Photoinhibition Of PSII Research Articles

Betaine Improves Freezing Tolerance in Wheat

The accumulation of the osmolyte betaine was found to be correlated with the development of freezing tolerance (FT) of two wheat cultivars where it increases by about three fold during the cold' acclimation period. Exogenous betaine application resulted in a large increase in total osmolality mostly due to betaine accumulation. Plants that accumulated betaine are more tolerant to freezing stress since a four day exposure to 250 mM betaine resulted in a LT50 of -8° C (in spring wheat Glenlea) and -9° C (in winter wheat Fredrick) compared to — 3°C (Glenlea) and — 4°C (Fredrick) for control non-exposed plants. Betaine treatment (250 mM) during cold acclimation increased FT in an additive manner since the LT50 reached — 14°C (Glenlea) and -22° C (Fredrick) compared to -8° C (Glenlea) and — 16°C (Fredrick) for plants that are cold acclimated in the absence of betaine. These results show that betaine treatment can improve FT by more than 5°C in both non-acclimated and cold-acclimated plants. The betaine treatment resulted in the induction of a subset of low temperature responsive genes, such as the wcor410, and wcor413, that are also induced by salinity or drought stresses. In addition to these genetic responses, betaine treatment was also able to improve the tolerance to photoinhibition of PSII and the steady-state yield of electron transport over PSII in a manner that mimicked cold-acclimated plants. These data also suggest that betaine improves FT by eliciting some of the genetic and physiological responses associated with cold acclimation.

Temperature/light dependent development of selective resistance to photoinhibition of photosystem I

Exposure of winter rye leaves grown at 20°C and an irradiance of either 50 or 250 μmol m −2 s −1 to high light stress (1600 μmol m −2 s −1, 4 h) at 5°C resulted in photoinhibition of PSI measured in vivo as a 34% and 31% decrease in ΔA 820/A 820 (P700 +). The same effect was registered in plants grown at 5°C and 50 μmol m −2 s −1. This was accompanied by a parallel degradation of the PsaA/PsaB heterodimer, increase of the intersystem e − pool size as well as inhibition of PSII photochemistry measured as F v/F m. Surprisingly, plants acclimated to high light (800 μmol m −2 s −1) or to 5°C and moderate light (250 μmol m −2 s −1) were fully resistant to photoinhibition of PSI and did not exhibit any measurable changes at the level of PSI heterodimer abundance and intersystem e − pool size, although PSII photochemistry was reduced to 66% and 64% respectively. Thus, we show for the first time that PSI, unlike PSII, becomes completely resistant to photoinhibition when plants are acclimated to either 20°C/800 μmol m −2 s −1 or 5°C/250 μmol m −2 s −1 as a response to growth at elevated excitation pressure. The role of temperature/light dependent acclimation in the induction of selective tolerance to PSI photoinactivation is discussed.

The cause of PSI photoinhibition at low temperatures in leaves of Cucumis sativus, a chilling‐sensitive plant

In a chilling‐sensitive plant, cucumber, chilling of leaves in the light results in irreversible damage to PSI. Recent in vitro studies suggested that hydroxyl radicals, which are formed in the presence of H2O2 and reduced Fe‐S centers, are involved in the PSI inhibition. We therefore examined this possibility in vivo. Chilling of leaves at 5°C in the light caused a temporary increase in H2O2 concentration, which was probably due to the net H2O2 production in vivo. The activity, measured at 5°C, of the thylakoid ascorbate peroxidase (APX), a key enzyme of the H2O2‐scavenging system, was about 20% of that measured at 25°C. The isolated thylakoids retaining high thylakoid APX activity did not show light‐dependent net H2O2 production at 25°C. However, at 5°C, net production of H2O2 was observed. Since the rate of electron flow to molecular oxygen in the isolated thylakoids was ca 5 mmol e− mol−1 Chl s−1 at 5°C, the H2O2‐scavenging capacity was below this level. When intact leaves were illuminated at 5°C at an irradiance of 100 µmol m−2 s−1, the rate of electron transport through PSII was ca 20 mmol e− mol−1 Chl s−1 and more than 80% of QA was in the reduced state. Since thylakoids are uncoupled in cucumber leaves at 5°C in the light. ATP is not formed and energy dissipation in the form of heat is suppressed. Therefore, the electron flow to molecular oxygen would be greater than 5 mmol e− mol−1 Chl s−1. Moreover, under such conditions, components in the electron transport chain, including Fe‐S centers in PSI, were probably reduced. These features indicate that, when cucumber leaves are chilled in the light, hydroxyl radicals can be produced by the Fenton reaction and cause damage to PSI.

