Cyclic electron flow around photosystem I in C(3) plants. In vivo control by the redox state of chloroplasts and involvement of the NADH-dehydrogenase complex

Cyclic electron flow around photosystem (PS) I has been widely described in vitro in chloroplasts or thylakoids isolated from C(3) plant leaves, but its occurrence in vivo is still a matter of debate. Photoacoustic spectroscopy and kinetic spectrophotometry were used to analyze cyclic PS I activity in tobacco (Nicotiana tabacum cv Petit Havana) leaf discs illuminated with far-red light. Only a very weak activity was measured in air with both techniques. When leaf discs were placed in anaerobiosis, a high and rapid cyclic PS I activity was measured. The maximal energy storage in far-red light increased to 30% to 50%, and the half-time of the P(700) re-reduction in the dark decreased to around 400 ms; these values are comparable with those measured in cyanobacteria and C(4) plant leaves in aerobiosis. The stimulatory effect of anaerobiosis was mimicked by infiltrating leaves with inhibitors of mitochondrial respiration or of the chlororespiratory oxidase, therefore, showing that changes in the redox state of intersystem electron carriers tightly control the rate of PS I-driven cyclic electron flow in vivo. Measurements of energy storage at different modulation frequencies of far-red light showed that anaerobiosis-induced cyclic PS I activity in leaves of a tobacco mutant deficient in the plastid Ndh complex was kinetically different from that of the wild type, the cycle being slower in the former leaves. We conclude that the Ndh complex is required for rapid electron cycling around PS I.     

Chlororespiration and poising of cyclic electron transport. Plastoquinone as electron transporter between thylakoid NADH dehydrogenase and peroxidase

Polypeptides encoded by plastid ndh genes form a complex (Ndh) which could reduce plastoquinone with NADH. Through a terminal oxidase, reduced plastoquinone would be oxidized in chlororespiration. However, isolated Ndh complex has low activity with plastoquinone and no terminal oxidase has been found in chloroplasts, thus the function of Ndh complex is unknown. Alternatively, thylakoid hydroquinone peroxidase could oxidize reduced plastoquinone with H(2)O(2). By immunoaffinity chromatography, we have purified the plastid Ndh complex of barley (Hordeum vulgare L.) to investigate the electron donor and acceptor specificity. A detergent-containing system was reconstructed with thylakoid Ndh complex and peroxidase which oxidized NADH with H(2)O(2) in a plastoquinone-dependent process. This system and the increases of thylakoid Ndh complex and peroxidase activities under photooxidative stress suggest that the chlororespiratory process consists of the sequence of reactions catalyzed by Ndh complex, peroxidase (acting on reduced plastoquinone), superoxide dismutase, and the non-enzymic one-electron transfer from reduced iron-sulfur protein (FeSP) to O(2). When FeSP is a component of cytochrome b(6).f complex or of the same Ndh complex, O(2) may be reduced with NADH, without requirement of light. Chlororespiration consumes reactive species of oxygen and, eventually, may decrease their production by lowering O(2) concentration in chloroplasts. The common plastoquinone pool with photosynthetic electron transport suggests that chlororespiratory reactions may poise reduced and oxidized forms of the intermediates of cyclic electron transport under highly fluctuating light intensities.     

Studies of the Ndh complex and photosystem II from mesophyll and bundle sheath chloroplasts of the C4-type plant Zea mays

In C(4) plants, granal mesophyll (MS) chloroplasts contain higher photosystem (PS) II and lower PS I activity than agranal bundle sheath (BS) chloroplasts. The maize NAD(P)H dehydrogenase or NAD(P)H-plastoquinone oxidoreductase (also named Ndh complex) from MS and BS chloroplasts, contains at least 11 subunits (NdhA-K) and is homologous to NADH dehydrogenase or Complex I from mitochondria and bacteria. The amount of Ndh complex is higher in BS compared with MS chloroplasts. However, there is little information about the interdependence of the PS II and Ndh complex in chlororespiration and linear and cyclic electron transport in C(4) plants. To characterize the expression of the PS II and Ndh complex in maize plastids, we used cytochrome b559 (cyt b559) antibodies and Ndh immunoglobulins (IgG) to analyze the Ndh complex and PS II in both MS and BS chloroplasts from maize leaves by Western blotting and immunolabeling. In Western blot experiments, it was found that the amount of cyt b559 (a marker for PS II) is 7-8 times higher in MS than BS chloroplasts. Conversely, the NdhH, -J, -K and -E content is 2.5-3 times higher in BS than MS chloroplasts. Similar results were obtained in immunolabeling experiments using Ndh IgGs and cyt b559 antibodies in MS and BS chloroplasts. These data suggest that in BS chloroplasts, ATP could be produced mainly by cyclic electron transport around PS I and Ndh complexes. Conversely, the linear electron transport in BS chloroplasts via PS II could have a lower production of ATP. These results also suggest that the contribution of the Ndh complex in the production of ATP in MS chloroplasts is minimal and that instead, this complex could have a chlororespiratory role.     

