Chlorophyll a fluorescence induction: a personal perspective of the thermal phase, the J–I–P rise

The fast (up to 1?s) chlorophyll (Chl) a fluorescence induction (FI) curve, measured under saturating continuous light, has a photochemical phase, the O-J rise, related mainly to the reduction of Q(A), the primary electron acceptor plastoquinone of Photosystem II (PSII); here, the fluorescence rise depends strongly on the number of photons absorbed. This is followed by a thermal phase, the J-I-P rise, which disappears at subfreezing temperatures. According to the mainstream interpretation of the fast FI, the variable fluorescence originates from PSII antenna, and the oxidized Q(A) is the most important quencher influencing the O-J-I-P curve. As the reaction centers of PSII are gradually closed by the photochemical reduction of Q(A), Chl fluorescence, F, rises from the O level (the minimal level) to the P level (the peak); yet, the relationship between F and [Q(A) (-)] is not linear, due to the presence of other quenchers and modifiers. Several alternative theories have been proposed, which give different interpretations of the O-J-I-P transient. The main idea in these alternative theories is that in saturating light, Q(A) is almost completely reduced already at the end of the photochemical phase O-J, but the fluorescence yield is lower than its maximum value due to the presence of either a second quencher besides Q(A), or there is an another process quenching the fluorescence; in the second quencher hypothesis, this quencher is consumed (or the process of quenching the fluorescence is reversed) during the thermal phase J-I-P. In this review, we discuss these theories. Based on our critical examination, that includes pros and cons of each theory, as well mathematical modeling, we conclude that the mainstream interpretation of the O-J-I-P transient is the most credible one, as none of the alternative ideas provide adequate explanation or experimental proof for the almost complete reduction of Q(A) at the end of the O-J phase, and for the origin of the fluorescence rise during the thermal phase. However, we suggest that some of the factors influencing the fluorescence yield that have been proposed in these newer theories, as e.g., the membrane potential ΔΨ, as suggested by Vredenberg and his associates, can potentially contribute to modulate the O-J-I-P transient in parallel with the reduction of Q(A), through changes at the PSII antenna and/or at the reaction center, or, possibly, through the control of the oxidation-reduction of the PQ-pool, including proton transfer into the lumen, as suggested by Rubin and his associates. We present in this review our personal perspective mainly on our understanding of the thermal phase, the J-I-P rise during Chl a FI in plants and algae.     

Measurement of the chlorophyll fluorescence induction kinetics with a 10 µs time resolution and its application in the forest decline research

Summary Measurement methods are described which determine the initial phase of the fluorescence induction kinetics with a maximum time resolution of 10 µs simultaneously for the two fluorescence componentsF ₆₈₅(t) andF ₁₃₀(t) selected by filters at the wavelengths 685 nm and 730 nm, respectively. The excitation light provided by a He-Ne laser (632.8 nm) is switched on within 0.3 µs (maximum intensityI _e=12 mW/cm²).F _o,F _p, andF _s, the initial-, peak-, and steady-state intensity and the initial valueR _o of the ratioR(t)=F ₇₃₀(t)/F ₆₈₅(t) can accurately be determined as well as the initial time derivativeF _o ^* of the fluorescence intensity.F _o andF _o ^* are related to the quantum yield _a of the antenna and to the photochemical quantum yield _pc, respectively. Spruce, oak, birch, poplar, and soy bean show a decline ofR(t) fromR _o to a first minimumR _b at some 10 ms which has a similar value as the second minimumR _p in the time range of seconds. Furthermore, the initial valueR _o and the steady-state valueR _S ofR(t) are also very similar. Measurements on spruce with water deficiency and with varying excitation light intensityI _e show effects on the initial phase of the fluorescence induction kinetics. Further measurements on spruce of different damage classes indicate that for the current year's needles the ratioF _p/F_o, is the most sensitive parameter to differentiate between the damage classes and thatF _o/F_s andR _o/R_b are also affected. As demonstrated by measurements on leaves of soy beans, the initial decrease ofR(t) fromR _o toR _b originates from a change of the fluorescence spectrum because no change of the leaf transmission can be observed in the time range between 10 µs and 1 ms.     