Photoinhibition of Photosystem I: Its Physiological Significance in the Chilling Sensitivity of Plants

Photoinhibition was defined originally as the decrease in photosynthetic activity that occurs upon excess illumination. The site of photoinhibition has generally been considered to be located in PSII. However, a novel type of photoinhibition has recently been characterized in chillingsensitive plants. This photoinhibition occurs under relatively weak illumination at chilling temperatures and the main site of damage is in PSI. The photoinhibition of PSI is initiated by the inactivation of the acceptor side, with the subsequent destruction of the reaction center and the degradation of the product of the psaB gene, which is one of the two major subunit polypeptides of the PSI reaction center complex. Chilling and oxidative stress (the presence of reactive species of oxygen) are characteristic requirements for the photoinhibition of PSI in vivo.

Over-production of the D1:2 protein makes Synechococcus cells more tolerant to photoinhibition of photosystem II.

Over-expression of the psbAIII gene encoding for the D1 protein (form II; D1:2) of the photosystem II reaction centre in the Synechococcus sp. PCC 7942 was studied using a tac promoter and the lacIQ system. Over-expression was induced with 40 microgram/ml IPTG in the growth medium for either 6 or 12 h at growth irradiance (50 mumol photons m-2 s-1). This treatment doubled the amount of psbAII/III mRNA and the D1:2 protein in membranes but decreased the amount of psbAI messages and the D1:1 protein. The total amount of both heterodimeric reaction centre proteins, D1 and D2, remained constant under growth light conditions, indicating that the number of PSII centres in the membranes was not affected, only the form of the D1 protein was changed from D1:1 to D1:2 in most centres. When the cells were photoinhibited either at 500 or 1000 mumol photons m-2 s-1, in the presence or absence of the protein synthesis inhibitor lincomycin, the D1:2 protein remained at a higher level in cells in which over-expression had been induced by IPTG. These cells were also less prone to photoinhibition of PSII. It is suggested that the tolerance of cells to photoinhibition increases when most PSII reaction centres contain the D1:2 protein at the beginning of high irradiance. This tolerance is further strengthened by maintaining psbAIII gene over-expression during the photoinhibitory treatment.

Abscisic acid induced protection against photoinhibition of PSII correlates with enhanced activity of the xanthophyll cycle

The exogenous application of abscisic acid (ABA) to barley seedlings resulted in partial protection of the PSII photochemistry against photoinhibition at low temperature, the effect being most pronounced at 10 −5 M ABA. This was accompanied by higher photochemical quenching (qP) in ABA-treated leaves. A considerable increase (122%) in the amount of total carotenoids and xanthophylls (antheraxanthin, violaxanthin and zeaxanthin) was also found in the seedlings subjected to ABA. The activity of the xanthophyll cycle measured by the epoxidation state of xanthophylls under high-light treatment was higher in ABA-treated plants compared with the control. This corresponds to a higher value (0.411) of non-photochemical quenching (qNP) observed in ABA-treated than in control (0.306) leaves.

Regulation of D1-protein degradation during photoinhibition of photosystem II in vivo: Phosphorylation of the D1 protein in various plant groups

