Chloroplast translocations induced by light pulses

The effect of single blue-light pulses on chloroplast rearrangement was studied in the leaves of Tradescantia albiflora, Chlorophytum elatum, and Lemna trisulca. For measuring translocations in terrestrial plants the method of transmission changes was used; translocations in the water plant Lemna were studied by direct microscopic observation and counting. Strong light (30 W m^-2) applied in the form of short pulses, shorter than a lag period of translocations, induces some transient effects in the following dark period. With short pulses, transient rearrangements of chloroplasts to a weak-light position were found. With longer pulse duration, biphasic responses took place in Tradescantia and Lemna: The initial movement to a partial strong-light position was followed by a wave of translocation to a weak-light arrangement. In Chlorophytum this type of response appeared only in a narrow fluence range. The validity of the reciprocity law in relation to fluence rate and time of irradiation was confirmed for Tradescantia. The results may give us an insight into the kinetics of the primary effects of light in the translocation process.     

Pb-Induced Avoidance-Like Chloroplast Movements in Fronds of Lemna trisulca L.

Lead ions are particularly dangerous to the photosynthetic apparatus, but little is known about the effects of trace metals, including Pb, on regulation of chloroplast redistribution. In this study a new effect of lead on chloroplast distribution patterns and movements was demonstrated in mesophyll cells of a small-sized aquatic angiosperm Lemna trisulca L. (star duckweed). An analysis of confocal microscopy images of L. trisulca fronds treated with lead (15 μM Pb²⁺, 24 h) in darkness or in weak white light revealed an enhanced accumulation of chloroplasts in the profile position along the anticlinal cell walls, in comparison to untreated plants. The rearrangement of chloroplasts in their response to lead ions in darkness was similar to the avoidance response of chloroplasts in plants treated with strong white light. Transmission electron microscopy X-ray microanalysis showed that intracellular chloroplast arrangement was independent of the location of Pb deposits, suggesting that lead causes redistribution of chloroplasts, which looks like a light-induced avoidance response, but is not a real avoidance response to the metal. Furthermore, a similar redistribution of chloroplasts in L. trisulca cells in darkness was observed also under the influence of exogenously applied hydrogen peroxide (H₂O₂). In addition, we detected an enhanced accumulation of endogenous H₂O₂ after treatment of plants with lead. Interestingly, H₂O₂-specific scavenger catalase partly abolished the Pb-induced chloroplast response. These results suggest that H₂O₂ can be involved in the avoidance-like movement of chloroplasts induced by lead. Analysis of photometric measurements revealed also strong inhibition (but not complete) of blue-light-induced chloroplast movements by lead. This inhibition may result from disturbances in the actin cytoskeleton, as we observed fragmentation and disappearance of actin filaments around chloroplasts. Results of this study show that the mechanisms of the toxic effect of lead on chloroplasts can include disturbances in their movement and distribution pattern.     

Contractile elements of Lemna trisulca L. glycerinated cell models during chloroplast translocations.

Electron microscopy of Lemna glycerinated cell models depicts contractile elements during chloroplast translocations. One contractile element, the thin ectoplasmic layer, is < or = 0.4 microm thick, pressed against plasma membrane-cell wall. Thin ectoplasmic layer contains numerous oriented filaments and some appear to be actin and myosin. Another contractile element is the outer chloroplast membrane which envelops each chloroplast and joins or fuses with the thin ectoplasmic layer. Chloroplast interconnections are formed between two or more chloroplasts by outer chloroplast membranes; they enhance chloroplast communications, translocations, and molecular exchanges.     

Chloroplast movement in Mougeotia induced by blue light pulses

The profile-to-face chloroplast movement in the green alga Mougeotia has been induced by strong blue and near-ultraviolet light pulses (6 J m^-2). Simultaneously, strong red or far-red light (10 W m^-2) was applied perpendicularly to the inducing beam. The response was measured photometrically. Against the far-red background the reciprocity law was found to hold for pulse durations varying two orders of magnitude. The action spectrum exhibited a maximum near 450 nm and a distinct increase in near-ultraviolet. The time-course and the spectral dependence of pulse responses of chloroplasts in Mougeotia were similar to those recorded for other plants which are sensitive only to blue. This points to an alternative sensor system active in the short-wavelength region in addition to the phytochrome system.Abbreviations FR far-red light - Pr red absorbing form of phytochrome - Pfr far-red absorbing form of phytochrome - R red light This paper is dedicated to the memory of Professor Jan Zurzycki     

Influence of calcium on blue-light-induced chloroplast movement in Lemna trisulca L.