N,N,N′,N′-tetramethyl-p-phenylenediamine initiates the appearance of a well-resolved I peak in the kinetics of chlorophyll fluorescence rise in isolated thylakoids

Addition of N,N,N′,N′-tetramethyl-p-phenylendiamine (TMPD) to thylakoid membranes isolated from pea leaves initiates the appearance of peak I in the polyphasic rise of chlorophyll (Chl) fluorescence observed during strong illumination, making it similar to that observed in leaves or intact chloroplasts. This effect depends on TMPD concentration and incubation period of isolated thylakoids with TMPD. The resolution of I-peak in the presence of weak concentrations of TMPD which reduced the overlap between I- and P-peaks, resulted from a decreased reduction of both fast and slow plastoquinone (PQ) pools of the granal and stromal thylakoids, respectively, as TMPD effectively accepts electrons from reduced PQ. High concentrations of TMPD markedly decreased the J–I–P phase of fluorescence rise and greatly retarded the I–P step rise. Accumulation of oxidized TMPD in the thylakoid lumen accelerated the re-oxidation of the acceptor side of Photosystem II (PSII) as illustrated by a two-fold increase in the magnitude of the fast component and complete suppression of the middle component of the variable Chl fluorescence (F_v) decay in the dark. Evidently, exogenous addition of high concentrations of TMPD prevented the light-induced reduction of the slow PQ pool.     

The fluorescence yield of the trimeric fucoxanthin–chlorophyll–protein FCPa in the diatom Cyclotella meneghiniana is dependent on the amount of bound diatoxanthin

The fluorescence yield of isolated fucoxanthin chlorophyll proteins, serving as light harvesting proteins in diatoms, was compared to the amount of diatoxanthin bound. Diatoxanthin was earlier shown to be involved in the xanthophyll cycle in diatoms as a functional analogue of zeaxanthin in higher plants. By growing cells under different light conditions, the amount of diatoxanthin in both the trimeric FCPa as well as the oligomeric FCPb of the diatom Cyclotella meneghiniana was increased. In the trimeric FCPa, the fluorescence yield decreased with increasing diatoxanthin content, whereas in the oligomeric FCPb fluorescence was generally lower, albeit constant. No pH dependence of fluorescence yield could be demonstrated except for artificially aggregated FCPa. Thus, diatoxanthin is able to quench fluorescence in FCPa, but the yield is also influenced by pH when the protein becomes aggregated.     

Analysis of chlorophyll fluorescence induction kinetics exhibited by DCMU-inhibited thylakoids and the origin of α and β centres

Analyses of room-temperature chlorophyll fluorescence curves from DCMU-inhibited thylakoids were used to investigate the proposed PS II structural heterogeneity of α and β centres. The kinetics of the area growth curves, representative of Q_A photoreduction, could be modified in the presence of DCMU by exogenous electron acceptors and by added reductants of the PQ pool. The effect of altered DCMU levels (range 0.2–100 μM) on the induction curve kinetics was to modify preferentially the slow-β component, while having only a very small effect on the total variable fluorescence yield. Over the DCMU concentration range used, the unnormalized area of the induction curve (A_max) decreased with increasing herbicide concentration by approx. 45%, indicating that less quanta were required to reduce Q_A. It was found that the dark reoxidation of Q_A in the presence of DCMU and Ant 2p after a light pretreatment regenerated the slow kinetic component. When chlorophyll fluorescence emission at 685 and 731 nm was measured, no difference was observed in the kinetics of the induction curve. The analysis of PS II-enriched, oxygen-evolving membranes indicated the presence of both the fast and slow kinetic components, although this type of preparation showed a modified fast phase. The above observations led to the conclusion that several of the previously proposed characteristics of PS II_α and PS II_β centres do not hold and that a type of PS II heterogeneity involving different degrees of DCMU inhibition is sufficient to explain many of the observations made.     

The rise in chlorophyll a fluorescence yield and decay in delayed light emission in Tris-washed chloroplasts in the 6–100 μs time range after an excitation flash

Parallel measurements of the rise in chlorophyll a fluorescence yield and delayed light emission decay, after a 10 ns saturating excitation flash, have been made in tris(hydroxymethyl)aminomethane-washed chloroplasts. Various electron donor systems (Mn²⁺; ascorbate; reduced phenylenediamine and benzidine) were used in conjuction with different preillumination regimes to alter [P⁺-680], the oxidized form of the Photosystem II reaction center chlorophyll a. Conditions giving rise to high [P⁺-680] resulted in only a small rise in fluorescence yield, an inhibition of a 6 μs delayed light component, and an enhancement of a 60 μs component of delayed light emission. These results confirm the hypothesis that P⁺-680 acts as a quencher of fluorescence and that delayed light emission in the microsecond time range is due to the back reaction of P⁺-680 and Q^?. (Q is the first “stable” electron acceptor of Photosystem II.) Two preillumination flashes are required before the full effect of Tris washing is observed in the delayed light emission decay and fluorescence yield rise; this suggests that a capacity to hold two charges exists between the Tris block and P⁺-680. Tris washing has no direct effect on the movement of electrons from Z (the first electron donor to P⁺-680) to P⁺-680. Finally, Mn²⁺ donates electrons to P⁺-680 via Z.     

