Improvement of Cerium of Photosynthesis Functions of Maize Under Magnesium Deficiency

Rare earth elements can promote photosynthesis, but their mechanisms are still poorly understood under magnesium deficiency. The present study was designed to determine the role of cerium in magnesium-deficient maize plants. Maize was cultivated in Hoagland’s solution added with cerium with and without adequate quantities of magnesium. Under magnesium-deficient conditions, cerium can prevents inhibition of synthesis of photosynthetic pigment, improves light energy absorption and conversion, oxygen evolution, and the activity of photo-phosphorelation and its coupling factor Ca²⁺-ATPase. These results suggest that cerium could partly substitute magnesium, improving photosynthesis and plant growth.     

The roles of weak light,oxygen and dithiothreitol in the photoreactivation of the oxygen evolving center of tris-washed and 2, 6-dichlorophenol indophenol-treated chloroplasts

The inactivated O₂-evolving center of Tris-washed chloroplasts was reactivated by DCPIP-treatment and photoreactivation in the presence of Mn²⁺, Ca²⁺, DTT and weak light. Many electron donors (Asc and reduced DCPIP, etc.) were found to be suitable substitutes for DTT. By studying the anaerobic inhibition of the reactivation, the electron acceptors O₂, NADP⁺, etc. were also found to be essential factors in photoreactivation. Weak light stimulated the chloroplast electron transport from the above-mentioned electron donors to the electron acceptor and effected the photoreactivation. More than 280 electrons were transported to NADP⁺ in the anaerobic photoreactivation of one unit of an O₂-evolving center with 400 Chl. Electron transport in the reactivation was inhibited by omitting DTT or Mn²⁺ ion, and by adding DCMU. The photoreactivated chloroplasts incorporated about 2 Mn by 400 Chl. Omission of DTT in the reactivation caused chloroplasts in the weak light to bind large amounts of excess Mn.Abbreviations Asc ascorbate - Chl chlorophyll - DCPIP 2, 6-dichlorophenol indophenol - DPC diphenyl carbazide - DTT dithiothreitol - Fd ferredoxin - STN a chloroplast preparation medium, containing 0.4 M sucrose, 0.05 M Tris-Cl and 0.01 M NaCl (pH 7.8 and 8.0) - TMPD tetramethyl-p-phenylenediamine     

Inhibition of the oxygen-evolving complex of photosystem II and depletion of extrinsic polypeptides by nickel

The toxic effect of Ni²⁺ on photosynthetic electron transport was studied in a photosystem II submembrane fraction. It was shown that Ni²⁺ strongly inhibits oxygen evolution in the millimolar range of concentration. The inhibition was insensitive to NaCl but significantly decreased in the presence of CaCl₂. Maximal chlorophyll fluorescence, together with variable fluorescence, maximal quantum yield of photosystem II, and flash-induced fluorescence decays were all significantly declined by Ni²⁺. Further, the extrinsic polypeptides of 16 and 24 kDa associated with the oxygen-evolving complex of photosystem II were depleted following Ni²⁺ treatment. It was deduced that interaction of Ni²⁺ with these polypeptides caused a conformational change that induced their release together with Ca²⁺ from the oxygen-evolving complex of photosystem II with consequent inhibition of the electron transport activity.     

Electron spin resonance and photoreaction of Mn(II) in lettuce chloroplasts.

