Plastoquinone Pool Research Articles

Regulation of electron transport is essential for photosystem I stability and plant growth.

Photosynthetic electron transport is regulated by cyclic and pseudocyclic electron flow (CEF and PCEF) to maintain the balance between light availability and metabolic demands. CEF transfers electrons from photosystem I to the plastoquinone pool with two mechanisms, dependent either on PGR5/PGRL1 or on the type I NADH dehydrogenase-like (NDH) complex. PCEF uses electrons from photosystem I to reduce oxygen and in many groups of photosynthetic organisms, but remarkably not in angiosperms, it is catalyzed by flavodiiron proteins (FLVs). In this study, Physcomitrella patens plants depleted in PGRL1, NDH and FLVs in different combinations were generated and characterized, showing that all these mechanisms are active in this moss. Surprisingly, in contrast to flowering plants, Physcomitrella patens can cope with the simultaneous inactivation of PGR5- and NDH-dependent CEF but, when FLVs are also depleted, plants show strong growth reduction and photosynthetic activity is drastically reduced. The results demonstrate that mechanisms for modulation of photosynthetic electron transport have large functional overlap but are together indispensable to protect photosystem I from damage and they are an essential component for photosynthesis in any light regime.

Photobiochemical changes in Chlorella g120 culture during trophic conversion (metabolic pathway shift) from heterotrophic to phototrophic growth regime

Physiological and photobiochemical changes and growth in the heterotrophic strain Chlorella vulgaris g120 were studied during trophic conversion from heterotrophic to phototrophic growth regime. After the exposure of the Chlorella g120 culture to light, it revealed a significant activity of the electron transport (450–700 μmol e− m−2 s−1 as measured by chlorophyll fluorescence) and high PSII photochemical yield Fv/Fm between 0.7 and 0.8. Fast fluorescence induction kinetics showed that PSII electron acceptors in the plastoquinone pool remained partly oxidized, indicating no downregulation of PSII electron transport. The data further revealed that high photobiochemical activity is lost in futile (protective) processes of non-photochemical quenching and respiration which indicate that surplus energy is dissipated in these processes. Pigment analysis showed low chlorophyll content − 0.35–1.15% as compared with exclusively phototrophic strain Chlorella vulgaris R-117. Nevertheless, the carotenoid content in g120 was relatively high − 0.20–0.33% of dry weight which resulted in a considerably high ratio of carotenoid/chlorophyll − 0.31–0.44. This strain probably does not possess the fully competent photosynthetic apparatus and can only partially adapt to phototrophy. We show that the heterotrophically grown g120 strain can undergo metabolic shift from heterotrophic to phototrophic growth regime. It might be an interesting strain from biotechnological point of view as a source of carotenoids, especially lutein.

Revisiting cyanobacterial state transitions

Photosynthetic organisms are exposed to a fluctuating environment in which light intensity and quality change continuously. Specific illumination of either photosystem (PSI or PSII) creates an energy imbalance, leading to the reduction or oxidation of the intersystem electron transport chain. This redox imbalance could trigger the formation of dangerous reactive oxygen species. Cyanobacteria, like plants and algae, have developed a mechanism to re-balance this preferential excitation of either reaction center, called state transitions. State transitions are triggered by changes in the redox state of the membrane-soluble plastoquinone (PQ) pool. In plants and green algae, these changes in redox potential are sensed by Cytochrome b6f, which interacts with a specific kinase that triggers the movement of the main PSII antenna (the light-harvesting complex II). By contrast, although cyanobacterial state transitions have been studied extensively, there is still no agreement about the molecular mechanism, the PQ redox state sensor and the signaling pathways involved. In this review, we aimed to critically evaluate the results published on cyanobacterial state transitions, and discuss the “new” and “old” models in the subject. The phycobilisome and membrane contributions to this physiological process were addressed and the current hypotheses regarding its signaling transduction pathway were discussed.