Photosynthetic regulation under fluctuating light in field-grown Cerasus cerasoides: A comparison of young and mature leaves

Photosystem I (PSI) is a potential target of photoinhibition under fluctuating light. However, photosynthetic regulation under fluctuating light in field-grown plants is little known. Furthermore, it is unclear how young leaves protect PSI against fluctuating light under natural field conditions. In the present study, we examined chlorophyll fluorescence, P700 redox state and the electrochromic shift signal in the young and mature leaves of field-grown Cerasus cerasoides (Rosaceae). Within the first seconds after any increase in light intensity, young leaves showed higher proton gradient (ΔpH) across the thylakoid membranes than the mature leaves, preventing over-reduction of PSI in the young leaves. As a result, PSI was more tolerant to fluctuating light in the young leaves than in the mature leaves. Interestingly, after transition from low to high light, the activity of cyclic electron flow (CEF) in young leaves increased first to a high level and then decreased to a stable value, while this rapid stimulation of CEF was not observed in the mature leaves. Furthermore, the over-reduction of PSI significantly stimulated CEF in the young leaves but not in the mature leaves. Taken together, within the first seconds after any increase in illumination, the stimulation of CEF favors the rapid lumen acidification and optimizes the PSI redox state in the young leaves, protecting PSI against photoinhibition under fluctuating light in field-grown plants.     

Identification of the Ndh (NAD(P)H-plastoquinone-oxidoreductase) complex in etioplast membranes of barley: changes during photomorphogenesis of chloroplasts

In the last few years the presence in thylakoid membranes of chloroplasts of a NAD(P)H-plastoquinone oxidoreductase complex (Ndh complex) homologous to mitochondrial complex I has been well established. Herein, we report the identification of the Ndh complex in barley etioplast membranes. Two plastid DNA-encoded polypeptides of the Ndh complex (NDH-A and NDH-F) were relatively more abundant in etioplast membranes than in thylakoids from greening chloroplasts. Conversion of etioplast into chloroplast, after light exposure of barley seedlings grown in the dark, was accompanied by a decrease in the NADH dehydrogenase activity associated to plastid membranes. Using native-PAGE and immunolabelling techniques we have determined that a NADH specific dehydrogenase activity associated with plastid membranes, which was more active in etioplasts than in greening chloroplasts, contained the NDH-A and NDH-F polypeptides. These results complemented by those obtained through blue-native-PAGE indicated that NDH-A and NDH-F polypeptides are part of a 580 kDa NADH dependent dehydrogenase complex present in etioplast membranes. This finding proves that accumulation of the Ndh complex is independent of light. The decrease in the relative levels and specific activity of this complex during the transition from etioplast to chloroplasts was accompanied by a parallel decrease in the specific activity of peroxidase associated to plastid membranes. Based on the mentioned observations it is proposed that an electron transport chain from NADH to H2O2 could be active in barley etioplasts.     

New subunits NDH-M, -N, and -O, encoded by nuclear genes, are essential for plastid Ndh complex functioning in higher plants

In higher plants, the Ndh complex reduces plastoquinones and is involved in cyclic electron flow around photosystem I, supplying extra-ATP for photosynthesis, particularly under environmental stress conditions. Based on plastid genome sequences, the Ndh complex would contain 11 subunits (NDH-A to -K), but homologies with bacterial complex indicate the probable existence of additional subunits. To identify missing subunits, tobacco (Nicotiana tabacum) NDH-H was His tagged at its N terminus using plastid transformation. A functional Ndh subcomplex was purified by Ni(2+) affinity chromatography and its subunit composition analyzed by mass spectrometry. Five plastid encoded subunits (NDH-A, -H, -I, -J, and -K) were identified as well as three new subunits (NDH-M, -N, and -O) homologous to cyanobacterial and higher plant proteins. Arabidopsis thaliana mutants missing one of these new subunits lack a functional Ndh complex, and NDH-M and NDH-N are not detected in a tobacco transformant lacking the Ndh complex. We discuss the involvement of these three nuclear-encoded subunits in the functional integrity of the plastidial complex.     

