The Arabidopsis PsbO2 protein regulates dephosphorylation and turnover of the photosystem II reaction centre D1 protein

The extrinsic photosystem II (PSII) protein of 33 kDa (PsbO), which stabilizes the water-oxidizing complex, is represented in Arabidopsis thaliana (Arabidopsis) by two isoforms. Two T-DNA insertion mutant lines deficient in either the PsbO1 or the PsbO2 protein were retarded in growth in comparison with the wild type, while differing from each other phenotypically. Both PsbO proteins were able to support the oxygen evolution activity of PSII, although PsbO2 was less efficient than PsbO1 under photoinhibitory conditions. Prolonged high light stress led to reduced growth and fitness of the mutant lacking PsbO2 as compared with the wild type and the mutant lacking PsbO1. During a short period of treatment of detached leaves or isolated thylakoids at high light levels, inactivation of PSII electron transport in the PsbO2-deficient mutant was slowed down, and the subsequent degradation of the D1 protein was totally inhibited. The steady-state levels of in vivo phosphorylation of the PSII reaction centre proteins D1 and D2 were specifically reduced in the mutant containing only PsbO2, in comparison with the mutant containing only PsbO1 or with wild-type plants. Phosphorylation of PSII proteins in vitro proceeded similarly in thylakoid membranes from both mutants and wild-type plants. However, dephosphorylation of the D1 protein occurred much faster in the thylakoids containing only PsbO2. We conclude that the function of PsbO1 in Arabidopsis is mostly in support of PSII activity, whereas the interaction of PsbO2 with PSII regulates the turnover of the D1 protein, increasing its accessibility to the phosphatases and proteases involved in its degradation.     

Acclimation of the Photosynthetic Apparatus to Growth Irradiance in a Mutant Strain of Synechococcus Lacking Iron Superoxide Dismutase

The acclimation of the photosynthetic apparatus to growth irradiance in a mutant strain of Synechococcus sp. PCC 7942 lacking detectable iron superoxide dismutase activity was studied. The growth of the mutant was inhibited at concentrations of methyl viologen 4 orders of magnitude smaller than those required to inhibit the growth of the wild-type strain. An increased sensitivity of photosynthetic electron transport near photosystem I (PSI) toward photooxidative stress was also observed in the mutant strain. In the absence of methyl viologen, the mutant exhibited similar growth rates compared with those of the wild type, even at high growth irradiance (350 [mu]E m-2 s-1) where chronic inhibition of photosystem II (PSII) was observed in both strains. Under high growth irradiance, the ratios of PSII to PSI and of [alpha]-phycocyanin to chlorophyll a were less than one-third of the values for the wild type. In both strains, cellular contents of chlorophyll a, [alpha]-phycocyanin, and [beta]-carotene, as well as the length of the phycobilisome rods, declined with increasing growth irradiance. Only the cellular content of the carotenoid zeaxanthin seemed to be independent of growth irradiance. These results suggest an altered acclimation to growth irradiance in the sodB mutant in which the stoichiometry between PSI and PSII is adjusted to compensate for the loss of PSI efficiency occurring under high growth irradiance. Similar shortening of the phycobilisome rods in the sodB mutant and wild-type strain suggest that phycobilisome rod length is regulated independently of photosystem stoichiometry.     

Acclimation of Arabidopsis thaliana to the light environment: regulation of chloroplast composition