Realtime kinetics of the light driven steps of photosynthetic water oxidation in living organisms by “stroboscopic” fluorometry

We develop a rapid “stroboscopic” fluorescence induction method, using the fast repetition rate fluorometry (FRRF) technique, to measure changes in the quantum yield of light emission from chlorophyll in oxygenic photosynthesis arising from competition with primary photochemical charge separation (P680* ➔ P680+QA−). This method determines the transit times of electrons that pass through PSII during the successive steps in the catalytic cycle of water oxidation/O2 formation (S states) and plastoquinone reduction in any oxygenic phototroph (in vivo or in vitro). We report the first measurements from intact living cells, illustrated by a eukaryotic alga (Nannochloropsis oceanica). We demonstrate that S state transition times depend strongly on the redox state of the PSII acceptor side, at both QB and the plastoquinone pool which serve as the major locus of regulation of PSII electron flux. We provide evidence for a kinetic intermediate S3′ state (lifetime 220 μs) following formation of S3 and prior to the release of O2. We compare the FRRF-detected kinetics to other previous spectroscopic methods (optical absorbance, EPR, and XES) that are applicable only to in vitro samples.

Photosynthetic characterization of flavodoxin-expressing tobacco plants reveals a high light acclimation-like phenotype

Flavodoxins are electron carrier flavoproteins present in bacteria and photosynthetic microorganisms which duplicate the functional properties of iron-sulphur containing ferredoxins and replace them under adverse environmental situations that lead to ferredoxin decline. When expressed in plant chloroplasts, flavodoxin complemented ferredoxin deficiency and improved tolerance to multiple sources of biotic, abiotic and xenobiotic stress. Analysis of flavodoxin-expressing plants grown under normal conditions, in which the two carriers are present, revealed phenotypic effects unrelated to ferredoxin replacement. Flavodoxin thus provided a tool to alter the chloroplast redox poise in a customized way and to investigate its consequences on plant physiology and development. We describe herein the effects exerted by the flavoprotein on the function of the photosynthetic machinery. Pigment analysis revealed significant increases in chlorophyll a, carotenoids and chlorophyll a/b ratio in flavodoxin-expressing tobacco lines. Results suggest smaller antenna size in these plants, supported by lower relative contents of light-harvesting complex proteins. Chlorophyll a fluorescence and P700 spectroscopy measurements indicated that transgenic plants displayed higher quantum yields for both photosystems, a more oxidized plastoquinone pool under steady-state conditions and faster plastoquinone dark oxidation after a pulse of saturating light. Many of these effects resemble the phenotypes exhibited by leaves adapted to high irradiation, a most common environmental hardship faced by plants growing in the field. The results suggest that flavodoxin-expressing plants would be better prepared to cope with this adverse situation, and concur with earlier observations reporting that hundreds of stress-responsive genes were induced in the absence of stress in these lines.

Minimizing an Electron Flow to Molecular Oxygen in Photosynthetic Electron Transfer Chain: An Evolutionary View.

Recruitment of H2O as the final donor of electrons for light-governed reactions in photosynthesis has been an utmost breakthrough, bursting the evolution of life and leading to the accumulation of O2 molecules in the atmosphere. O2 molecule has a great potential to accept electrons from the components of the photosynthetic electron transfer chain (PETC) (so-called the Mehler reaction). Here we overview the Mehler reaction mechanisms, specifying the changes in the structure of the PETC of oxygenic phototrophs that probably had occurred as the result of evolutionary pressure to minimize the electron flow to O2. These changes are warranted by the fact that the efficient electron flow to O2 would decrease the quantum yield of photosynthesis. Moreover, the reduction of O2 leads to the formation of reactive oxygen species (ROS), namely, the superoxide anion radical and hydrogen peroxide, which cause oxidative stress to plant cells if they are accumulated at a significant amount. From another side, hydrogen peroxide acts as a signaling molecule. We particularly zoom in into the role of photosystem I (PSI) and the plastoquinone (PQ) pool in the Mehler reaction.

Molecular Mechanism of Oxidation of P700 and Suppression of ROS Production in Photosystem I in Response to Electron-Sink Limitations in C3 Plants.

Photosynthesis fixes CO2 and converts it to sugar, using chemical-energy compounds of both NADPH and ATP, which are produced in the photosynthetic electron transport system. The photosynthetic electron transport system absorbs photon energy to drive electron flow from Photosystem II (PSII) to Photosystem I (PSI). That is, both PSII and PSI are full of electrons. O2 is easily reduced to a superoxide radical (O2−) at the reducing side, i.e., the acceptor side, of PSI, which is the main production site of reactive oxygen species (ROS) in photosynthetic organisms. ROS-dependent inactivation of PSI in vivo has been reported, where the electrons are accumulated at the acceptor side of PSI by artificial treatments: exposure to low temperature and repetitive short-pulse (rSP) illumination treatment, and the accumulated electrons flow to O2, producing ROS. Recently, my group found that the redox state of the reaction center of chlorophyll P700 in PSI regulates the production of ROS: P700 oxidation suppresses the production of O2− and prevents PSI inactivation. This is why P700 in PSI is oxidized upon the exposure of photosynthesis organisms to higher light intensity and/or low CO2 conditions, where photosynthesis efficiency decreases. In this study, I introduce a new molecular mechanism for the oxidation of P700 in PSI and suppression of ROS production from the robust relationship between the light and dark reactions of photosynthesis. The accumulated protons in the lumenal space of the thylakoid membrane and the accumulated electrons in the plastoquinone (PQ) pool drive the rate-determining step of the P700 photo-oxidation reduction cycle in PSI from the photo-excited P700 oxidation to the reduction of the oxidized P700, thereby enhancing P700 oxidation.