Functional characterization of the thylakoid Ndh complex phosphorylation by site-directed mutations in the ndhF gene

To investigate the phosphorylation of the NDH-F subunit of the thylakoid Ndh complex, we constructed three site-directed mutant transgenic tobaccos (Nicotiana tabacum) (T181A, T181S and T181D) in which the ₅₄₁ACT₅₄₃ triplet encoding the Thr-181 has been substituted by GCT, TCT or GAT encoding alanine, serine and aspartic acid, respectively. Western blots with phospho-threonine antibody detected the 73 kD NDH-F phosphorylated polypeptide in control but not in mutant tobaccos. Differences in Ndh activity, chlorophyll fluorescence and photosynthesis among mutants and control plant demonstrate the key role of the phosphorylation of conserved Thr-181 in the activity and function of the Ndh complex. The substitution of aspartic acid for threonine in T181D mimics the presumable activation effects of the threonine phosphorylation in Ndh activity, post-illumination increase of chlorophyll fluorescence and photosynthesis rapid responses to changing light intensities. A tentative role of the phosphorylation-activated Ndh complex is suggested to poise the redox level and, consequently, optimizing the rate of cyclic electron transport under field conditions.     

Inactivation of a plastid evolutionary conserved gene affects PSII electron transport, life span and fitness of tobacco plants

Chloroplasts contain a plastoquinone-NADH-oxidoreductase (Ndh) complex involved in protection against stress and the maintenance of cyclic electron flow. Inactivation of the Ndh complex delays the development of leaf senescence symptoms. Chlorophyll a fluorescence measurements, blue native gel electrophoresis, immunodetection and other techniques were employed to study tobacco (Nicotiana tabacum) Ndh-defective mutants (DeltandhF). The DeltandhF mutants compared with wild-type plants presented: (i) higher photosystem II : photosystem I (PSII : PSI) ratios; (ii) similar or higher levels of ascorbate, carotenoids, thylakoid peroxidase and superoxide dismutase, yield (Phi(PSII)) and maximal photochemical efficiency of PSII levels (F(v)/F(m)) than wild-type plant leaves of the same age; (iii) lower values of nonphotochemical quenching yield (Phi(NPQ)), but not at very high light intensities or during induced leaf senescence; (iv) a similar decrease of antioxidants during senescence; (v) no significant differences in the total foliar area and apical growth rate; and (vi) a production of viable seeds significantly higher than wild-type plants. These results suggest that the Ndh complex is involved in one of the redundant mechanisms that play a safety role in photosynthesis under stress, which has been conserved during evolution, but that its deletion increases fitness when plants are grown under favourable controlled conditions.     

Identification of a functional respiratory complex in chloroplasts through analysis of tobacco mutants containing disrupted plastid ndh genes.

The plastid genomes of several plants contain homologues, termed ndh genes, of genes encoding subunits of the NADH:ubiquinone oxidoreductase or complex I of mitochondria and eubacteria. The functional significance of the Ndh proteins in higher plants is uncertain. We show here that tobacco chloroplasts contain a protein complex of 550 kDa consisting of at least three of the ndh gene products: NdhI, NdhJ and NdhK. We have constructed mutant tobacco plants with disrupted ndhC, ndhK and ndhJ plastid genes, indicating that the Ndh complex is dispensible for plant growth under optimal growth conditions. Chlorophyll fluorescence analysis shows that in vivo the Ndh complex catalyses the post-illumination reduction of the plastoquinone pool and in the light optimizes the induction of photosynthesis under conditions of water stress. We conclude that the Ndh complex catalyses the reduction of the plastoquinone pool using stromal reductant and so acts as a respiratory complex. Overall, our data are compatible with the participation of the Ndh complex in cyclic electron flow around the photosystem I complex in the light and possibly in a chloroplast respiratory chain in the dark.     