The regulation by light of the composition of the photosynthetic apparatus was investigated in Arabidopsis thaliana (L.) Heynh. cv. Landsberg erecta. When grown in high- and low-irradiance white light, wild-type plants and photomorphogenic mutants showed large differences in their maximum photosynthetic rate and chlorophyll a/b ratios; such changes were abolished by growth in red light. Photosystem I (PSI) and PSII levels were measured in wild-type plants grown under a range of light environments; the results indicate that regulation of photosystem stoichiometry involves the specific detection of blue light. Supplementing red growth lights with low levels of blue light led to large increases in PSII content, while further increases in blue irradiance had the opposite effect; this latter response was abolished by the hy4 mutation, which affects certain events controlled by a blue-light receptor. Mutants defective in the phytochrome photoreceptors retained regulation of photosystem stoichiometry. We discuss the results in terms of two separate responses controlled by blue-light receptors: a blue-high-fluence response which controls photosystem stoichiometry; and a blue-low-fluence response necessary for activation of such control. Variation in the irradiance of the red growth light revealed that the blue-high-fluence response is attenuated by red light; this may be evidence that photosystem stoichiometry is controlled not only by photoreceptors, but also by photosynthetic metabolism.Abbreviations BHF blue-high-fluence - BLF blue-low-fluence - Chl chlorophyll - FR far-red light - LHCII light-harvesting complex of PSII - P_max maximum photosynthetic rate - R red light - Rubisco ribulose-1,5-bisphosphate carboxylase/oxygenase This work was supported by Natural Environment Research Council Grant No. GR3/7571A. We would like to thank H. Smith (Botany Department, University of Leicester) and E. Murchie (INRA, Versailles) for helpful discussions.     

A small zinc finger thylakoid protein plays a role in maintenance of photosystem II in Arabidopsis thaliana

This work identifies LOW QUANTUM YIELD OF PHOTOSYSTEM II1 (LQY1), a Zn finger protein that shows disulfide isomerase activity, interacts with the photosystem II (PSII) core complex, and may act in repair of photodamaged PSII complexes. Two mutants of an unannotated small Zn finger containing a thylakoid membrane protein of Arabidopsis thaliana (At1g75690; LQY1) were found to have a lower quantum yield of PSII photochemistry and reduced PSII electron transport rate following high-light treatment. The mutants dissipate more excess excitation energy via nonphotochemical pathways than wild type, and they also display elevated accumulation of reactive oxygen species under high light. After high-light treatment, the mutants have less PSII-light-harvesting complex II supercomplex than wild-type plants. Analysis of thylakoid membrane protein complexes showed that wild-type LQY1 protein comigrates with the PSII core monomer and the CP43-less PSII monomer (a marker for ongoing PSII repair and reassembly). PSII repair and reassembly involve the breakage and formation of disulfide bonds among PSII proteins. Interestingly, the recombinant LQY1 protein demonstrates a protein disulfide isomerase activity. LQY1 is more abundant in stroma-exposed thylakoids, where key steps of PSII repair and reassembly take place. The absence of the LQY1 protein accelerates turnover and synthesis of PSII reaction center protein D1. These results suggest that the LQY1 protein may be involved in maintaining PSII activity under high light by regulating repair and reassembly of PSII complexes.     

Towards understanding the functional difference between the two PsbO isoforms in Arabidopsis thaliana—insights from phenotypic analyses of psbo knockout mutants

The extrinsic PsbO subunit of the water-oxidizing photosystem II (PSII) complex is represented by two isoforms in Arabidopsis thaliana, namely PsbO1 and PsbO2. Recent analyses of psbo1 and psbo2 knockout mutants have brought insights into their roles in photosynthesis and light stress. Here we analyzed the two psbo mutants in terms of PsbOs expression pattern, organization of PSII complexes and GTPase activity. Both PsbOs are present in wild-type plants, and their expression is mutually controlled in the mutants. Almost all PSII complexes are in the monomeric form not only in the psbo1 but also in the psbo2 mutant grown under high-light conditions. This results either from an enhanced susceptibility of PSII to photoinactivation or from malfunction of the repair cycle. Notably, the psbo1 mutant displays such problems even under growth-light conditions. These results together with the finding that PsbO2 has a threefold higher GTPase activity than PsbO1 have significance for the turnover of the PSII D1 subunit in Arabidopsis.     

Light-intensity-dependent expression of Lhc gene family encoding light-harvesting chlorophyll-a/b proteins of photosystem II in Chlamydomonas reinhardtii

