Pierre Joliot

UN NOUVEAU MODELE DES CENTRES PHOTOCHIMIQUES DU SYSTEME II

Abstract— The amount of oxygen evolved by Chlorella cells or by isolated chloroplasts has been measured after illumination by short saturating flashes. In all conditions, the amount of oxygen evolved by one flash is proportional to the fraction of the photochemical centers susceptible to produce oxygen.

A complex containing PGRL1 and PGR5 is involved in the switch between linear and cyclic electron flow in Arabidopsis.

During photosynthesis, two photoreaction centers located in the thylakoid membranes of the chloroplast, photosystems I and II (PSI and PSII), use light energy to mobilize electrons to generate ATP and NADPH. Different modes of electron flow exist, of which the linear electron flow is driven by PSI and PSII, generating ATP and NADPH, whereas the cyclic electron flow (CEF) only generates ATP and is driven by the PSI alone. Different environmental and metabolic conditions require the adjustment of ATP/NADPH ratios and a switch of electron distribution between the two photosystems. With the exception of PGR5, other components facilitating CEF are unknown. Here, we report the identification of PGRL1, a transmembrane protein present in thylakoids of Arabidopsis thaliana. Plants lacking PGRL1 show perturbation of CEF, similar to PGR5-deficient plants. We find that PGRL1 and PGR5 interact physically and associate with PSI. We therefore propose that the PGRL1-PGR5 complex facilitates CEF in eukaryotes.

Cyclic electron transfer in plant leaf

The turnover of linear and cyclic electron flows has been determined in fragments of dark-adapted spinach leaf by measuring the kinetics of fluorescence yield and of the transmembrane electrical potential changes under saturating illumination. When Photosystem (PS) II is inhibited, a cyclic electron flow around PSI operates transiently at a rate close to the maximum turnover of photosynthesis. When PSII is active, the cyclic flow operates with a similar rate during the first seconds of illumination. The high efficiency of the cyclic pathway implies that the cyclic and the linear transfer chains are structurally isolated one from the other. We propose that the cyclic pathway operates within a supercomplex including one PSI, one cytochrome bf complex, one plastocyanin, and one ferredoxin. The cyclic process induces the synthesis of ATP needed for the activation of the Benson–Calvin cycle. A fraction of PSI (∼50%), not included in the supercomplexes, participates in the linear pathway. The illumination would induce a dissociation of the supercomplexes that progressively increases the fraction of PSI involved in the linear pathway.

Evidence for two active branches for electron transfer in photosystem I.

All photosynthetic reaction centers share a common structural theme. Two related, integral membrane polypeptides sequester electron transfer cofactors into two quasi-symmetrical branches, each of which incorporates a quinone. In type II reaction centers [photosystem (PS) II and proteobacterial reaction centers], electron transfer proceeds down only one of the branches, and the mobile quinone on the other branch is used as a terminal acceptor. PS I uses iron-sulfur clusters as terminal acceptors, and the quinone serves only as an intermediary in electron transfer. Much effort has been devoted to understanding the unidirectionality of electron transport in type II reaction centers, and it was widely thought that PS I would share this feature. We have tested this idea by examining in vivo kinetics of electron transfer from the quinone in mutant PS I reaction centers. This transfer is associated with two kinetic components, and we show that mutation of a residue near the quinone in one branch specifically affects the faster component, while the corresponding mutation in the other branch specifically affects the slower component. We conclude that both electron transfer branches in PS I are active.

STUDIES OF SYSTEM II PHOTOCENTERS BY COMPARATIVE MEASUREMENTS OF LUMINESCENCE, FLUORESCENCE, AND OXYGEN EMISSION*

Abstract— New results are presented on the emission of oxygen by algae and chloroplasts illuminated by a sequence of short saturating flashes. These results favor the four‐state hypothesis of Kok and co‐workers, in which formation of oxygen requires the accumulation of four oxidants produced by four successive photoreactions. Deactivation of the more oxidized precursor states in the dark is studied under different conditions of preillumination. Our results suggest that both a one step and a two step mechanism of deactivation exist. In order to understand the biological significance of Koks parameter α—the fraction of photochemical centers unable to react on each flash (“misses”)‐we study reoxidation of acceptor Q after one flash by fluorescence techniques. It appears that a fraction of Q‐ is reoxidized by a back reaction which cancels the effect of the preilluminating flash and is probably responsible for the misses. The results of some luminescence experiments are also reported. These experiments demonstrate that delayed emission of light is associated with the deactivation of states S2 and S3. It is possible that excitons produced by deactivation can be reabsorbed by active photochemical centers, which can modify considerably the deactivation process.