The efficiency of electron transfer from QA − to the donor side of Photosystem II decreases during induction of photosynthesis: Evidences from chlorophyll fluorescence and photoacoustic techniques

The amplitudes ratio of the fast and slow phases (A_fast/A_slow) in the kinetics of the dark relaxation of variable chlorophyll fluorescence (F_V) was studied after various periods of illumination of dark-adapted primary barley leaves. Simultaneously, photosynthetic activity was monitored using the photoacoustic technique and the photochemical and non-photochemical fluorescence quenching parameters. The ratio A_fast/A_slow changed with the preceding illumination time in a two-step manner. During the first stage of photosynthetic induction (0–20 s of illumination), characterized by a drop in O₂-dependent photoacoustic signal following an initial spike and by a relatively stable small value of photochemical F_V quenching, the ratio A_fast/A_slow remained practically unaltered. During the second stage (20–60 s of illumination), when both the rate of O₂ evolution and the photochemical F_V quenching were found to be sharply developed, a marked increase in the above ratio was also observed. A linear correlation was found between the value of the photochemical quenching and the ratio A_fast/A_slow during the second phase of photosynthetic induction. It is concluded that the slow phase appearing in the kinetics of F_V dark relaxation is not due to the existence of Photosystem II reaction centres lacking the ability to reduce P700⁺ with high rates, but is instead related to the limitation of electron release from Photosystem I during the initial stage of the induction period of photosynthesis. This limitation keeps the intersystem electron carriers in the reduced state and thus increases the probability of back electron transfer from Q_A ^– to the donor side of Photosystem II.Abbreviations A_fast/A_slow the ratio of magnitudes between the fast and slow phases of dark relaxation of variable fluorescence - F_O initial level of chlorophyll fluorescence - F_V variable chlorophyll fluorescence (F-F_O) - (F_V)_S the yield of variable chlorophyll fluorescence under saturating pulse in illuminated leaves - (F_V)_M the yield of variable chlorophyll fluorescence under saturating pulse in dark-adapted leaves - PA photoacoustic - PSI Photosystem I - PS II Photosystem II - qN non-photochemical quenching - qQ photochemical quenching     

The Origin of Life: Chemical Evolution of a Metabolic System in a Mineral Honeycomb?

For the RNA-world hypothesis to be ecologically feasible, selection mechanisms acting on replicator communities need to be invoked and the corresponding scenarios of molecular evolution specified. Complementing our previous models of chemical evolution on mineral surfaces, in which selection was the consequence of the limited mobility of macromolecules attached to the surface, here we offer an alternative realization of prebiotic group-level selection: the physical encapsulation of local replicator communities into the pores of the mineral substrate. Based on cellular automaton simulations we argue that the effect of group selection in a mineral honeycomb could have been efficient enough to keep prebiotic ribozymes of different specificities and replication rates coexistent, and their metabolic cooperation protected from extensive molecular parasitism. We suggest that mutants of the mild parasites persistent in the metabolic system can acquire useful functions such as replicase activity or the production of membrane components, thus opening the way for the evolution of the first autonomous protocells on Earth.     

Effects of enhanced UV-B radiation in the field on the concentration of phenolics and chlorophyll fluorescence in two boreal and arctic–alpine lichens

Lichens constitute a prominent part of the vegetation at high latitudes and altitudes, but the effects of UV-B radiation on these symbiotic organisms are not well known. In a northern boreal site (Abisko, northern Sweden), the usnic acid-producing lichens Flavocetraria nivalis and Nephroma arcticum were exposed to enhanced UV-B radiation, corresponding to 25% ozone depletion, for two and one growing seasons, respectively. They were compared with lichens grown under ambient UV-B and harvested fresh from the field. The treated thalli of F. nivalis had been transplanted from a site 24 km from the treatment site. From this source locality, untreated thalli were also harvested. Enhanced UV-B did not affect concentrations of usnic acid and the two depsides phenarctin and nephroarctin. A gradual decline of usnic acid, probably coupled to unusually long periods of dry, sunny weather, was observed both under enhanced and ambient UV-B and in untreated thalli. Photosystem II efficiency in both species was slightly reduced by enhanced UV-B. However, differences between seasons were larger than differences between treatments, which indicate that UV-B effects are minor in comparison to other climatic variables. Concentrations of UV-B-absorbing phenolics in lichens do not show a simple relationship to UV-B dose and therefore cannot be used as bioindicators of UV-B levels.     