Photoinhibition of PSII and turnover of the D1 reaction-centre protein in vivo were studied in pumpkin leaves (Cucurbita pepo L.) acclimated to different growth irradiances and in low-light-grown moss, (Ceratodon purpureus) (Hedw.) Brid. The low-light-acclimated pumpkins were most susceptible to photoinhibition. The production rate of photoinhibited PSII centres (kPI), determined in the presence of a chloroplast-encoded protein-synthesis inhibitor, showed no marked difference between the high- and low-light-grown pumpkin leaves. On the other hand, the rate constant for the repair cycle (kREC) of PSII was nearly three times higher in the high-light-grown pumpkin when compared to low-light-grown pumpkin. The slower degradation rate of the damaged D1 protein in the low-light-acclimated leaves, determined by pulsechase experiments with [35S]methionine suggested that the degradation of the Dl protein retards the repair cycle of PSII under photoinhibitory light. Slow degradation of the D1 protein in low-light-grown pumpkin was accompanied by accumulation of a phosphorylated form of the D1 protein, which we postulate as being involved in the regulation of D1-protein degradation and therefore the whole PSII repair cycle. In spite of low growth irradiance the repair cycle of PSII in the moss Ceratodon was rapid under high irradiance. When compared to the high- or low-light-acclimated pumpkin leaves, Ceratodon had the highest rate of D1-protein degradation at 1000 μmol photons m−2 s−1. In contrast to the higher plants, the D1 protein of Ceratodon was not phosphorylated either under high irradiance in vivo or under in-vitro conditions, which readily phosphorylate the D1 protein of higher plants. This is consistent with the rapid degradation of the D1 protein in Ceratodon. Screening experiments indicated that D1 protein can be phosphorylated in the thylakoid membranes of angiosperms and conifers but not in lower plants. The postulated regulation mechanism of D1-protein degradation involving phosphorylation and the role of thylakoid organization in the function of PSII repair cycle are discussed.

Mechanism of photosystem-I photoinhibition in leaves of Cucumis sativus L.

It was recently shown that the site of photoinhibition in leaves of Cucumis sativus L. at low temperatures is Photosystem I (PSI), not PSII (I. Terashima et al. 1994, Planta 193, 300–306). In the present study, the mechanisms of this PSI photoinhibition in vivo were examined. By lowering the photon flux density during the photoinhibitory treatment of leaves at 4°C for 5 h to less than 100 μmol·m−2s−1, we were able to separate the steps of the destruction of the electron-transfer components. Although P-700, the reaction-center chlorophyll, was almost intact in this low-light treatment, the quantum yield of the electron transfer through PSI and photochemically induced absorption change at 701 nm were markedly inhibited. This, along with the results from the measurements of the light-induced absorption changes in the presence of various concentrations of methyl viologen, an artificial electron acceptor, indicates that the component on the acceptor side of the PSI, A1 or Fx, is the first site of inactivation. When the photon flux density during the treatment was increased to 220 μmol·m−2s−1, the destruction of P-700 itself was also observed. Furthermore, the partial degradation of the chlorophyll-binding large subunits was observed in photoinhibited leaves. This degradation of the subunits was not detected when the treatment was carried out under nitrogen atmosphere, the condition in which the electron transfer is not inhibited. Thus, the photoinhibitory processes in the reaction center of PSI go through three steps, the inactivation of the acceptor side, the destruction of the reaction-center chlorophyll and the degradation of the reaction center subunit(s). The similarities and the differences between the mechanisms of PSI photoinhibition and those of PSII photoinhibition are discussed.

The site of photoinhibition in leaves of Cucumis sativus L. at low temperatures is photosystem I, not photosystem II

Maximum quantum yields (QY) of photosynthetic electron flows through PSI and PSII were separately assessed in thylakoid membranes isolated from leaves of Cucumis sativus L. (cucumber) that had been chilled in various ways. The QY(PSI) in the thylakoids prepared from the leaves treated at 4° C in moderate light at 220 μmol quanta·m−2·s−1 (400–700 nm) for 5 h, was about 20–30% of that in the thylakoids prepared from untreated leaves, while QY(PSII) decreased, at most, by 20% in response to the same treatment. The decrease in QY(PSI) was observed only when the leaves were chilled at temperatures below 10° C, while such a marked temperature dependency was not observed for the decrease in QY(PSII). In the chilling treatment at 4° C for 5 h, the quantum flux density that was required to induce 50% loss of QY (PSI) was ca. 50 umol quanta·m−2·s−1. When the chilling treatment at 4° C in the light was conducted in an atmosphere of N2, photoinhibition of PSI was largely suppressed, while the damage to PSII was somewhat enhanced. The ferricyanide-oxidised minus ascorbate-reduced difference spectra and the light-induced absorbance changes at 700 nm obtained with the thylakoid suspension, indicated the loss of P700 to extents that corresponded to the decreases in QY(PSI). Accordingly, the decreases in QY(PSI) can largely be attributed to destruction of the PSI reaction centre itself. These results clearly show that, at least in cucumber, a typical chillingsensitive plant, PSI is much more susceptible to aerobic photoinhibition than PSII.