Supramolecular organization of the photosynthetic apparatus of Rhodobacter sphaeroides.

Native tubular membranes were purified from the purple non‐sulfur bacterium Rhodobacter sphaeroides. These tubular structures contain all the membrane components of the photosynthetic apparatus, in the relative ratio of one cytochrome bc1 complex to two reaction centers, and ∼24 bacteriochlorophyll molecules per reaction center. Electron micrographs of negative‐stained membranes diffract up to 25 Å and allow the calculation of a projection map at 20 Å. The unit cell (a = 198 Å, b = 120 Å and γ = 103°) contains an elongated S‐shaped supercomplex presenting a pseudo‐2‐fold symmetry. Comparison with density maps of isolated reaction center and light‐harvesting complexes allowed interpretation of the projection map. Each supercomplex is composed of light‐harvesting 1 complexes that take the form of two C‐shaped structures of ∼112 Å in external diameter, facing each other on the open side and enclosing the two reaction centers. The remaining positive density is tentatively attributed to one cytochrome bc1 complex. These features shed new light on the association of the reaction center and the light‐harvesting complexes. In particular, the organization of the light‐harvesting complexes in C‐shaped structures ensures an efficient exchange of ubihydroquinone/ubiquinone between the reaction center and the cytochrome bc1 complex.

Regulation of cyclic and linear electron flow in higher plants

Cyclic electron flow is increasingly recognized as being essential in plant growth, generating a pH gradient across thylakoid membrane (ΔpH) that contributes to ATP synthesis and triggers the protective process of nonphotochemical quenching (NPQ) under stress conditions. Here, we report experiments demonstrating the importance of that ΔpH in protecting plants from stress and relating to the regulation of cyclic relative to linear flow. In leaves infiltrated with low concentrations of nigericin, which dissipates the ΔpH without significantly affecting the potential gradient, thereby maintaining ATP synthesis, the extent of NPQ was markedly lower, reflecting the lower ΔpH. At the same time, the photosystem (PS) I primary donor P700 was largely reduced in the light, in contrast to control conditions where increasing light progressively oxidized P700, due to down-regulation of the cytochrome bf complex. Illumination of nigericin-infiltrated leaves resulted in photoinhibition of PSII but also, more markedly, of PSI. Plants lacking ferredoxin (Fd) NADP oxidoreductase (FNR) or the polypeptide proton gradient regulation 5 (PGR5) also show reduction of P700 in the light and increased sensitivity to PSI photoinhibition, demonstrating that the regulation of the cytochrome bf complex (cyt bf) is essential for protection of PSI from light stress. The formation of a ΔpH is concluded to be essential to that regulation, with cyclic electron flow playing a vital, previously poorly appreciated role in this protective process. Examination of cyclic electron flow in plants with a reduced content of FNR shows that these antisense plants are less able to maintain a steady rate of this pathway. This reduction is suggested to reflect a change in the distribution of FNR from cyclic to linear flow, likely reflecting the formation or disassembly of FNR–cytochrome bf complex.