Characterization of the peridinin–chlorophyll a-protein complex in the dinoflagellate Symbiodinium

The water-soluble peridinin–chlorophyll a-proteins (PCPs) are one of the major light harvesting complexes in photosynthetic dinoflagellates. PCP contains the carotenoid peridinin as its primary pigment. In this study, we identified and characterized the PCP protein and the PCP gene organization in Symbiodinium sp. CS-156. The protein molecular mass is 32.7 kDa, revealing that the PCP is of the monomeric form. The intronless PCP genes are organized in tandem arrays. The PCP gene cassette is composed of 1095-bp coding regions and spacers in between. Despite the heterogeneity of PCP gene tandem repeats, we identified a single form of PCP, the sequence of which exactly matches the deduced sequence of PCP gene clone 7 (JQ395030) by LC–MS/MS analysis of tryptic digested PCP, revealing the mature PCP apoprotein is 312 amino acids in length. Pigment analysis showed a peridinin-to-Chl a ratio of 4. The peridinin-to-Chl a Q_y energy transfer efficiency is 95% in this complex.     

Effects of high temperature on the chlorophyll a fluorescence of Alhagi sparsifolia at the southern Taklamakan Desert

Climate change is expected to result in an increase in the frequency and magnitude of extreme weather events. Alhagi sparsifolia is an important factor for wind prevention and sand fixation in the forelands of the Taklamakan Desert. The effects of high temperature on desert plants remain widely unknown. In this work, chlorophyll a fluorescence induction kinetics were investigated at different time stresses of 5, 20, 40, and 60 min at temperature gradients of 38–44 °C at 2 °C intervals. A pronounced K-step was found, and the values of the maximum quantum yield for primary photochemistry, the quantum yield of electron transport, the density of reaction centers and the performance index on absorption basis were lowest after 60 min at 44 °C, thus indicating that the oxygen-evolving complex was damaged, the inactivated reaction centers increased, and the activity of the photosystem II (PSII) reaction center in leaves was seriously limited. Therefore, we suggest that under normal temperature (below 42 °C), the PSII of A. sparsifolia would be unaffected. When such temperature is maintained for 40 min, the activity of PSII would be limited, and when retained for 60 min, PSII may be severely damaged.     

The rise and fall of handicap principle: a commentary on the “Modelling and the fall and rise of the handicap principle”

The honesty of animal communication is in the spot lights in the last 30 years. During most of this time the field was dominated by one explanation: Zahavi’s handicap principle (Zahavi, J Theor Biol 67:603–605, 1975; Grafen, J Theor Biol 144:517–546, 1990). Grose (Biol Philos 2011) embarks to explain both the success of the theory and the empirical difficulties that exist despite this success. While I wholeheartedly agree with the criticism offered by Grose and with almost all the claims he makes, the treatment of the issue is far from complete and it still leaves much to be explained. Accordingly, my commentary consist of six sections: in the first section I clear up some technical issues left unexposed, most importantly the role of strategic cost in handicap signalling; in the second section I relate this to empirical testing; in the next section I comment on the historical development of the handicap principle; in the fourth section I review the biological models that came up with conclusions that contradict the handicap principle; in the fifth section I discuss the reasons behind the success of the handicap theory; finally, in the last section I discuss the application of the handicap theory to anthropology and human sciences.     

An in silico biomechanical analysis of the stent–esophagus interaction

Despite all technological innovations in esophageal stent design over the past 20 years, the association between the stent design’s mechanical behavior and its effect on the clinical outcome has not yet been thoroughly explored. A parametric numerical model of a commercially available esophageal bioresorbable polymeric braided wire stent is set up, accounting for stent design aspects such as braiding angle, strut material, wire thickness, degradation and friction between the wires comprising a predictive tool on the device’s mechanical behavior. Combining this tool with complex multilayered numerical models of the pathological in vivo stressed, actively contracting and buckling esophagus could provide clinicians and engineers with a patient-specific window into the mechanical aspects of stent-based esophageal intervention. This study integrates device and soft tissue mechanics in one computational framework to potentially aid in the understanding of the occurrence of specific symptoms and complications after stent placement.     