Excessive light conditions repressed the levels of mRNAs accumulation of multiple Lhc genes encoding light-harvesting chlorophyll-a/b (LHC) proteins of photosystem (PS)II in the unicellular green alga, Chlamydomonas reinhardtii. The light intensity required for the repression tended to decrease with lowering temperature or CO(2) concentration. The responses of six LhcII genes encoding the major LHC (LHCII) proteins and two genes (Lhcb4 and Lhcb5) encoding the minor LHC proteins of PSII (CP29 and CP26) were similar. The results indicate that the expression of these Lhc genes is coordinately repressed when the energy input through the antenna systems exceeds the requirement for CO(2) assimilation. The Lhc mRNA level repressed under high-light conditions was partially recovered by adding the electron transport inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea, suggesting that redox signaling via photosynthetic electron carriers is involved in the gene regulation. However, the mRNA level was still considerably lower under high-light than under low-light conditions even in the presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea. Repression of the Lhc genes by high light was prominent even in the mutants deficient in the reaction center(s) of PSII or both PSI and PSII. The results indicate that two alternative processes are involved in the repression of Lhc genes under high-light conditions, one of which is independent of the photosynthetic reaction centers and electron transport events.     

The role of ΔpH-dependent dissipation of excitation energy in protecting photosystem II against light-induced damage in Arabidopsis thaliana

The Arabidopsis thaliana subunit PsbS of photosystem II (PSII) is essential for the non-photochemical quenching of chlorophyll fluorescence and thus for ΔpH-dependent energy dissipation (qE). As a result of the excision of an En-transposon, a frameshift mutation in the psbS gene was obtained, which results in the complete absence of the PsbS protein and of qE. Two-dimensional gel analyses of thylakoid membranes indicated that the depletion of PsbS has no effect on PSII composition, excluding a structural role for PsbS in the organization of the PSII antenna. The susceptibility of mutant plants to photoinactivation of PSII was significantly increased during exposure to high light for up to 8 h. Divergence of mutant plants from wild-type levels of photoinactivation were most pronounced during the first 2 h of illumination, while after longer exposure times the rate of PSII inactivation were similar in both genotypes. The increased PSII inactivation in the mutant was not accompanied by an increased rate of D1 protein degradation, and recovery of PSII activity in the mutant under low light was similar or even faster in comparison to wild-type plants. However, growth under high light intensities resulted in decreased growth rates of psbs mutant plants. We conclude that energy dissipation in PSII related to qE is not primarily required for the protection of PSII against light-induced destruction, but may rather be involved in reducing the electron pressure on the photosynthetic electron transport chain at saturating light intensities.     

Dark heterotrophic growth conditions result in an increase in the content of photosystem II units in the filamentous cyanobacterium Anabaena variabilis ATCC 29413.

The filamentous nitrogen-fixing cyanobacterium Anabaena variabilis ATCC 29413 is capable of heterotrophic growth in complete darkness. After 6 months of continuous dark growth, both the autotrophic and heterotrophic cultures were found to have the same doubling time of 14 h. On a cellular basis, the chlorophyll content remained the same and the phycobilin content showed an increase in the dark-grown cultures. Fluorescence emission spectra at 77 K of dark-grown cells indicated that the phycobilisomes are functionally associated with photosystem II (PSII). Moreover, upon transfer to light, the dark-grown cells readily evolved oxygen. Although photosystem I (PSI) and whole chain-mediated electron transfer rates were comparable in both types of cultures, the rate of PSII-mediated electron transfer was found to be 20% higher in dark-grown cells. The PSI to PSII ratio changed from 6:1 in autotrophic cultures to 4:1 in the dark-grown cells. These changes in the rate of PSII electron transfer and in the stoichiometry between the two photosystems under dark, heterotrophic growth conditions were brought about by a preferential increase in the number of PSII units while the number of PSI units remained unchanged. The advantages of using this organism in the selection of PSI-deficient mutants are discussed.     

Effects of inactivating psbM and psbT on photodamage and assembly of photosystem II in Synechocystis sp. PCC 6803