Growth under Fluctuating Light Reveals Large Trait Variation in a Panel of Arabidopsis Accessions.

The oxidation of P700 in photosystem I (PSI) is a robust mechanism that suppresses the production of reactive oxygen species. We researched the contribution of photorespiration to the oxidation of P700 in wheat leaves. We analyzed the effects of changes in partial pressures of CO2 and O2 on photosynthetic parameters. The electron flux in photosynthetic linear electron flow (LEF) exhibited a positive linear relationship with an origin of zero against the dissipation rate (vH+) of electrochromic shift (ECS; ΔpH across thylakoid membrane), indicating that cyclic electron flow around PSI did not contribute to H+ usage in photosynthesis/photorespiration. The vH+ showed a positive linear relationship with an origin of zero against the H+ consumption rates in photosynthesis/photorespiration (JgH+). These two linear relationships show that the electron flow in LEF is very efficiently coupled with H+ usage in photosynthesis/photorespiration. Lowering the intercellular partial pressure of CO2 enhanced the oxidation of P700 with the suppression of LEF. Under photorespiratory conditions, the oxidation of P700 and the reduction of the plastoquinone pool were stimulated with a decrease in JgH+, compared to non-photorespiratory conditions. These results indicate that the reduction-induced suppression of electron flow (RISE) suppresses the reduction of oxidized P700 in PSI under photorespiratory conditions. Furthermore, under photorespiratory conditions, ECS was larger and H+ conductance was lower against JgH+ than those under non-photorespiratory conditions. These results indicate that photorespiration enhances RISE and ΔpH formation by lowering H+ conductance, both of which contribute to keeping P700 in a highly oxidized state.

Photosynthetic and biochemical traits change in the green-tide-forming macroalga Ulva pertusa during sporulation1.

The physiological and biochemical changes in the green macroalga Ulva pertusa during the progression of sporulation have been characterized. The transition from the vegetative to the sporulation stage was accompanied by an increase in chlorophyll a (Chl a), chlorophyll b (Chl b), and carotenoid content, as well as an increase in DPPH scavenging and responsiveness to diphenylamine. However, oxygen evolution and maximum electron transport rate decreased. The discrepancy between photosynthetic performance and pigment content might relate to the self-shading of spores within a sporangium. Spore-forming U.pertusa thalli were low-light-adapted, due to an increase in the number of photosynthetic units. Decreased electron transport during sporulation might trigger sporulation, as for some cyanobacteria and other Ulva spp., via oxidization of the plastoquinone pool and cyclic phosphorylation, thus producing ATP to generate carbon and nitrogen skeletons required for spores. It is thus concluded that carotenoids function both in spore initiation and/or maturation and in their photoprotection.

Comparative analysis of thylakoid protein complexes in state transition mutants nsi and stn7: focus on PSI and LHCII

The photosynthetic machinery of plants can acclimate to changes in light conditions by balancing light-harvesting between the two photosystems (PS). This acclimation response is induced by the change in the redox state of the plastoquinone pool, which triggers state transitions through activation of the STN7 kinase and subsequent phosphorylation of light-harvesting complex II (LHCII) proteins. Phosphorylation of LHCII results in its association with PSI (state 2), whereas dephosphorylation restores energy allocation to PSII (state 1). In addition to state transition regulation by phosphorylation, we have recently discovered that plants lacking the chloroplast acetyltransferase NSI are also locked in state 1, even though they possess normal LHCII phosphorylation. This defect may result from decreased lysine acetylation of several chloroplast proteins. Here, we compared the composition of wild type (wt), stn7 and nsi thylakoid protein complexes involved in state transitions separated by Blue Native gel electrophoresis. Protein complex composition and relative protein abundances were determined by LC–MS/MS analyses using iBAQ quantification. We show that despite obvious mechanistic differences leading to defects in state transitions, no major differences were detected in the composition of PSI and LHCII between the mutants. Moreover, both stn7 and nsi plants show retarded growth and decreased PSII capacity under fluctuating light as compared to wt, while the induction of non-photochemical quenching under fluctuating light was much lower in both nsi mutants than in stn7.