Photosynthesis and photoinhibition in leaves of chlorophyll b‐less barley in relation to absorbed light

The response of photosynthesis to absorbed light by intact leaves of wild‐type (Hordeum vulgare L. cv. Gunilla) and chlorophyll b‐less barley (H. vulgare L. cv. Dornaria, chlorina‐f22800) was measured in a light integrating sphere. Up to the section where the light response curve bends most sharply the responses of the b‐less and wild‐type barley were similar but not identical. Average quantum yield and convexity for the mutant light response curves were 0.89 and 0.90, respectively, times those of the wild‐type barley. The maximum quantum yield for PSII photochemistry was also 10% lower as indicated by fluorescence induction kinetics (Fv/Fm). Just above the region where the light curve bends most sharply, photosynthesis decreased with time in the mutant but not in the wild‐type barley. This decrease was associated with a decrease in Fv/Fm indicating photoinhibition of PSII. This photoinhibition occurred in the same region of the light response curve where zeaxanthin formation occurs. Zeaxanthin formation occurred in both the chlorophyll b‐less and wild‐type leaves. However, the epoxidation state was lower in the mutant than in the wild‐type barley. The results indicate that chlorophyll b‐less mutants will have reduced photosynthetic production as a result of an increased sensitivity to photoinhibition and possibly a lowered quantum yield and convexity in the absence of photoinhibition.

Temperature‐dependent changes in Photosystem II heterogeneity of attached leaves under high light

Attached leaves of pumpkin (Cucurbita pepo L. cv. Jattiläismeloni) were exposed to high light intensity at room temperature (ca 23°C) and at 1°C. Fluorescence parameters and electron transport activities measured from isolated thylakoids indicated faster photoinhibition of PSII at low temperature. Separation of the α and β components of the complementary area above the fluorescence induction curve of dichlorophenyl-dimethylurea-poisoned thylakoids revealed that at low temperature only the α-centers declined during exposure to high light intensity while the content of functional β-centers remained constant. Freeze-fracture electron microscopy showed no decrease in the density of particles on the appressed exoplasmic fracture face, indicating that the photoinhibited α-centers remained in the appressed membranes at 1°C. Because of the function of the repair and protective mechanisms of PSII, strong light induced less photoinhibition at room temperature, but more complicated changes occurred in the α/β-heterogeneity of PSII. During the first 30 min at high light intensity the decrease in α-centers was almost as large as at 1°C, but in contrast to the situation at low temperature the decrease in α-centers was compensated for by a significant increase in PSIIβ-centers. Changes in the density and size of freeze-fracture particles suggest that this increase in β-centers was due to migration of phosphorylated light-harvesting complex from appressed to non-appressed thylakoid membranes while the PSII core remained in the appressed membranes. This situation, however, was only transient and was followed by a rapid decrease in the functionalβ-centers.

Cytochrome b-559 may function to protect photosystem II from photoinhibition.

Although cytochrome b-559 is an integral component of the photosystem II complex (PSII), its function is unknown. Because cytochrome b-559 has been shown to be both photooxidized and photoreduced in PSII, one of several proposals is that it mediates cyclic electron transfer around PSII, possibly as a protective mechanism. We have used electron paramagnetic resonance spectroscopy to investigate the pathway of photooxidation of cytochrome b-559 in PSII and have shown that it proceeds via photooxidation of chlorophyll. We propose that this photooxidation of chlorophyll is the first step in the photoinhibition of PSII. The unique susceptibility of PSII to photoinhibition is probably due to the fact that it is the only reaction center in photosynthesis which generates an oxidant with a reduction potential high enough to oxidize chlorophyll. We propose that the function of cytochrome b-559 is to mediate cyclic electron transfer to rereduce photooxidized chlorophyll and protect PSII from photoinhibition. We also suggest that the chlorophyll(s) which are susceptible to photooxidation are analogous to the monomer chlorophylls found in the bacterial photosynthetic reaction center complex.