田间条件下棉花幼叶光合特性及光保护机制

通过比较棉花(Gossypium hirsutum)幼叶和完全展开叶气体交换参数及叶绿素荧光特性的差异, 探讨高光强下幼叶的光抑制程度及明确光保护机制间的协调机理。在田间自然条件下, 以棉花刚展平的幼嫩叶片(幼叶)和面积已达到最大的完全展开叶片为研究对象, 通过测定不同发育阶段叶片气体交换参数及叶绿素a荧光参数的变化, 并运用Dual-PAM100对不同发育阶段的叶片进行快速光响应曲线的拟合。结果表明: 幼叶和完全展开叶片在光合、荧光特性方面表现出明显的差异。与完全展开叶相比, 较低的叶绿素(Chl)含量和气孔导度(G_s)是幼叶较低净光合速率(P_n)的限制因素, 从而直接导致其光系统II (PSII)实际光化学效率(Φ_PSII)和光化学猝灭系数(q_P)的降低。在1800 μmol·m^-2·s^-1光强以下, 完全展开叶具有较强的围绕PSI循环的电子流(CEF), 有利于合成ATP, 是其具有较高光合能力的原因之一。相同光强下, 幼叶较低的光饱和点(LSP)更易受光抑制, 但其PSII原初光化学效率(F_v/F_m)的日变化幅度显著小于完全展开叶, 说明强光下幼叶通过类胡萝卜素(Car)猝灭单线态氧、光呼吸(P_r)、热耗散(NPQ)以及PSI-CEF等光保护机制能有效地耗散过剩的光能, 从而避免其光合机构发生光抑制。     

Light-harvesting chlorophyll a-b complex requirement for regulation of Photosystem II photochemistry by non-photochemical quenching

Recently, it has been suggested (Horton et al. 1992) that aggregation of the light-harvesting a-b complex (LHC II) in vitro reflects the processes which occur in vivo during fluorescence induction and related to the major non-photochemical quenching (qE). Therefore the requirement of this chlorophyll a-b containing protein complex to produce qN was investigated by comparison of two barley mutants either lacking (chlorina f2) or depressed (chlorina¹⁰⁴) in LHC II to the wild-type and pea leaves submitted to intermittent light (IL) and during their greening in continuous light. It was observed that qN was photoinduced in the absence of LHC II, i.e. in IL grown pea leaves and the barley mutants. Nevertheless, in these leaves qN had no (IL, peas) or little (barley mutants) inhibitory effect on the photochemical efficiency of Q_A reduction measured by flash dosage response curves of the chlorophyll fluorescence yield increase induced by a single turn-over flash During greening in continuous light of IL pea leaves, an inhibitory effect on Q_A photoreduction associated to qN developed as Photosystem II antenna size increased with LHC II synthesis. Utilizing data from the literature on connectivity between PS II units versus antenna size, the following hypothesis is put forward to explain the results summarized above. qN can occur in the core antenna or Reaction Center of a fraction of PS II units and these units will not exhibit variable fluorescence. Other PS II units are quenched indirectly through PS II-PS II exciton transfer which develops as the proportion of connected PS II units increases through LHC II synthesis.     

Location,expression and orientation of the putative chlororespiratory enzymes,Ndh and IMMUTANS,in higher-plant plastids

The chloroplasts of many plants contain not only the photosynthetic electron transport chain, but also two enzymes, Ndh and IMMUTANS, which might participate in a chloroplast respiratory chain. IMMUTANS encodes a protein with strong similarities to the mitochondrial alternative oxidase and hence is likely to be a plastoquinol oxidase. The Ndh complex is a homologue of complex I of mitochondria and eubacteria and is considered to be a plastoquinone reductase. As yet these components have not been purified to homogeneity and their expression and orientation within the thylakoid remain ill-defined. Here we show that the IMMUTANS protein, like the Ndh complex, is a minor component of the thylakoid membrane and is localised to the stromal lamellae. Protease digestion of intact and broken thylakoids indicates that both Ndh and IMMUTANS are orientated towards the stromal phase of the membrane in Spinacia oleracea L. Such an orientation is consistent with a role for the Ndh complex in the energisation of the plastid membrane. In expression studies we show that IMMUTANS and the Ndh complex are present throughout the development of both Pisum sativum L. cv Progress No. 9 and Arabidopsis thaliana (L.) Heynh. leaves, from early expansion to early senescence. Interestingly, both the Ndh complex and the IMMUTANS protein accumulate within etiolated leaf tissue, lacking the photosystem II complex, consistent with roles outside photosynthetic electron transport.Abbreviations PQ plastoquinone - PSI, PSII photosystem I, II     

Role of thylakoid Ndh complex and peroxidase in the protection against photo-oxidative stress: fluorescence and enzyme activities in wild-type and ndhF-deficient tobacco

