Phosphoglucomutase isozymes of red cells in Mus musculus: Additional polymorphism at PGM-1 compared by two electrophoretic systems

Phosphoglucomutase (PGM) of red cells was examined in 15 inbred strains of mice, using two different starch gel electrophoretic buffer systems. Two new alleles, Pgm-1 ^c and Pgm-1 ^d, were discovered at the Pgm-1 locus. Pgm-1 ^c was first identified in strain C3H/HeNWe and Pgm-1 ^d in 129/ReWl. No variation was observed at the Pgm-2 locus.Supported in part by USPHS Pre-doctoral Fellowship No. 5 F1 GM-32,680, USPHS Research Resources Grant No. 5-PO6 RR 00343-05, USPHS (National Cancer Institute) Chemotherapy Contract 71-2010, and Biomedical Sciences Support Grant FR 07037 to the University of Kansas.     

Measurement of gradients of absorbed light in spinach leaves from chlorophyll fluorescence profiles

Profiles of chlorophyll fluorescence were measured in spinach leaves irradiated with monochromatic light. The characteristics of the profiles within the mesophyll were determined by the optical properties of the leaf tissue and the spectral quality of the actinic light. When leaves were infiltrated with 10^?4M DCMU [3‐(3,4‐dichlorophenyl)‐1, 1‐dimethyl‐urea] or water, treatments that minimized light scattering, irradiation with 2000 μmol m^?2 s^?1 green light produced broad Gaussian‐shaped fluorescence profiles that spanned most of the mesophyll. Profiles for chlorophyll fluorescence in the red (680 ± 16 nm) and far red (λ > 710 nm) were similar except that there was elevated red fluorescence near the adaxial leaf surface relative to far red fluorescence. Fluorescence profiles were narrower in non‐infiltrated leaf samples where light scattering increased the light gradient. The fluorescence profile was broader when the leaf was irradiated on its adaxial versus abaxial surface due to the contrasting optical properties of the palisade and spongy mesophyll. Irradiation with blue, red and green monochromatic light produced profiles that peaked 50, 100 and 150 μm, respectively, beneath the irradiated surface. These results are consistent with previous measurements of the light gradient in spinach and they agree qualitatively with measurements of carbon fixation under monochromatic blue, red and green light. These results suggest that chlorophyll fluorescence profiles may be used to estimate the distribution of quanta that are absorbed within the leaf for photosynthesis.     

Dark Ammonium Assimilation Reduces the Plastoquinone Pool of Photosystem II in the Green Alga Selenastrum minutum

The impact of dark NH₄⁺ and NO₃⁻ assimilation on photosynthetic light harvesting capability of the green alga Selenastrum minutum was monitored by chlorophyll a fluorescence analysis. When cells assimilated NH₄⁺, they exhibited a large decline in the variable fluorescence/maximum fluorescence ratio, the fluorescence yield of photosystem II relative to that of photosystem I at 77 kelvin, and O₂ evolution rate. NH₄⁺ assimilation therefore poised the cells in a less efficient state for photosystem II. The analysis of complementary area of fluorescence induction curve and the pattern of fluorescence decay upon microsecond saturating flash, indicators of redox state of plastoquinone (PQ) pool and dark reoxidation of primary quinone electron acceptor (Q_A), respectively, revealed that the PQ pool became reduced during dark NH₄⁺ assimilation. NH₄⁺ assimilation also caused an increase in the NADPH/NADP⁺ ratio due to the NH₄⁺ induced increase in respiratory carbon oxidation. The change in cellular reductant is suggested to be responsible for the reduction of the PQ pool and provide a mechanism by which the metabolic demands of NH₄⁺ assimilation may alter the efficiency of photosynthetic light harvesting. NO₃⁻ assimilation did not cause a reduction in PQ and did not affect the efficiency of light harvesting. These results illustrate the role of cellular metabolism in the modulating photosynthetic processes.     