The presence of calcium is essential for chloroplast movement induced by blue light in Lemna trisulca L. The regulatory role of calcium was confirmed by the inhibition of chloroplast movement by cytochalasin B and trifluoperazine. The calcium concentration in tissues was modified by ethylene glycol-bis(2-aminoethylether)-N,N,N, N-tetraacetic acid (EGTA), the calcium ionophore A23187 and La³⁺. Only a long period of incubation (12h) in EGTA or La³⁺ caused distrubances in chloroplast movement. This indicates that calcium influx is not essential for chloroplast movement. Those conditions that dramatically changed the internal calcium concentration, either applications of calcium ionophore A23187 and EGTA, or ionophore and La³⁺, markedly decreased the amplitude of response to blue-light pulses. This demonstrates that disturbances of chloroplast movement are observable only when internal stores of calcium are affected by Ca²⁺-antagonists. We suggest that the calcium involved in blue-light-induced chloroplast movement is derived from intracellular stores. The addition of Mg²⁺ to EGTA buffer counteracted its effect, indicating that Mg²⁺, as well as Ca²⁺, might possibly be involved in chloroplast movement.Abbreviations EGTA ethylene glycol-bis(2-aminoethylether)-N,N,N,N-tetraacetic acid - Hepes 4(2-hydroxyethyl-1-piperazine) ethanesulfonic acid - A23187 calcium ionophore We express our gratitude to Professor W. Korohoda for valuable critical comments on this paper and stimulating discussion. We also thank Mr. P. Malec for help in preparing the experiment with trifluoperazine and Mr. A. Waloszek for taking the photographs. We are indebted to Mr. Tim Kline (International House, Krakow, Poland) for improving the English style. This research was supported by grant No. 1042/P2/92/03 from the State Committe for Scientific Research.     

Comparative utilization of inorganic and organic compounds as sole nitrogen sources by the submergent duckweed,Lemna trisulca L.

The capability of the submergentLemna trisulca L. to utilize various inorganic and organic sources of nitrogen was studied using both non-axenic and axenic cultures. When doubling time for frond production was measured, the nitrogen sources in order of effectiveness were urea, aspartic acid, nitrate, glutamic acid, arginine, ammonium and casein hydrolysate. Nitrite supported a relatively rapid growth rate after an initial lag of 7 days. Other parameters of growth such as fresh or dry weight per frond or ohlorophyll content did not oorrelate well with rate of frond production. Casein hydrolysate and urea were found to elicit a morphology different from that seen in cultures containing the other nitrogen sources. These preferences for source of nitrogen were different than those known for the emergent species ofLemnaceae. The unique value ofL. trisulca as a subject for plant physiological research is discussed. This study also provides a possible explanation for the existence of nutritional niches existing in aquatio ecosystems containing several different species ofLemnaceae.     

Control of Photosystem II in spinach leaves by continuous light and by light pulses given in the dark

The light-induced induction of components of non-photochemical quenching of chlorophyll fluorescence which are distinguished by different rates of dark relaxation (qNf, rapidly relaxing and qNs, slowly relaxing or not relaxing at all in the presence brief saturating light pulses which interrupt darkness at low frequencies) was studied in leaves of spinach.After dark adaptation of the leaves, a fast relaxing component developed in low light only after a lag phase. Quenching increased towards a maximum with increasing photon flux density. This fast component of quenching was identified as energy-dependent quenching qE. It required formation of an appreciable transthylakoid pH and was insignificant when darkened spinach leaves received 1 s pulses of light every 30 s even though zeaxanthin was formed from violaxanthin under these conditions.Another quenching component termed qNs developed in low light without a lag phase. It was not dependent on a transthylakoid pH gradient, decayed exponentially with a long half time of relaxation and was about 20% of total quenching irrespective of light intensity. When darkened leaves were flashed at frequencies higher than 0.004 Hz with 1 s light pulses, this quenching also appeared. Its extent was very considerable, and it did not require formation of zeaxanthin. Relaxation was accelerated by far-red light, and this acceleration was abolished by NaF.We suggest that qNs is the result of a so-called state transition, in which LHC II moves after its phosphorylation from fluorescent PS II to nonfluorescent PS I. This state transition was capable of decreasing in darkened leaves the potential maximum quantum efficiency of electron flow through Photosystem II by about 20%.Abbreviations PFD photon flux density - PS photosystem     