An Ndh-deficient mutant of tobacco ( Nicotiana tabacum cv. Petit Havana) was prepared by disrupting the ndhF gene in a transplastomic approach. The mutant (Δ ndhF ) showed 10% of the Ndh complex activity (EC 1.6.5.3) and 8% of the NDH-F polypeptide of that of non-transformed plants. However, in Δ ndhF , NDH-A, another Ndh polypeptide, was still present at 50% of the level in non-transformed plants. Δ ndhF tobacco showed higher sensitivity than non-transformed plants to photo-oxidative stress (as judged by chlorophyll bleaching) caused by increased light intensity and paraquat applications. These photo-oxidative treatments increased the amount and activity of the Ndh complex, thylakoid peroxidase, post-illumination chlorophyll fluorescence and non-photochemical quenching (NPQ) of chlorophyll fluorescence in non-transformed but not in Δ ndhF tobacco. Highly stressed non-transformed plants showed a rapid post-rise decline of chlorophyll fluorescence, probably indicating a re-oxidation of reduced plastoquinone. The results indicate that, in normal plants, the Ndh complex and thylakoid peroxidase (EC 1.11.1.7) provide and remove electrons, respectively, to balance the redox level of the intermediates of cyclic electron transport. In this way, they optimize the generation of the transmembrane H⁺ gradient of thylakoids and, as a consequence, increase the NPQ and the protection against photo-oxidative stress.     

Expression of the Plastid ndhF Gene Product in Photosynthetic and Non-Photosynthetic Tissues of Developing Barley Seedlings

A fragment of the NDH-F subunit of the plastid NAD(P)H dehydrogenasecomplex (NAD(P)H-plastoquinone-oxidoreductase) from barley wasexpressed as a fusion protein in Escherichia coli and an antibodyto the fusion protein was prepared. Western blot analysis usingthe anti-NDH-F antibody showed specificity towards a plastidpolypeptide of approximately 70 kDa present in both photosyntheticand non-photosynthetic barley tissue. The polypeptide was foundin thylakoid membranes of green leaves whereas in etiolatedleaves it was shown to be associated with the membrane fractionof etioplasts. NDH-F levels were higher in roots and etiolatedtissue than in greening or young leaves. During leaf ontogeny,NDH-F levels decreased from young to mature tissue but increasedduring senescence. The accumulation of NDH-F in thylakoids ofyoung leaves was stimulated by photooxidative treatment. Theresults indicate a high degree of expression of plastid ndhgenes (which encode NAD(P)H dehydrogenase sub-units) in non-photosyntheticplastids and under conditions which impair the photosyntheticactivity of chloroplasts. In addition to its putative implicationin photosynthetic electron transport, a non-photosynthetic role,such as chloro-respiration, is proposed for the plastid NAD(P)Hdehydrogenase complex. (Received May 20, 1997; Accepted October 8, 1997)     

Electron flow to photosystem I from stromal reductants in vivo: the size of the pool of stromal reductants controls the rate of electron donation to both rapidly and slowly reducing photosystem I units

Electron donation from stromal reductants to photosystem I (PSI) was studied using the kinetics of P700(+) (the oxidized primary donor of PSI) reduction in the dark after irradiation of barley ( Hordeum vulgare L.) leaves. The leaves were treated with diuron and methyl viologen to abolish both the electron flow from PSII and PSI-driven cyclic electron transport. The redox state of P700 was monitored using the absorbance changes at 830 nm (Delta A(830)). Two exponentially decaying components with half-times of about 3 s (the slow component) and about 0.6 s (the fast one) were distinguished in the kinetic curves of Delta A(830) relaxation after a 1-s pulse of far-red light. The complex kinetics of P700(+) reduction thus manifested two types of PSI unit differing in the rate of electron input from stromal reductants. The rates of both kinetic components assayed after 1-s pulses were increased about 20-fold by a short (2-5 min) heat-pretreatment of leaves, indicating the accelerated input of electrons to both types of PSI unit. The increased rates of electron flow to P700(+) were even observed 1.5 h after the action of heat had been completed. Both kinetic components were dramatically slowed down upon irradiation of heat-treated leaves for 20-30 s. Their rates were restored after a short (20-30 s) period of darkness. A 5-min leaf exposure at 38 degrees C was sufficient to stimulate by severalfold the reduction of P700(+) pre-oxidized by a brief light pulse. In contrast, the acceleration of P700(+) reduction after a 1-min irradiation was observed only if leaves were subjected to temperatures above 40 degrees C. Neither heat treatment of leaves nor light-dark modulations in the rates of the fast and the slow components of P700(+) dark reduction influenced the relative magnitudes of the two kinetic components, providing strong additional evidence in favor of two distinct types of PSI existing per se in barley leaves. The key role in the control of the activity of electron donation to P700(+) in both rapidly and slowly reducing PSI units was attributed to the amount of stromal reductants available for P700(+) reduction. The latter was expected to be reduced under illumination in the presence of methyl viologen, while increased again in the dark. The regeneration of the pool of stromal reductants in the dark was likely provided by starch breakdown within the chloroplast stroma, but not by import of reducing equivalents from the cytosol. This was evidenced by much lower rates, compared with 1-h dark-adapted leaves, of dark reduction of both components of P700(+) in leaves stored for 24 h in the dark and thus depleted of starch but containing large amounts of glucose, the respiratory substrate.     