PsbM and PsbT have been assigned to electron densities on both photosystem II (PSII) monomers at the PSII dimer interface in X-ray crystallographic structures from Thermosynechoccocus elongatus and T. vulcanus. Our results show that removal of either or both proteins from Synechocystis sp. PCC 6803 resulted in photoautotrophic strains but the DeltaPsbM:DeltaPsbT mutant did not form stable dimers. A CP43-less PSII monomer accumulated in both single mutants, although absence of PsbT destabilized PSII to a greater extent than removing PsbM. Additionally, DeltaPsbT cells exhibited slowed electron transfer between the plastoquinone electron acceptors, Q(A) and Q(B); however, S-state cycling in both mutants was similar to wild type. Oxygen evolution in these mutants rapidly inactivated following exposure to high light where recovery required protein synthesis and could proceed in the dark in DeltaPsbM cells but required light in DeltaPsbT cells. Interestingly, the extent of recovery of oxygen-evolving activity was greatest in the DeltaPsbM:DeltaPsbT strain. We also found recovery required Psb27 in DeltaPsbT cells although, under our conditions, the DeltaPsb27 strain remained similar to wild type. In contrast, the DeltaPsbM:DeltaPsb27 mutant could not assemble PSII beyond a CP43-minus intermediate. Our results suggest essential roles for Psb27 in biogenesis in the DeltaPsbM strain and for repair from photodamage in cells lacking PsbT.     

Requirement of carotene isomerization for the assembly of photosystem II in Synechocystis sp. PCC 6803

Carotene isomerase mutant (crtH mutant) cells of Synechocystis sp. PCC 6803 can accumulate beta-carotene under light conditions. However, the mutant cells grown under a light-activated heterotrophic growth condition contained detectable levels of neither beta-carotene nor D1 protein of the photosystem (PS) II reaction center, and no oxygen-evolving activity of PSII was detected. beta-Carotene and D1 protein appeared and a high level of PSII activity was detected after the cells were transferred to a continuous light condition. The PSI activities of thylakoid membranes from mutant cells were almost the same as those of thylakoid membranes from wild-type cells, both before and after transfer to the continuous light condition. These results suggest that beta-carotene is required for the assembly of PSII but not for that of PSI.     

The PsbY protein is not essential for oxygenic photosynthesis in the cyanobacterium Synechocystis sp. PCC 6803

A tetra-manganese cluster in the photosystem II (PSII) pigment-protein complex plays a critical role in the photosynthetic oxygen evolution process. PsbY, a small membrane-spanning polypeptide, has recently been suggested to provide a ligand for manganese in PSII (A.E. Gau, H.H. Thole, A. Sokolenko, L. Altschmied, R.G. Herrmann, E.K. Pistorius [1998] Mol Gen Genet 260: 56-68). We have constructed a mutant strain of the cyanobacterium Synechocystis sp. PCC 6803 with an inactivated psbY gene (sml0007). Southern-blot and polymerase chain reaction analysis showed that the mutant had completely segregated. However, the DeltapsbY mutant cells grew normally under photoautotrophic conditions. Moreover, growth of the wild-type and mutant cells were similar under high-light photoinhibition conditions, as well as in media without any added manganese, calcium, or chloride, three required inorganic cofactors for the oxygen-evolving complex of PSII. Analysis of steady-state and flash-induced oxygen evolution, fluorescence induction, and decay kinetics, and thermoluminescence profiles demonstrated that the DeltapsbY mutant cells have normal photosynthetic activities. We conclude that the PsbY protein in Synechocystis 6803 is not essential for oxygenic photosynthesis and does not provide an important binding site for manganese in the oxygen-evolving complex of PSII.     

MPH1 is a thylakoid membrane protein involved in protecting photosystem II from photodamage in land plants

Photosystem II (PSII) is highly susceptible to photoinhibition caused by environmental stimuli such as high light; therefore plants have evolved multifaceted mechanisms to efficiently protect PSII from photodamage. We previously published data suggesting that Maintenance of PSII under High light 1 (MPH1, encoded by AT5G07020), a PSII-associated proline-rich protein found in land plants, participates in the maintenance of normal PSII activity under photoinhibitory stress. Here we provide additional evidence for the role of MPH1 in protecting PSII against photooxidative damage. Two Arabidopsis thaliana mutants lacking a functional MPH1 gene suffer from severe photoinhibition relative to the wild-type plants under high irradiance light. The mph1 mutants exhibit significantly decreased PSII quantum yield and electron transport rate after exposure to photoinhibitory light. The mutants also display drastically elevated photodamage to PSII reaction center proteins after high-light treatment. These data add further evidence that MPH1 is involved in PSII photoprotection in Arabidopsis. MPH1 homologs are found across phylogenetically diverse land plants but are not detected in algae or prokaryotes. Taken together, these results suggest that MPH1 protein began to play a role in protecting PSII against excess light following the transition from aquatic to terrestrial conditions.     