Retinal alterations induced by continuous light in immature rats

Summary Immature albino rats were subjected to (a) continuous illumination for 5–9 days, or (b) continuous illumination followed by prolonged darkness. Their electroretinographic responses and the ultrastructural characteristics of the rod outer segments, as revealed by a mixture of zinc iodine-osmium tetroxide (ZIO) at different temperatures, were studied and compared with those of a control group maintained in a cyclic rhythm of light and darkness.Noteworthy differences in the distribution of ZIO reactive sites were observed in the rats exposed for 5–9 days to continuous illumination (no electroretinographic responses) as compared with normal controls. At 4°C, ZIO staining was negative in the rods of illuminated rats whereas at 20 and 60°C it was positive inside the tubular and vesicular structures.After prolonged darkness, in rats with a partial electroretinographic recovery and ultrastructural restoration, the ZIO reaction showed a similar pattern to that observed in the control group, ZIO deposits being found both in the intra and extradiscal spaces.Supported by Grants from the Consejo Nacional de Investigaciones Cientificas y Técnicas, Argentina and the National Institutes of Health (51 NS 06953-09 NEUA), U.S.A., and by the Universidad Nacional de Buenos Aires and Fight for Sight Inc., N.Y., USA.We are indebted to Miss Margarita López for her skilful technical assistance and to Mr. Alberto Saenz for the electron micrographs.     

Retinal alterations induced by continuous light in immature rats

Immature albino rats were exposed to continuous illumination for 5-93 days and the light induced ultrastructural and electroretinographic changes were studied. Another group was exposed to continuous light for 7-9 days and then kept in complete darkness, or in cyclic light-dark up to 90 days. By comparison with the results obtained in adult animals, lesions appeared faster in the immature group. Tubular transformation of rods, phagocytosis of altered outer and inner segments with resulting changes in retinal organization, synaptic degeneration in the outer plexiform layer, and cell lysis of some photoreceptor cell perikarya are described. ERG recovery, following the period of darkness or cyclic light-dark was only partial, the amplitude of the "b" wave reached only 50-60% of the control preillumination values. However, the fine structure of the recovered outer segments was similar to that found in normal retinae.     

Photoinhibition of photosynthesis in Lemna gibba as induced by the interaction between light and temperature. I. Photosynthesis in vivo

The floating angiosperm Lemna gibba L. was exposed for 2 h to various combinations of photosynthetic photon flux densities and temperature. The extent of photoinhibition of photosynthesis was assayed by measuring the net CO₂ uptake before and after a photoinhibitory treatment, and the time course for photoinhibition was studied. It was found that the maximum quantum yield and the light-saturated rate of CO₂ uptake were affected by the interaction between light and temperature during the photoinhibitory treatment. At a constant photon flux density of 650 μmol m⁻² s⁻¹ the extent of photoinhibition increased with decreasing temperature showing that even a chilling-resistant plant like L. gibba is much more susceptible to photoinhibition at chilling temperatures. About 60% photoinhibition of the quantum yield for CO₂ uptake could be obtained either by a high photon flux density of 1 750 μmol m⁻² s⁻¹ and 25°C or by a moderate photon flux density of 650 μmol m⁻² s⁻¹ and 3°C. The time courses of recovery from 60% photoinhibition produced by either of these two treatments were similar, indicating that the nature of the photoinhibition was intrinsically similar. The extent of photoinhibition was related to the amount of light absorbed in excess to what could be handled by photosynthesis at that temperature. The vital importance of photosynthesis in alleviating photoinhibition is discussed.     

Growth dynamics of a methylotroph (Methylomonas L3) in continuous cultures. I. Fast transients induced by methanol pulses and methanol accumulation