Electron spin resonance (esr) of lettuce chloroplasts yields three types of signals: (i) a broad (~900 G) signal around g = 2.22 (apparently due to Cu²⁺ complexes); (ii) an Mn²⁺ spectrum around g = 2.003 consisting of six hyperfine lines (A = 94.5 G) of ~30 G width; and (iii) a sharp signal at g = 2.00 due to photosignals I and II. The present work is concerned with the Mn²⁺ signal and its relation to the photosynthetic process. Intensity measurements were performed by comparing the intensities of the Mn²⁺ signals of two identical chloroplast preparations, one of which was slightly acidified. The integrated intensity of the signal in the normal preparation was approximately one-fourth of that in the acidified sample, suggesting that only the?$\text{1}\text{2}\text{?}\text{1}\text{2}$ fine structure band is observed in untreated chloroplasts. This indicates that the manganese in the chloroplasts is bound in an asymmetric environment, apparently in protein complexes. The Mn²⁺ signal is light sensitive, decreasing on illumination and reappearing in the dark. Typical values for the half-lives of the light and dark processes in normal chloroplasts are 0.25 and 2.1s, respectively. The effect is interpreted in terms of the photooxidation of Mn²⁺ to higher oxidation states which are invisible to esr spectroscopy. In order to determine whether this process is related to photosynthesis the effect of certain reagents and treatments that are known to affect the photosynthetic system was studied. It was found that the oxygen evolution inhibitors 3-(3,4 dichlorophenyl)-1,1-dimethylurea (DCMU) and carbonylcyanide-p-trifluoromethoxyphenylhydrazone (FCCP) as well as the electron donors, phenylenediamine and sodium ascorbate, reduce or completely eliminate the light effect on the Mn²⁺ signal. Heat treatment and Tris washing caused deceleration of both the light and dark reactions. These effects indicate that the photooxidation of the Mn²⁺ is related to the photosynthetic cycle, the most probable site being the water splitting apparatus of photosystem II.     

Studies on the manganese of the chloroplast

Manganese deficiency of green plants is known to affect preferentially the activity of the oxygen evolving system in the photosynthetic apparatus. Our studies showed that the time needed to reactivate photosynthesis in Mn-deficient algae varies with each culture, and is often very short when Mn is added not before illumination but during the light period. The recent finding by Cheniae and Martin that the reactivation requires light, is confirmed. The plain incorporation of ⁵⁴Mn into deficient algae as distinguished from reactivation was barely affected by light, yet was inhibited by uncouplers of phosphorylation. Higher plants responded to manganese deficiency either by adjusting the number of chloroplasts per cell to the limited Mn supply, or by forming disorganized chloroplasts with low chlorophyll content. These 2 types of responses produced chlorotic plants which had either a few photosynthetically active or many disabled chloroplasts. Photosystem I mediated photophosphorylation turned out to be much more sensitive to manganese deficiency than the system I dependent photoreduction of NADP⁺.     

Characteristics of the Mg-ATPase Activity Associated with the Membrane-Bound Maize Coupling Factor

The membrane-bound coupling factor of maize mesophyll thylakoids is a latent ATPase. Mg²⁺-ATPase activity can be induced in the light with either dithiothreitol or low concentrations of trypsin. Maize thylakoids that are activated with light plus trypsin exhibit considerably higher levels of activity in Na₂SO₃-dependent Mg²⁺-ATPase assays compared to thylakoids that are light and dithiothreitol activated (1400 micromoles per milligram of chlorophyll per hour versus 200 micromoles per milligram of chlorophyll per hour). Treatment with light and dithiothreitol or light plus trypsin were also required to demonstrate high levels of octyl glucoside-dependent Mg²⁺-ATPase activity in maize mesophyll thylakoids. Only small differences in octyl glucoside-dependent Mg²⁺-ATPase activity were observed in preparations that were activated in the light with either trypsin or dithiothreitol. Mg²⁺-ATPase activity can also be induced in maize mesophyll chloroplasts by illuminating intact preparations under appropriate conditions. Little or no ATPase activity was observed in the absence of illumination or in the presence of light plus methyl viologen. The active state decayed in the dark with a t_½ of 6 to 7 minutes at room temperature. Based on the effect of the thiol oxidant, o-iodosobenzoate, and the uncoupler, nigericin, on the kinetics of deactivation of ATPase activity in intact maize chloroplasts, it appears that the activation process requires a transmembrane proton gradient and reduction of a key disulfide bridge in the gamma of chloroplast coupling factor one.     