The occurrence of a thylakoid-localized small zinc finger protein in land plants

Previous studies showed that LOW QUANTUM YIELD OF PHOTOSYSTEM II 1 (LQY1), a small thylakoid zinc finger protein was involved in maintenance and repair of Photosystem II (PSII). Here the author provide additional evidence for the role of LQY1 in PSII maintenance and repair and further commentary on the occurrence of LQY1 protein among land plants. After exposure to high light, Arabidopsis thaliana mutants lacking functional LQY1 gene (At1g75690) are more photoinhibited than wild-type control plants; display higher total non-photochemical quenching and photoinhibitory quenching. These results are consistent with the initial observation that lqy1 mutants have lower PSII efficiency than wild-type plants after high-light treatment. The low-PSII-efficiency phenotype can be suppressed upon complementation of lqy1 mutants with the LQY1 gene from wild-type plants. This further demonstrates that LQY1 is important in maintaining the activity of photosystem II in Arabidopsis. LQY1 homologs are present in land plants but are absent from sequenced genomes of green algae and cyanobacteria, which may reflect plant adaptation to excess light stress during the transition to land.     

Effect of light quality on chloroplast-membrane organization and function in pea

The effect of light quality during plant growth of chloroplast membrane organization and function in peas (Pisum sativum L. cv. Alaska) was investigated. In plants grown under photosystem (PS) I-enriched (far-red enriched) illumination both the PSII/PSI stoichiometry and the electrontransport capacity ratios were high, about 1.9. In plants grown under PSII-enriched (far-red depleted) illumination both the PSII/PSI stoichiometry and the electron-transport capacity ratios were significantly lower, about 1.3. In agreement, steady-state electron-transport measurements under synchronous illumination of PSII and PSI demonstrated an excess of PSII in plants grown under far-red-enriched light. Sodium dodecylsulfate polyacrylamide gel electrophoretic analysis of chlorophyll-containing complexes showed greater relative amounts of the PSII reaction center chlorophyll-protein complex in plants grown under farred-enriched light. Additional changes were observed in the ratio of light-harvesting chlorophyll a/b protein to PSII reaction center chlorophyll-protein under the two different light-quality regimes. The results demonstrate the dynamic nature of chloroplast structure and support the notion that light quality is an important factor in the regulation of chloroplast membrane organization and-function.Abbreviations and symbols Chl chlorophyll - CPa PSII reaction center chlorophyll protein complex - CPI PSI chlorophyll protein complex - FR-D light depleted in far-red sensitizing primarily PSII - FR-E light enriched in far-red sensitizing primarily PSI - LHCP PSII light-harvesting chlorophyll a/b protein complex - P ₇₀₀ primary electron donor of PSI - PSI, PSII photosystems I and II, respectively - Q primary electron acceptor of PSII     

Photosynthetic Properties of Photosystem II in Arabidopsis thaliana lpa1 Mutant

In a previous study, we characterized a high chlorophyll fluorescence lpa1 mutant of Arabidopsis thaliana, in which approximately 20% photosystem (PS) II protein is accumulated. In the present study, analysis of fluorescence decay kinetics and thermoluminescence profiles demonstrated that the electron transfer reaction on either the donor or acceptor side of PSII remained largely unaffected in the lpa1 mutant. In the mutant, maximal photochemical efficiency (Fv/Fm, where Fm is the maximum fluorescence yield and Fv is variable fluorescence) decreased with increasing light intensity and remained almost unchanged in wild-type plants under different light conditions. The Fv/Fm values also increased when mutant plants were transferred from standard growth light to low light conditions. Analysis of PSII protein accumulation further confirmed that the amount of PSII reaction center protein is correlated with changes in Fv/Fm in lpa1 plants. Thus, the assembled PSII in the mutant was functional and also showed increased photosensitivity compared with wild-type plants.(Author for correspondence. Tel: +86 (0)10 6283 6256; Fax: +86 (0)10 8259 9384; E-mail: zhanglixin@ibcas.ac.cn)