The dynamic behavior of the Ribulose Monophosphate-type Methylomonas L3 in continuous cultures was studied, using methanol pulses to induce fast transients in steady-state cultures of single (methanol) and mixed (methanol plus formaldehyde) substrates. In several experiments, the methanol-uptake rate (MUR) profiles displayed negative MUR values for a time period following the methanol pulse, and significant amounts of methanol disappeared immediately following the pulse. These phenomena suggested the accumulation of methanol in the cells upon pulsing, apparently due to an active transport system. Accordingly, and in order to estimate the potential of the transport system for methanol accumulation, accumulation profiles were calculated for several pulse experiments. The calculations are based on a methanol balance and experimentally determined values of the cell volume and the true transient biomass yields. It is calculated that methanol accumulates up to 200-fold to very high intracellular concentrations. The accumulation is calculated to be much higher in single- (methanol) substrate cultures of low dilution rate than in cultures of high dilution rate or of mixed substrates. The specific growth rate immediately following the methanol pulse decreased in single-substrate cultures and increased in mixed-substrate ones. The biomass yield decreased after the methanol addition in all experiments; however, the drop was less severe in the mixed-substrate experiments. It is also suggested that formaldehyde as a methanol cosubstrate may be an effective means of providing more stable biomass yields and growth rates in reactors with imperfect mixing, and of protecting the reactor against accidentally induced methanol accumulation.     

Amaranthin accumulation in continuous red and blue light by seedlings ofAmaranthus caudatus L.

Four days oldAmaranthus seedlings responded to light treatment with an increase of amaranthin accumulation. With increasing irradiation time, red light caused a saturation effect. Blue light induced a high irradiation response. The blue light effect was reversible to a certain extent by far-red irradiation given at the end of the treatment with blue light. Intermittent red light (3 h red light, 3 h dark, …) caused a higher amaranthin accumulation than 24 h continuous red light. Results obtained with red and blue light are discussed on the basis of the phytochrome system.     

Photoinhibition of photosynthesis in Lemna gibba as induced by the interaction between light and temperature. II. Photosynthetic electron transport

Photoinhibition of photosynthesis in Lemna gibba L. was induced by exposing intact plants to a high photosynthetic photon flux density of 1 750 μmol m⁻² s⁻¹ at a low temperature of 3°C. Subsequently isolated chloroplasts showed pronounced reductions in the capacity of whole chain electron transport, measured as Hill activity, and in the efficiency of electron transport to the primary electron acceptor Q of photosystem II, measured as variable chlorophyll fluorescence at 20°C. These changes proceeded with similar kinetics (probably of the first-order reaction), suggesting that the site of photoinhibition is in the electron transfer to Q. A partial uncoupling of the whole chain electron transport also occured. The capacity of electron transport mediated by photosystem I was unaffected. The extent of photoinhibition of photosynthetic electron transport, as produced by a 2 h exposure of L. gibba to three different combinations of photon flux density and temperature was studied. It was shown that intrinsically similar states of photoinhibition, on the evidence of their time courses of recovery, were induced by either a high photon flux density and 25°C or by a moderate photon flux density and 3°C.     

Chloroplast structure in normal and pigment-deficient soybeans grown in continuous red or far-red light

Clark L1, a normal green soybean [ Glycine max (L.) Merrill] and Clark y₉y₉, a backross-developed isoline exhibiting pigment deficiency, were grown under continuous red (11 W m⁻² and far-red (9 W m⁻²) light. Chloroplast thylakoids from the unifoliolate leaf (9–10 days old) were isolated and analyzed for pigments, pigment-protein, membrane polypeptides, electron transport and ultrastructural differences. Chloroplasts of soybean plants grown under far-red light have decreased chlorophyll a to chlorophyll b ratio, increased light-harvesting complexes, and grana structure with few stroma-type thylakoids. Photosystem II/photosystem I ratios (PSII/PSI) are higher in far-red due to decreased synthesis of PSI reaction center and/or less antenna associated with PSI.     

Rapidly reversible chlorophyll fluorescence quenching induced by pulses of supersaturating light in vivo

The saturation pulse method provides a means to distinguish between photochemical and non-photochemical quenching, based on the assumption that the former is suppressed by a saturating pulse of light (SP) and that the latter is not affected by the SP. Various types of non-photochemical quenching have been distinguished by their rates of dark relaxation in the time ranges of seconds, minutes, and hours. Here we report on a special type of non-photochemical quenching, which is rapidly induced by a pulse of high-intensity light, when PS II reaction centers are closed, and rapidly relaxes again after the pulse. This high-intensity quenching, HIQ, can be quantified by pulse-amplitude-modulation (PAM) fluorimetry (MULTI-COLOR-PAM, high sensitivity combined with high time resolution) via the quasi-instantaneous post-pulse fluorescence increase that precedes recovery of photochemical quenching in the 100–400-µs range. The HIQ amplitude increases linearly with the effective rate of quantum absorption by photosystem II, reaching about 8% of maximal fluorescence yield. It is not affected by DCMU, is stimulated by anoxic conditions, and is suppressed by energy-dependent non-photochemical quenching (NPQ). The HIQ amplitude is close to proportional to the square of maximal fluorescence yield, Fm′, induced by an SP and varied by NPQ. These properties are in line with the working hypothesis of HIQ being caused by the annihilation of singlet excited chlorophyll a by triplet excited carotenoid. Significant underestimation of maximal fluorescence yield and photosystem II quantum yield in dark-acclimated samples can be avoided by use of moderate SP intensities. In physiologically healthy illuminated samples, NPQ prevents significant lowering of effective photosystem II quantum yield by HIQ, if excessive SP intensities are avoided.