Heat inactivation of oxygen evolution in Photosystem II particles and its acceleration by chloride depletion and exogenous manganese

Heat inactivation of oxygen evolution by isolated Photosystem II particles was accelerated by Cl⁻ depletion and exogenous Mn²⁺. Weak red light also accelerated heat inactivation. Heat treatment released the 33, 24 and 18 kDa proteins and Mn from the Photosystem II particles. The protein release was stimulated by Cl⁻ depletion and exogenous Mn²⁺, and the Mn release was also stimulated by Cl⁻ depletion. A 50% loss of Mn corresponded to full inactivation of oxygen evolution, whereas no direct correlation seemed to exist between the loss of any one protein and inactivation of oxygen evolution. Removal of the 24 and 18 kDa proteins from photosystem II particles only slightly decreased the heat stability of oxygen evolution.     

Regulation of chloroplast photosynthetic activity by exogenous magnesium

Magnesium was most inhibitory to photosynthetic reactions by intact chloroplasts when the magnesium was added in the dark before illumination. Two millimolar MgCl₂, added in the dark, inhibited CO₂-dependent O₂ evolution by Hordeum vulgare L. and Spinacia oleracea L. (C₃ plants) chloroplasts 70 to 100% and inhibited (pyruvate + oxaloacetate)-dependent O₂ evolution by Digitaria sanguinalis L. (C₄ plant) mesophyll chloroplasts from 80 to 100%. When Mg²⁺ was added in the light, O₂ evolution was reduced only slightly. O₂ evolution in the presence of phosphoglycerate was less sensitive to Mg²⁺ inhibition than was CO₂-dependent O₂ evolution.

Magnesium prevented the light activation of several photosynthetic enzymes. Two millimolar Mg²⁺ blocked the light activation of NADP-malate dehydrogenase in D. sanguinalis mesophyll chloroplasts, and the light activation of phosphoribulokinase, NADP-linked glyceraldehyde-3-phosphate dehydrogenase, and fructose 1,6-diphosphatase in barley chloroplasts. The results suggest that Mg²⁺ inhibits chloroplast photosynthesis by preventing the light activation of certain enzymes.

     

Differential efficiency of photosynthetic oxygen evolution in flashing light in triazine-resistant and triazine-susceptible biotypes of Senecio vulgaris L.

Photosynthetic oxygen evolution in response to flashing light was studied in triazine-susceptible and triazine-resistant biotypes of Senecio vulgaris L. Studies were conducted to determine if the modification of the herbicide-binding site which confers s-triazine resistance also affects the oxygen-evolving system. Oxygen evolution was measured using a Joliot-type oxygen-specific electrode on broken, stroma-free chloroplasts of both biotypes. We observed abnormal patterns of oxygen evolution in resistant chloroplasts. The S′₁ → S₂ transition is slower while the S₂ decay is faster. The S′₂ → S₃ transition, in contrast, is slightly faster in resistant chloroplasts, while the decay of the S₃ state is the same as in susceptible chloroplasts. These altered kinetics may be due to altered Q → B (B^?) electron flow in resistant chloroplasts. These results are also consistent with the hypothesis that back-reactions from the reducing (acceptor) side of Photosystem II to the oxidizing (donor) side occur with greater frequency in resistant than susceptible chloroplasts. These events are responsible for lower oxygen yield and increased ‘misses’ and ‘double hits,’ resulting in abnormal yield patterns and lower quantum yield of CO₂ fixation in resistant chloroplasts compared to the susceptible ones.     

Photoinhibition in vivo of Photosystem II reactions during developmentof the photosystems of wheat seedlings

Photoinhibition of O₂ evolution and reactions leading to millisecond-delayed light emission (ms-DLE) of chlorophyll by illumination of leaves with excess white light were investigated in wheat seedlings greened for different times in a special chamber with constant conditions (20°C; CO₂ and humidity). A sharp reduction in initial and steady state rates of O₂ evolution and in the intensity of different components of ms-DLE under excess light on the stage of lag-phase of chlorophyll biosynthesis (4–6h of greening) were observed. An increasing stability of the oxygen-evolving process and ms-DLE of chlorophyll during formation of the thylakoid membrane photosystems (12–24 h of greening) was shown. Rifampicin did not influence the stability of oxygen evolution whereas cycloheximide led to the intensification of photoinhibition of the initial and steady-state rates of oxygen evolution under the inhibitory light action. The early stages of photosystems formation during short time of greening of etiolated seedlings were more sensitive to the action of inhibitory light, possibly due to a weak interaction of the oxygen-evolving system components and connection with reaction centers of Photosystem II.     