24-Epibrassinolide alleviates the toxic effects of NaCl on photosynthetic processes in potato plants.

Brassinosteroids are promising agents for alleviating the negative effects of salinity on plants, but the mechanism of their protective action is far from being understood. We investigated the effect of pretreatment with 24-epibrassinolide (24-EBL) on the photosynthetic and physiological parameters of potato plants under progressive salinity stress caused by root application of 100mM NaCl. Salinity clearly inhibited primary photosynthetic processes in potato plants by reducing the contents of photosynthetic pigments, photosynthetic electron transport and photosystem II (PSII) maximal and effective quantum yields. These negative effects of salinity on primary photosynthetic processes were mainly due to toxic ionic effects on the plant's ability to oxidize the plastoquinone pool. Pretreatment with 24-EBL alleviated this stress effect and allowed the maintenance of plastoquinone pool oxidation and the efficiency of photosystem II photochemistry to be at the same levels as those in unstressed plants; however, the pretreatment did not affect the photosynthetic pigment content. 24-EBL pretreatment clearly alleviated the decrease in leaf osmotic potential under salinity stress. The stress-induced increases in lipid peroxidation and proline contents were not changed under brassinosteroid pretreatment. However, 24-EBL pretreatment increased the peroxidase activity and improved the K+/Na+ ratio in potato leaves, which were likely responsible for the protective 24-EBL action under salt stress.

An evolutionarily conserved iron-sulfur cluster underlies redox sensory function of the Chloroplast Sensor Kinase

Photosynthetic efficiency depends on equal light energy conversion by two spectrally distinct, serially-connected photosystems. The redox state of the plastoquinone pool, located between the two photosystems, is a key regulatory signal that initiates acclimatory changes in the relative abundance of photosystems. The Chloroplast Sensor Kinase (CSK) links the plastoquinone redox signal with photosystem gene expression but the mechanism by which it monitors the plastoquinone redox state is unclear. Here we show that the purified Arabidopsis and Phaeodactylum CSK and the cyanobacterial CSK homologue, Histidine kinase 2 (Hik2), are iron-sulfur proteins. The Fe-S cluster of CSK is further revealed to be a high potential redox-responsive [3Fe-4S] center. CSK responds to redox agents with reduced plastoquinone suppressing its autokinase activity. Redox changes within the CSK iron-sulfur cluster translate into conformational changes in the protein fold. These results provide key insights into redox signal perception and propagation by the CSK-based chloroplast two-component system.

Regulation of the size of photosystem II light harvesting antenna represents a universal mechanism of higher plant acclimation to stress conditions.

We investigated acclimatory responses of Arabidopsis plants to drought and salinity conditions before the appearance of obvious signs of damage caused by these factors. We detected changes indicating an increase in the reduction level of the chloroplast plastoquinone pool (PQ pool) 5-7 days after introduction of the stress factors. After 10-14 days, a decrease in the size of PSII light harvesting antenna was observed in plants under conditions of drought and salinity. This was confirmed by a decrease in content of PSII antenna proteins and by downregulation of gene expression levels of these proteins under the stress conditions. No changes in values of performance index and maximum quantum yield of PSII were detected. Under drought and salinity, the content of hydrogen peroxide in leaves was higher than in control leaves. Thus, we propose that reduction of the size of PSII antenna represents one of the universal mechanisms of acclimation of higher plants to stress factors and the downsizing already begins to manifest under mild stress conditions. Both the PQ pool reduction state and the hydrogen peroxide content are important factors needed for the observed rearrangement.

Acclimation to low ultraviolet-B radiation increases photosystem I abundance and cyclic electron transfer with enhanced photosynthesis and growth in the cyanobacterium Nostoc sphaeroides.

