1 proteins in fully dark-adapted soybean leaves and could be completely dephosphorylated by FSBA (1 mM) treatment for 2 hours. (2) Dephosphorylation of D1* resulted in neither substantial net loss of D1 proteins nor the significant changes of chlorophyll a fluorescence parameters. (3) The electron transport activity of PS II (H2O ® 1,4-BQ) had no significant change after D1* was completely dephosphorylated. Based on these results, it is concluded that phosphorylation/dephosphorylation of D1 protein alone has no significant effect on the function of PS II reaction centers in soybean leaves.