Effects of lanthanum and calcium on photoelectron transport activity and the related protein complexes in chloroplast of cucumber leaves

The effects of lanthanum and calcium ions on electron transport, dichlorephenol indophenol (DCIP) photoreduction, and oxygen evolution activities in chloroplast from cucumber (Cucumis satives L.) were determined. The lanthanum inhibited the whole electron chain-transport activity of chloroplast. DCIP photoreduction and oxygen evolution activities of the photosystem I (PSII) also decrease after treatment with La³⁺. But the diminished activities of PSII and chloroplast caused by La³⁺ could be reversed by Ca²⁺ and even became higher than the control level. The concentration analysis of related protein complexes to photoelectron transport in chloroplast included that La³⁺ induced the concentration of chlorophyll protein complexes increasing but caused some nonchlorophyll protein complexes to decompose partially. This increasing effect of La³⁺ on chlorophyll protein complexes results in the improvement of chlorophyll content, which will improve the absorption of photoelectron and energy transport in the process of photosynthesis.     

铅胁迫对玉米幼苗叶片光系统功能及光合作用的影响

通过同时测定玉米叶片的叶绿素荧光快速诱导动力学曲线和对820 nm光的吸收、分析叶片的气体交换过程以及叶绿体活性氧清除关键酶的活性,研究了不同浓度的铅(Pb)胁迫对玉米光系统Ⅰ(PSⅠ)、光系统Ⅱ(PSⅡ)的光化学活性和光合作用的影响,并分析了Pb胁迫下两个光系统的相互关系.结果表明:铅胁迫显著抑制了玉米地上部分和地下部分的生长、降低了叶片光合色素含量、并通过非气孔因素限制了光合作用、导致过剩激发能的增加;铅胁迫显著抑制了超氧化物歧化酶(SOD)和抗坏血酸过氧化物酶(APX)的活性、伤害了PSII反应中心、PSII的受体侧和供体侧(放氧复合体)以及PSI光化学活性.     

Manganese binding to sodium cyanide-treated chloroplasts: Effects of light and redox-potentials on the binding

Mn²⁺-binding to Mn-depleted chloroplasts by the treatment withcyanide was inhibited by light. Quantitative study indicatesthat only the binding with a high affinity constant was inhibitedby light. This photoinhibition required electron transport activityand disappeared on addition of 2,6-dichlorophenolindophenol.The binding of Mn²⁺ was also inhibited by addition of reductantin the dark. The high affinity binding at various redox potentialsindicates the participation of a two electrons-transfer componenthaving a midpoint potential of 336 mV. Mn²⁺-binding was decreasedon treatment of chloroplasts with proteolytic enzyme in thedark but not in the light. We propose that the high affinitybinding site is concealed in the membrane matrix of chloroplastsat low oxidation-reduction potential. (Received December 14, 1976; )     

Further characterization of a photosystem ii particle isolated from spinach chloroplasts by triton treatment delayed light emission