     

Growth stimulation inPhaseolus vulgaris L. induced by gamma irradiation of seeds

Gamma radiation in doses 0.13 to 0.77 C kg^-1 (0.5 to 3.0 kR) significantly (P ≥ 0.01) stimulated seed germination, seedling height, and length of primary leaves of French bean cv. ‘Blue Lake’; these doses did not affect chlorophyll content per leaf area unit. Doses of 1.16 to 1.93 C kg^-1 (4.5 to 7.5 kR) induced inhibition of the four parameters studied.     

Photoinhibition of photosynthesis in Lemna gibba as induced by the interaction between light and temperature. III. Chlorophyll fluorescence at 77 K

Photoinhibition in Lemna gibba L. was studied by interpreting chlorophyll fluorescence characteristics at 77 K on the basis of the bipartite model of Butler and co-workers (Butler 1978). Application of this analysis to chloroplasts (isolated from plants before and after exposure to a photosynthetic photon flux density of 1 750 μmol m⁻² s⁻¹ at 3°C for 2 h) revealed that photoinhibition had the following effect on primary events in photosynthesis. Firstly, the fluorescence of PS II increased (44%) in the state of open traps (F_o) and decreased (32%) in the state of closed traps (F_m). It is suggested, that the F_o-decrease reflects increased quenching by radiationless decay, both effects occurring at PS II reaction centers. Secondly, the rate constant for transfer of excitation energy from PS II to PS I (k_T(μ→J)) increased by 34%. However, in the state of closed traps, the flux of excitation energy via this transfer process decreased, most likely because of increased quenching by radiationless decay at PS II reaction centers. Thirdly, the probability for fluorescence from PS I decreased (19%). This indicates increased quenching by radiationless decay.     

Efficiency of photosynthesis in continuous and pulsed light emitting diode irradiation

The light utilization efficiency and relative photon requirement of photosynthesis in pulsed and continuous light from light emitting diodes (LEDs) has been measured. First, we chacterized the photon requirement of photosynthesis from light of LEDs that differ in spectral quality. A photon requirement of 10.3±0.4 was measured using light from a 658 nm peak wavelength (22 nm half band width) LED over the range of 0–50 mol photons m^–2 s^–1 in 2 kPa O₂ in leaves of tomato (Lycopersicon esculentum Mill., cv. VF36). Because the conversion of electrical power to photons increased with wavelength, LED lamps with peak photon output of 668 nm were most efficient for converting electricity to photosynthetically fixed carbon. The effect of pulsed irradiation on photosynthesis was then measured. When all of the light to make the equivalent of 50 mol photons m^–2 s^–1 was provided during 1.5 s pulses of 5000 mol photons m^–2 s^–1 followed by 148.5 s dark periods, photosynthesis was the same as in continuous 50 mol photons m^–2 s^–1. When the pulse light and dark periods were lengthened to 200 s and 19.8 ms, respectively, photosynthesis was reduced, although the averaged photon flux density was unchanged. Under these conditions, the light pulses delivered 10¹⁷ photons m^–2, which we calculate to be equivalent to the capacitance of PS I or PS II. Data support the theory that photons in pulses of 100 s or shorter are absorbed and stored in the reaction centers to be used in electron transport during the dark period. When light/dark pulses were lengthened to 2 ms light and 198 ms dark, net photosynthesis was reduced to half of that measured in continuous light. Pigments of the xanthophyll cycle were not affected by any of these pulsed light treatments even though zeaxanthin formation occurred when leaves were forced to dissipate an equal amount of continuous light.Abbreviations CWF cool white fluorescent - EPS xanthophyll epoxidation state - LED light emitting diode - LUE light utilization efficiency - PFD photon flux density - PR photon requirement (for CO₂ fixation) - PS II primary donor in Photosystem II - RPR relative photon requirement