CO2 assimilation, xanthophyll cycle pigments and PSII efficiency in pumpkin plants as affected by ozone fumigation

CO₂ assimilation, xanthophyll cycle pigments and PSII efficiency were analyzed in two different ages of pumpkin leaves (Cucurbita pepo L. cv. Ambassador) exposed to 150 nmol mol^-1 of ozone (5 days, 5 h day^-1). Gas-exchange measurements revealed a reduction in CO₂ assimilation and stomatal conductance, accompanied by an increase in the intercellular CO₂ concentration both in young and in mature leaves as compared to their respective controls. In both leaves, F₀ remained unchanged, while F_m and the F_v/F_m ratio decreased after O₃ fumigation, indicating that ozone may induce an alteration in the capability of photosystem II (PSII) to reduce the primary acceptor Q_A. In the mature leaves the photochemical quenching (q_p) was significantly lowered by the pollutant, but this was not the case in the young leaves where q_p did not change. In both mature and young ozonated pumpkin leaves, the development of non-photochemical quenching caused a decrease in the PSII photochemical rate, as shown by the correlation between F_v/F_m and the de-epoxidation state of dark-adapted leaves. Decreases in the F_v/F_m ratio are generally attributed to damage to the PSII reaction centre, apart from the down-regulation of the capacity of PSII electron transport. While in young ozonated leaves the decrease in the F_v/F_m ratio was not associated with damage to the D1 protein, in mature ozonated pumpkin leaves, the decrease in the F_v/F_m was accompanied by a significant decline in the D1 content. In conclusion, ozone exposure induces alterations in the light reactions of photosynthesis in both young and mature leaves. However, in young leaves the engagement of the xanthophyll cycle appears to counteract ozone effects against the photosynthetic apparatus as demonstrated by the absence of damage to the D1 protein. On the other hand, the loss of D1 protein in mature fumigated leaves suggests that the activation of the xanthophyll cycle is not sufficient to prevent photoinhibition, probably because a physiological state of senescence adds to the oxidative stress.     

Competition between linear and cyclic electron flow in plants deficient in Photosystem I

Photosynthetic electron transport can involve either a linear flow from water to NADP, via Photosystems (PS) II and I or a cyclic flow just involving PSI. Little is known about factors regulating the relative flow through each of these pathways. We have examined photosynthetic electron transport through each system in plants of Arabidopsis thaliana in which either the PSI-D1 or PSI-E1 subunits of PSI have been knocked out. In both cases, this results in an imbalance in the turnover of PSI and PSII, such that PSII electron transport is limited by PSI turnover. Phosphorylation of light-harvesting complex II (LHCII) and its migration to PSI is enhanced but only partially reversible and not sufficient to balance photosystem turnover. In spite of this, cyclic electron flow is able to compete efficiently with PSI across a range of conditions. In dark-adapted leaves, the efficiency of cyclic relative to linear flow induced by far-red light is increased, implying that the limiting step of cyclic flow lies in the re-injection of electrons into the electron transport chain. Illumination of leaves with white light resulted in transient induction of a significant non-photochemical quenching in knockout plants which is probably high energy state quenching induced by cyclic electron flow. At high light and at low CO(2), non-photochemical quenching was greater in the knockout plants than in the wildtype. Comparison of PSI and PSII turnover under such conditions suggested that this is generated by cyclic electron flow around PSI. We conclude that, when the concentration of PSI is limiting, cyclic electron flow is still able to compete effectively with linear flow to maintain a high DeltapH to regulate photosynthesis.