Delayed light emission from the Triton-fractionated Photosystem II subchloroplast fragments (TSF-IIa) was measured between 0.5 and 10 ms after the termination of illumination. The delayed light emission was diminished by Photosystem II inhibitors, DCMU and o-phenanthroline, which act between the reduced primary acceptor and the plastoquinone pool.Secondary electron donors to Photosystem II, diphenylcarbazide, phenylenediamine, Mn²⁺, and ascorbate inhibited delayed light emission. Secondary electron acceptors such as ferricyanide, dichlorophenol indophenol, and dimethyl benzoquinone enhanced delayed light emission. The addition of secondary electron acceptors to TSF-IIa particles containing Mn²⁺ restored delayed light emission to almost the control level. The plastoquinone antagonist, 2,5-dibromo-3-methyl-6-isopropyl p-benzoquinone, increased delayed light emission at low concentrations but decreased it at higher concentrations. Silicomolybdate enhanced the delayed light emission of TSF-IIa particles markedly, and reversed the inhibition by DCMU. Silicomolybdate showed a similar stimulatory effect on the delayed-light intensity in broken spinach chloroplasts at shorter times after the termination of illumination. Carbonyl cyanide m-chloro (or p-trifluoromethoxy) phenylhydrazones inhibited the delayed light emission, but NH₄Cl had no effect.     

Modifications of the functioning of Photosystem II induced in the dark by alkaline pH in the presence of cations

Submission of chloroplasts to alkaline pH, in the range pH 7.5–9.5, leads to changes in their oxygen-evolving capacities. These changes are enhanced by the addition of divalent cations and also monovalent cations at high concentrations. (1) Dark incubation of chloroplasts at pH ? 9 gives rise to a time-dependent inactivation of electron transport from water to 2,6-dichlorophenolindophenol measured at neutral pH. The rate of inactivation is increased by adding cations. (2) The variable fluorescence is decreased with a dependence on incubation time and concentration of cations similar to that of the Hill reaction. Addition of the electron donor NH₂OH removes most of the fluorescence quenching, (3) EPR measurements indicate that the inactivations are accompanied by loss of Mn²⁺ and the appearance of signal II fast. (4) At lower pH (7.5) the oscillations of oxygen evolved per flash during a sequence of flashes show an increase in damping when 20 mM MgCl₂ is present instead of 100 mM KCI. These changes are not seen at pH 6. (5) None of these Mg²⁺-induced modifications are prevented by glutaraldehyde fixation. We conclude that the effects of alkaline pH and MgCl₂ do not involve major protein structural changes, and that both act on the manganese-containing protein of the oxygen-evolving site.     

The release of polypeptides and manganese from oxygen-evolving photosystem II preparations following zinc-treatment

Inhibition of oxygen evolution in photosystem II membrane fragments from pea chloroplasts by washing with Zn₂₊ causes appearance of the EPR signal of Mn(H₂O)₆²⁺. This Mn²⁺ remains associated with the membrane fraction. Release of Mn²⁺ into the medium was correlated with the amount of the 23 kDa protein removed from the membrane. This suggests that this protein may function as a ‘gate’ to an aqueous compartment into which Mn²⁺ is released. Inhibition by Zn²⁺ correlated with the release of 1 Mn²⁺ per reaction centre, out of a total stoichiometry of 4 Mn atoms per reaction centre. By comparing the release of Mn following Zn-treatment of NaCI or CaC1₂ washed membranes, it is concluded that the 33 kDa protein is involved in binding of 2 Mn.     

Effect of Water Stress on Photochemical Activity of Chloroplasts during Greening of Etiolated Barley Seedlings

Water stress retards accumulation of chlorophyll and chlorophylla/b protein complex during greening of barley seedlings in light.The rate of 2,6-dichlorophenol indophenol (DCPIP) photoreductionin isolated chloroplast which decreases under water stress isenhanced significantly in the presence of electron donors, diphenylcarbazide (DPQ) and Mn²⁺. Under water stress, the decrease ofthe rate of oxygen evolution measured in continuous white lightwas 40–73% and that of oxygen uptake (as a measure ofelectron transport through PS I from reduced DCPIP) was onlyabout 20%. During greening, under water stress, (i) a differentialinhibition of PS II biosynthesis as compared to PS I occurs,(ii) the site of electron donation by DPC seems to be closerto the reaction center ofPS II, (iii) the oxidizing side ofPS II near the oxygen-evolving system is affected maximallyby water stress. (Received March 11, 1980; Accepted November 13, 1980)     