Erratum to: An in silico biomechanical analysis of the stent–esophagus interaction

Biomechanics and Modeling in Mechanobiology - In the original publication of the article, Tables 2 and 3 were published with error. The correct tables are provided below (Tables 2, 3). The...     

Leaf characteristics and diurnal variation of chlorophyll fluorescence in leaves of the ‘bana’ vegetation of the amazon region

In six dominant species of the Amazonian ‘Bana’ vegetation, leaf blade characteristics, pigment composition, and chlorophyll (Chl) fluorescence parameters were measured in young and mature leaves under field conditions. Leaf δ¹³C was comparable in the six species, which suggested that both expanding and expanded leaves contained organic matter fixed under similar intercellular and ambient CO₂ concentration (C _i/C _a). High leaf C/N and negative δ¹⁵N values found in this habitat were consistent with the extreme soil N-deficiency. Analysis of Chl and carotenoids showed that expanding leaves had an incomplete development of photosynthetic antenna when compared to adult leaves. Dynamic inactivation of photosystem 2 (PS2) at midday was observed at both leaf ages as F_v/F_m decreased compared to predawn values. Adult leaves reached overnight F_v/F_m ratios typical of healthy leaves. Overnight recovery of F_v/F_m in expanding leaves was incomplete. F₀ remained unchanged from midday to predawn and F_v tended to increase from midday to predawn. The recovery from midday depression observed in adult leaves suggested an acclimatory down-regulation associated with photo-protection and non-damage of PS2.     

The enigma of the rise of angiosperms: can we untie the knot?

Multiple hypotheses have been put forward to explain the rise of angiosperms to ecological dominance following the Cretaceous. A unified scheme incorporating all these theories appears to be an inextricable knot of relationships, processes and plant traits. Here, we revisit these hypotheses, categorising them within frameworks based on plant carbon economy, resistance to climatic stresses, nutrient economy, biotic interactions and diversification. We maintain that the enigma remains unresolved partly because our current state of knowledge is a result of the fragmentary nature of palaeodata. This lack of palaeodata limits our ability to draw firm conclusions. Nonetheless, based on consistent results, some inferences may be drawn. Our results indicate that a complex multidriver hypothesis may be more suitable than any single‐driver theory. We contend that plant carbon economy and diversification may have played an important role during the early stages of gymnosperms replacement by angiosperms in fertile tropical sites. Plant tolerance to climatic stresses, plant nutrition, biotic interactions and diversification may have played a role in later stages of angiosperm expansion within temperate and harsh environments. The angiosperm knot remains partly tied, but to unravel it entirely will only be feasible if new discoveries are made by scientific communities.     

Changes with age in the absorption spectrum of chlorophyll α in a diatom

Summary Absorption spectra of a young and an old culture of the diatom Pheodactylum tricornutum were measured in thin layers between two opal glass sheets. The spectra at 24° and at -196°C were replotted to give equal areas from 730–625 m to allow direct comparison. At 24°C the spectrum for the difference between the two cultures had a negative component of 18 m half width centered at 675 m and a positive region of W_0.5=26 m near 700 m.The spectra at -196°C may be somewhat distorted by clumping of the cells during freezing but nevertheless the 16 day culture clearly showed a smaller proportion of Ca 670 to Ca 680. This older culture has a shoulder due to a 707 m component. The difference curve at -196°C shows the decrease of an unsymmetrical band peaking at 669 m and an increase at 695 m in addition to the 707 m component. Due to the possibility of distortion, the presence of an actual component at 695 is doubtful in these particular cultures.The room temperature spectrum in the chloropyhll a region for the 5 day culture can be closely fitted by a single probability curve at 675 m having a half-width of 31 m. The sum of two components, with widths more reasonable for chlorophylls, also matched the data well enough. These two probability curves, of 22 m half width, centered on 669 and 683.2 m and had a height ratio, h₆₆₉/h₆₈₃ of 1.18. In the 16 day culture the ratio for these bands changed to 1.11 and there was extra absorption around 700 m.Dedicated to Professor C. B. van Niel on the occasion of his 70th birthday