Ultraviolet-B radiation is known to harm most photosynthetic organisms with the exception of several studies of photosynthetic eukaryotes in which UV-B showed positive effects. In this study, we investigated the effect of acclimation to low UV-B radiation on growth and photosynthesis of the cyanobacterium Nostoc sphaeroides. Exposure to 0.08 W m-2 UV-B plus low visible light for 14 d significantly increased the growth rate and biomass production by 16% and 30%, respectively, compared with those under visible light alone. The UV-B acclimated cells showed an approximately 50% increase in photosynthetic efficiency (α) and photosynthetic capacity (Pmax ), a higher PSI/PSII fluorescence ratio, an increase in PSI content and consequently enhanced cyclic electron flow, relative to those of non-acclimated cells. Both the primary quinone-type acceptor and plastoquinone pool re-oxidation were up-regulated in the UV-B acclimated cells. In parallel, the UV-B acclimated colonies maintained a higher rate of D1 protein synthesis following exposure to elevated intensity of UV-B or visible light, thus functionally mitigating photoinhibition. The present data provide novel insight into photosynthetic acclimation to low UV-B radiation and suggest that UV-B may act as a positive ecological factor for the productivity of some photosynthetic prokaryotes, especially during twilight periods or in shaded environments.

Is a blue–red light a good elicitor of phenolic compounds in the family Droseraceae? A comparative study

Plants from the family Droseraceae, especially Drosera sp. and Dionaea sp., are naturally rich in phenolic derivatives such as plumbagin, among others. Plumbagin is known both for its pharmacological significance and its protective properties against light stress. Light stress – high light intensity or/and light spectral composition – activates plants' response mechanisms including, among others, hormonal (salicylic acid, jasmonic acid) pathways and secondary metabolite (phenolic compounds, proline) pathways. Short-wavelength radiation, due to its high energy, will induce the synthesis of protective secondary metabolites, including those with pharmaceutical properties. The aim of the study was to describe and compare acclimation strategies of Drosera peltata and Dionaea muscipula to blue–red light in the context of phenolic compound accumulation, and salicylic acid, jasmonic acid and proline synthesis.For the first time, differences in the responses of D. muscipula and D. peltata to blue–red light (in the ratio 6:1) were established. In Dionaea sp., it was associated with the use of redox equivalents (in particular, plastoquinone pool) for the synthesis of primary metabolites used in the process of growth and development. In Drosera sp., a rapid adjustment of redox state led to the synthesis of secondary metabolites, constituting a reservoir of carbon skeletons and allowing for a quick defence response to stress factors. In both species, blue–red light did not induce the jasmonic acid pathway. However, the salicylic acid pathway was induced as an alternative to the phenolic compound synthesis pathway. Nevertheless, the applied blue–red light was not an effective elicitor of phenolic compounds in the plants examined.

Comparative photosynthetic responses of Norway spruce and Scots pine seedlings to prolonged water deficiency

Stressors of different natures, including drought stress, substantially compromise the ability of plants to effectively and safely utilize light energy. We investigated the influence of water stress on the photosynthetic processes in Picea abies and Pinus sylvestris, two species with contrasting drought sensitivities. Spruce and pine seedlings were exposed to polyethylene glycol 6000-induced water deficits of different intensities and durations. The maintenance of photosystem I (PSI) oxidation in spruce required increased photosynthetic control and led to the increased reduction of the plastoquinone pool, which was not the case in pine seedlings. As a result of increased excitation pressure, photosystem II (PSII) inactivation was observed in spruce plants, whereas in pine, the decreased PSII photochemistry was likely due to sustained non-photochemical quenching. Downregulation of PSII photochemistry and maintenance of PSI in an oxidized state were linked with the prevention of oxidative stress, even under severe water deficit. The decreased photosynthetic pigment content and photosynthetic gene expression suggested the coordinated downregulation of photosynthetic apparatus components under water stress to reduce light energy absorption. In summary, the observed adaptative mechanisms of pine and spruce to water stress may be similar to the well-studied adaptative mechanisms to winter stress, which may indicate the universality of protective mechanisms under various stresses in conifers.

Magnesium-Deficiency Effects on Pigments, Photosynthesis and Photosynthetic Electron Transport of Leaves, and Nutrients of Leaf Blades and Veins in Citrus sinensis Seedlings.