Uptake of ATP analogs by isolated pea chloroplasts and their effect on CO2 fixation and electron transport

1. The ATP analog, adenylyl-imidodiphosphate rapidly inhibited CO₂-dependent oxygen evolution by isolated pea chloroplasts. Both α, β- and β, γ-methylene adenosine triphosphate also inhibited oxygen evolution. The inhibition was relieved by ATP but only partially relieved by 3-phosphoglycerate. Oxygen evolution with 3-phosphoglycerate as substrate was inhibited by adenylyl-imidodiphosphate to a lesser extent than CO₂-dependent oxygen evolution. The concentration of adenylyl-imidodiphosphate required for 50% inhibition of CO₂-dependent oxygen evolution was 50 μM.2. Although non-cyclic photophosphorylation by broken chloroplasts was not significantly affected by adenylyl-imidodiphosphate, electron transport in the absence of ADP was inhibited by adenylyl-imidodiphosphate to the same extent as by ATP, suggesting binding of the ATP analog to the coupling factor of phosphorylation.3. The endogenous adenine nucleotides of a chloroplast suspension were labelled by incubation with [¹⁴C]ATP and subsequent washing. Addition of adenylyl-imidodiphosphate to the labelled chloroplasts resulted in a rapid efflux of adenine nucleotides suggesting that the ATP analog was transported into the chloroplasts via the adenine nucleotide translocator.4. It was concluded that uptake of ATP analogs in exchange for endogenous adenine nucleotides decreased the internal ATP concentration and thus inhibited CO₂ fixation. Oxygen evolution was inhibited to a lesser extent in spinach chloroplasts which apparently have lower rates of adenine nucleotide transport than pea chloroplasts.     

Effect of Mn2+ and Ca2+ on O2 evolution and on the variable fluorescence yield associated with Photosystem II in preparations of Anacystis nidulans

Extraction with EDTA of lyophilized and lysozyme treated preparations of the blue-green algae Anacystis nidulans resulted in loss of the capacity for photoevolution of O₂. Reactivation was achieved by the addition of both cations: Mn²⁺ and Ca²⁺ (or to a smaller extent by Mn²⁺ and Sr²⁺). The dual requirement for Mn²⁺ and Ca²⁺ could be demonstrated when the O₂ evolution under short saturating light flashes and the variable chlorophyll fluorescence associated with the reduction of the primary acceptor of Photosystem II was examined. The fluorescence experiments in addition showed that incorporation of the cations was a light dependent step, since the fluorescence rise only started after a lag period.     

Reversible inactivation of photosystem II reaction centers in cation-depleted chloroplast membranes

Isolated pea chloroplasts were washed once in 10 mm NaCl and were then suspended in “low-salt” medium. Approximately one-half of the photosystem II reaction centers of these salt-depleted membranes were found to be photochemically inactive. These units became active in the presence of low concentrations of divalent cations (5–10 mm Mg²⁺) or high concentrations of monovalent cations (150–200 mm Na⁺), as evidenced by a twofold increase in the steady-state flash yield of oxygen evolution under short (~10-μs) saturating repetitive flashes (two per second). The half-maximal increase in flash yield occurred at ~2 mM Mg²⁺ or ~75 mm Na⁺. The flash yield of hydroxylamine oxidation in these low-salt chloroplasts increased twofold after Mg²⁺ addition, indicating that the cation action was close to the reaction-center chlorophyll complex. The relation between flash yield and dark time between flashes was not changed significantly by Mg²⁺, indicating that the rate-limiting step of the overall electron transport (H₂0 —→ ferricyanide) was not affected significantly. When the rate-limiting step was bypassed using silicomolybdate as the photosystem II electron acceptor (in the presence of diuron), the reduction rate doubled in the presence of Mg²⁺, even under continuous, saturating light. In glutaraldehyde-fixed chloroplasts, Mg²⁺ did not increase the flash yield of O² evolution; this suggests that protein conformational changes in the chloroplast membranes were involved in Mg²⁺ activation of photosystem II centers.