Flash-induced 519 nm absorption change in green algae

Abstract 1. The 515 nm absorption change induced by isolated flashes was studied in Chlorella and Chlamydomonas, using a new spectrophotometric method. A strong actinic flash is followed by a weak monochromatic detecting flash (duration 3 μs) which samples the absorption level. 2. The time course of the absorption change after one actinic flash in darkadapted algae shows four phases: Phase a: a large absorption increase ( 2 3 of the maximum) occurs in less than 1 μs. 60–80% of this absorption increase is linked to Photosystem I activity, and 20–40% to Photosystem II. Phase b: a slow absorption increase occurs in the time range of 1–50 ms. Phase b is derived only from Photosystem I. Phase c: a fast absorption decrease is observed between 50 and 500 ms. Phase c is absent in Chlamydomonas mutant (F54) which is blocked at a terminal stage of phosphorylation. Phase d: a slow absorption decrease is observed between 500 ms and 10 s. 3. The turnover times of the reactions driving the absorption change were studied. The turnover times of both photosystems can be distinguished; a fast turnover time ( t 1 2 = 100 μ s ) is observed for Photosystem I. 4. The experimental results are discussed in terms of the chemiosmotic theory of Mitchell (Mitchell, P. (1961) Nature 191, 144–148) and the electrochromic hypothesis of Witt et al. Our results suggest that phosphorylation in Chlorella can be driven by an electrical potential only. This is in agreement with formal results obtained in spinach chloroplasts (Witt, H. T. (1971) Q. Rev. Biophys. 4, 365–477). 5. In agreement with Junge et al., we interpret the biphasicity of the absorption decay by the existence of a critical potential for phosphorylation; but we observe that the value of the critical potential depends on physiological conditions, and can be close to zero.

Cinétiques des réactions liées a l'émission d'oxygène photosynthétique

Abstract The transient kinetics of O2 emission by Chlorella pyrenoidosa have been studied by measuring dissolved O2 with a sensitive and rapid amperometric method. Highly precise measurements during the first few seconds following illumination show an O2 gush with a span of about 1 sec; the gush is preceded by a short activation phase during which the velocity of O2 emission increases rapidly. The kinetics are governed only by one photochemical rate constant (System II) and several thermal rate constants which have been evaluated. Measurement of the initial velocity of O2 emission permits to obtain a very precise action spectrum of System II. The concentration of the photochemical complex taking part in the reaction is proportional to the quantity of O2 evolved following a short electronic flash, whereas O2 evolved during the gush gives a measure of the concentration of an intermediate which leads to regenerate this complex. The compounds are reformed in the presence of light and their concentrations attain maximal values parallely with the velocity of O2 emission. Their formation is linked to a photochemical reaction sensibilised at high wavelength (System I) and their destruction to the one giving rise to O2 formation (System II). The author proposes an interpretation of the ensemble of kinetics phenomena. The concentration of various supposed intermediates implicated, as well as certain thermal rate constants involved in the process of O2 emission, have been evaluated.

Plastoquinone compartmentation in chloroplasts. I: Evidence for domains with different rates of photo-reduction

The photo-reduction of plastoquinones by Photosystem II reaction centers was investigated using fluorescence and oxygen-evolution measurements in thylakoids deprived of Photosystem I acceptors. The process appears biphasic under limiting, as well as saturating, illumination. The ‘fast’ pool fraction (about 6 PQ molecules per PS II center) represents 50–70% of the total. Its half reduction time under saturating light was found about 25–60 ms, while that of the ‘slow’ pool was 0.8–1 s. When the photo-reduction process is interrupted after reduction of the fast pool and resumed after a dark period, a redistribution of the reduced plastoquinones towards the slow pool is observed, with t12 ≈ 6 s. We interpret these results as expressing a limitation of PQ diffusion in the membrane and propose that the fast pool reflects the fraction present in the grana region where most PS II centers are located, while the slow pool corresponds to quinones from the stromal region. The relationship between the redox states of PQ and of the primary acceptor QA during photo-reduction of the fast pool expresses marked discrepancies with respect to a quasi-equilibrium relationship. This failure to achieve equilibrium on a rapid time-scale and the slow diffusion rate of quinones over long distances are accounted for by small size domains bounded by membrane proteins. In agreement with this view, we found that the amount of fast photo-reducible quinones is decreased when a fraction of PS II centers is inhibited, indicating that the domains contain, on average, about 3–4 PS II centers. We conclude that PQ cannot be responsible for the long range diffusion involved in rapid electron transfer from granal (PS II) to stromal (PS II) regions, a role that must be fulfilled by plastocyanin.