Citrus sinensis seedlings were irrigated with nutrient solution at a concentration of 0 (Mg-deficiency) or 2 (Mg-sufficiency) mM Mg (NO3)2 for 16 weeks. Mg-deficiency-induced interveinal chlorosis, vein enlargement and corkiness, and alterations of gas exchange, pigments, chlorophyll a fluorescence (OJIP) transients and related parameters were observed in middle and lower leaves, especially in the latter, but not in upper leaves. Mg-deficiency might impair the whole photosynthetic electron transport, including structural damage to thylakoids, ungrouping of photosystem II (PSII), inactivation of oxygen-evolving complex (OEC) and reaction centers (RCs), increased reduction of primary quinone electron acceptor (QA) and plastoquinone pool at PSII acceptor side and oxidation of PSI end-electron acceptors, thus lowering energy transfer and absorption efficiency and the transfer of electrons to the dark reactions, hence, the rate of CO2 assimilation in Mg-deficiency middle and lower leaves. Although potassium, Mg, manganese and zinc concentration in blades displayed a significant and positive relationship with the corresponding element concentration in veins, respectively, great differences existed in Mg-deficiency-induced alterations of nutrient concentrations between leaf blades and veins. For example, Mg-deficiency increased boron level in the blades of upper leaves, decreased boron level in the blades of lower leaves, but did not affect boron level in the blades of middle leaves and veins of upper, middle and lower leaves. To conclude, Mg-deficiency-induced interveinal chlorosis, vein enlargement, and corkiness, and alterations to photosynthesis and related parameters increased with increasing leaf age. Mg-deficiency-induced enlargement and corkiness of veins were not caused by Mg-deficiency-induced boron-starvation.

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

Here, we developed a method for measuring the invivo 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.

Energetics of the exchangeable quinone, QB, in Photosystem II

Photosystem II (PSII), the light-driven water/plastoquinone photooxidoreductase, is of central importance in the planetary energy cycle. The product of the reaction, plastohydroquinone (PQH2), is released into the membrane from the QB site, where it is formed. A plastoquinone (PQ) from the membrane pool then binds into the QB site. Despite their functional importance, the thermodynamic properties of the PQ in the QB site, QB, in its different redox forms have received relatively little attention. Here we report the midpoint potentials (Em ) of QB in PSII from Thermosynechococcus elongatus using electron paramagnetic resonance (EPR) spectroscopy: Em QB/QB•- ≈ 90 mV, and Em QB•-/QBH2 ≈ 40 mV. These data allow the following conclusions: 1) The semiquinone, QB•-, is stabilized thermodynamically; 2) the resulting Em QB/QBH2 (∼65 mV) is lower than the Em PQ/PQH2 (∼117 mV), and the difference (ΔE ≈ 50 meV) represents the driving force for QBH2 release into the pool; 3) PQ is ∼50× more tightly bound than PQH2; and 4) the difference between the Em QB/QB•- measured here and the Em QA/QA•- from the literature is ∼234 meV, in principle corresponding to the driving force for electron transfer from QA•- to QB The pH dependence of the thermoluminescence associated with QB•- provided a functional estimate for this energy gap and gave a similar value (≥180 meV). These estimates are larger than the generally accepted value (∼70 meV), and this is discussed. The energetics of QB in PSII are comparable to those in the homologous purple bacterial reaction center.

Characterization of the transient fluorescence wave phenomenon that occurs during H2 production in Chlamydomonas reinhardtii.

The redox state of the plastoquinone (PQ) pool in sulfur-deprived, H2-producing Chlamydomonas reinhardtii cells was studied using single flash-induced variable fluorescence decay kinetics. During H2 production, the fluorescence decay kinetics exhibited an unusual post-illumination rise of variable fluorescence, giving a wave-like appearance. The wave showed the transient fluorescence minimum at ~60 ms after the flash, followed by a rise, reaching the transient fluorescence maximum at ~1 s after the flash, before decaying back to the initial fluorescence level. Similar wave-like fluorescence decay kinetics have been reported previously in anaerobically incubated cyanobacteria but not in green algae. From several different electron and proton transfer inhibitors used, polymyxin B, an inhibitor of type II NAD(P)H dehydrogenase (NDA2), had the effect of eliminating the fluorescence wave feature, indicating involvement of NDA2 in this phenomenon. This was further confirmed by the absence of the fluorescence wave in the Δnda2 mutant lacking NDA2. Additionally, Δnda2 mutants have also shown delayed and diminished H2 production (only 23% if compared with the wild type). Our results show that the fluorescence wave phenomenon in C. reinhardtii is observed under highly reducing conditions and is induced by the NDA2-mediated electron flow from the reduced stromal components to the PQ pool. Therefore, the fluorescence wave phenomenon is a sensitive probe for the complex network of redox reactions at the PQ pool level in the thylakoid membrane. It could be used in further characterization and improvement of the electron transfer pathways leading to H2 production in C. reinhardtii.