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Under conditions of iron-stress, the Photosystem II associated chlorophyll a protein complex designated CP 43', which is encoded by the isiA gene, becomes the major pigment-protein complex in Synechococcus sp. PCC 7942. The isiB gene, which is located immediately downstream of isiA, encodes the protein flavodoxin, which can functionally replace ferredoxin under conditions of iron stress. We have constructed two cyanobacterial insertion mutants which are lacking (i) the CP 43' apoprotein (designated isiA (-)) and (ii) flavodoxin (designated isiB (-)). The function of CP 43' was studied by comparing the cell characteristics, PS II functional absorption cross-sections and Chl a fluorescence parameters from the wild-type, isiA (-) and isiB (-) strains grown under iron-stressed conditions. In all strains grown under iron deprivation, the cell number doubling time was maintained despite marked changes in pigment composition and other cell characteristics. This indicates that iron-starved cells remained viable and that their altered phenotype suggests an adequate acclimation to low iron even in absence of CP 43' and/or flavodoxin. Under both iron conditions, no differences were detected between the three strains in the functional absorption crossection of PS II determined from single turnover flash saturation curves of Chl a fluorescence. This demonstrates that CP 43' is not part of the functional light-harvesting antenna for PS II. In the wild-type and the isiB (-) strain grown under iron-deficient conditions, CP 43' was present in the thylakoid membrane as an uncoupled Chl-protein complex. This was indicated by (1) an increase of the yield of prompt Chl a fluorescence (Fo) and (2) the persistence after PS II trap closure of a fast fluorescence decay component showing a maximum at 685 nm.

The enzyme superoxide dismutase is ubiquitous in aerobic organisms where it plays a major role in alleviating oxygen-radical toxicity. An insertion mutation introduced into the iron superoxide dismutase locus (designated sodE) of the cyanobacterium Synechococcus sp. PCC 7942 created a mutant strain devoid of detectable iron superoxide dismutase activity. Both wild-type and mutant strains exhibited similar photosynthetic activity and viability when grown with 17 jmolm-2-s'1 illumination in liquid culture supplemented with 3% carbon dioxide. In contrast, the sodB mutant exhibited significantly greater damage to its photosynthetic system than the wild-type strain when grown under increased oxygen tension or with methyl viologen. Although damage occurs at both photosystems I and II, it is primarily localized at photosystem I in the sodB mutant. Growth in 100% molecular oxygen for 24 hr decreased photoacoustically measured energy storage in 3-(3,4-dichlorophenyl)-1,1-dimethylurea and abolished the fluorescence state 2 to state 1 transition in the sodB mutant, indicating interruption of cyclic electron flow around photosystem I. Analysis of the flash-induced absorption transient at 705 nm indicated that the interruption of cyclic electron flow occurred in the return part of the cycle, between the two [4 Fe-4 SI centers of photosystem I, FA and FB, and cytochrome f. Even though the sodB mutant was more sensitive to damage by active oxygen than wild-type cells, both strains were equally sensitive to the photoinhibition of photosystem II caused by exposure to strong light.

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Photosystem H reaction centers in plants, algae, and cyanobacteria are susceptible to damage by excess light that irreversibly impairs activity and eventually results in the proteolytic degradation of at least one of the core proteins. Oxygenic photosynthetic organisms are vulnerable to damage by a number of environmental factors, including visible light, UV-B radiation, heat, cold, drought, and atmospheric pollutants (1). In some cases, the site of damage is the photosystem II (PSII) reaction center, a membrane-bound, multisubunit complex that catalyzes the oxidation of water and the reduction of plastoquinone (2). The PSII reaction center provides approximately half the energy used for biomass production in plants and is responsible for the release of molecular oxygen into the atmosphere. The reaction center core is composed of the a and f8 subunits of cytochrome b-559 (Cyt b559) and the two proteins, D1 and D2, that bind the redox components known to be required for electron transfer from the water-oxidizing manganese cluster to plastoquinone. These include the primary donor (P680), the secondary donor (Yz), the primary acceptor pheophytin (Pheo), and the primary and secondary plastoquinone acceptors (QA and QB).Exposure of PSII to supersaturating levels of visible light causes the loss of water oxidation (3) and eventually leads to removal and degradation of the D1 polypeptide (4). There is disagreement concerning the early events that lead to photoinhibition of the reaction center (reviewed in ref. 5). Using thylakoid membranes as model systems, some experiments show that the initial site of damage is on the oxidizing side of PSII, whereas other experiments, often done under different conditions, show that the primary site of damage is on the reducing side of PSII. These results form the basis for two models of photoinhibition: those that focus on damage done by the highly oxidizing cation radicals and those that focus on damage done by the highly reducing anion radicals.This study was initiated to investigate the molecular mechanism of PSII photoinhibition and protective reactions. Potentiometric titrations reveal that a one-electron redox component plays a critical role in the process. The rate of PSII damage is dramatically increased when the component is reduced. We suggest that the redox component is the lowpotential form of Cyt b559 (Cyt b559LP), which protects the reaction center from photoinhibition by providing an alternative electron pathway that can oxidize the damaging radical state P680/Pheo-/QA-MATERIALS AND METHODS Membrane Isolation. Spinach leaves (Spinacia oleracea) were either harvested from plants grown hydroponically (6) or bought from a local market. Thylakoid membranes were isolated from leaves as described (7) and stored on ice at a chlorophyll concentration of 1-2 mM in medium containing 5 mM Hepes-NaOH (pH 7.5), 200 mM sorbitol, 2 mM MgCl2, and fatty acid-free bovine serum albumin (0.5 mg/ml). The membranes were used for experiments within 7 h ofisolation. Chlorophyll conce...

Light that exceeds the photosynthetic capacity of a plant can impair the ability of photosystem II to oxidize water. The light-induced inhibition is initiated by inopportune electron transport reactions that create damaging redox states. There is evidence that secondary electron transport pathways within the photosystem II reaction center can protect against potentially damaging redox states. Experiments using thylakoid membranes poised at different ambient redox potentials demonstrate that light-induced damage to photosystem II can be controlled by a redox component within the reaction center [Nedbal, L., et al. (1992) Proc. Natl. Acad. Sci. U.S.A. 89, 7929-7933]. The rate of photoinhibition is slow when the redox component is oxidized, but increases by more than 10-fold when the redox component is reduced. Here, using spinach thylakoid membranes, we provide evidence that the redox component is cytochrome b559, an intrinsic heme protein of the photosystem II reaction center. The results support a model in which the low-potential (LP) form of cytochrome b559 protects photosystem II by deactivating a rarely formed, but hazardous redox state of photosystem II, namely, P680/Pheo-/ QA-. Cytochrome b559LP is proposed to deactivate this potentially lethal redox state by accepting electrons from reduced pheophytin. The key observations supporting this proposal are as follows: (1) The oxidation-reduction potential of cytochrome b559LP is in the range predicted by redox titrations of photoinhibition. (2) If cytochrome b559LP is reduced prior to illumination, the rate of photoinhibition is fast, whereas if the cytochrome is oxidized prior to illumination, the rate of photoinhibition is slow.(ABSTRACT TRUNCATED AT 250 WORDS)

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