On the advantages of using green light to study fluorescence yield changes in leaves

In photosynthetic chains, the kinetics of fluorescence yield depends on the photochemical rates at the level of both Photosystem I and II and thus on the absorption cross section of the photosynthetic units as well as on the coupling between light harvesting complexes and photosynthetic traps. A new set-up is described which, at variance with the commonly used set-ups, uses of a weakly absorbed light source (light-emitting diodes with maximum output at 520 nm) to excite the photosynthetic electron chain and probe the resulting fluorescence yield changes and their time course. This approach optimizes the homogeneity of the exciting light throughout the leaf and we show that this homogeneity narrows the distribution of the photochemical rates. Although the exciting light is weakly absorbed, the possibility to tune the intensity of the light emitting diodes allows one to reach photochemical rates ranging from 10⁴ s^− 1 to 0.25 s^− 1 rendering experimentally accessible different functional regimes. The variations of the fluorescence yield induced by the photosynthetic activity are qualitatively and quantitatively discussed. When illuminating dark-adapted leaves by a weak light, the kinetics of fluorescence changes displays a pronounced plateau which precedes the fluorescence increase reflecting the full reduction of the plastoquinone pool. We ascribe this plateau to the time delay needed to reduce the photosystem I electron acceptors.     

Adaptation of the photosynthetic apparatus of Nitellopsis obtusa to changing light intensity at the molecular level: different pathways of a singlet excitation quenching

The effect of prolonged illumination (60 min) with photosynthetically active monochromatic radiation of low intensity (3 μmol m⁻² s⁻¹) and high intensity (60 μmol m⁻² s⁻¹), corresponding to the physiological conditions and light stress conditions, respectively, was studied in the algae Nitellopsis obtusa. Illumination of Nitellopsis obtusa cells with strong light was associated with activation of the xanthophyll cycle, manifested by the deepoxidation of violaxanthin and accumulation of antheraxanthin and zeaxanthin. At the same time, the efficient singlet excitation quenching in the photosynthetic apparatus was activated, as demonstrated by the decrease in the intensity of the chlorophyll a fluorescence emission by ca 50 %. The difference of the fluorescence excitation spectra recorded before and after the light treatment match the difference absorption spectrum of the xanthophyll cycle pigments. The illumination with low light intensity resulted also in the chlorophyll a fluorescence quenching but the effect was very small (less than 10 %). The fluorescence quenching is interpreted in terms of the energy transfer between the Q_y energy level of chlorophyll a and the 2¹ A_g ⁻ energy level of zeaxanthin. The singlet energy levels of carotenoids, corresponding to the green spectral region, are also taken into consideration in the interpretation of the excitation energy exchange between the carotenoids and chlorophylls. Possible molecular mechanisms involved in the activation of the strong and the weak excitation quenching, including violaxanthin isomerization, and possible physiological functions of such pathways of energy transfer are discussed.     

Simultaneous gas exchange and fluorescence measurements indicate differences in the response of sunflower,bean and maize to water stress

Gas exchange and fluorescence measurements of attached leaves of water stressed bean, sunflower and maize plants were carried out at two light intensities (250 mol quanta m^-2s^-1 and 850 mol quanta m^-2s^-1). Besides the restriction of transpiration and CO₂ uptake, the dissipation of excess light energy was clearly reflected in the light and dark reactions of photosynthesis under stress conditions. Bean and maize plants preferentially use non-photochemical quenching for light energy dissipation. In sunflower plants, excess light energy gave rise to photochemical quenching. Autoradiography of leaves after photosynthesis in ¹⁴CO₂ demonstrated the occurrence of leaf patchiness in sunflower and maize but not in bean. The contribution of CO₂ recycling within the leaves to energy dissipation was investigated by studies in 2.5% oxygen to suppress photorespiration. The participation of different energy dissipating mechanisms to quanta comsumption on agriculturally relevant species is discussed.Abbreviations F_o minimal fluorescence - F_m maximal fluorescence - F_p peak fluorescence - g leaf conductance - P_N net CO₂ uptake - q_N coefficient of non-photochemical quenching - q_P coefficient of photochemical quenching     

Acclimation of Arabidopsis thaliana to the light environment: changes in photosynthetic function

Arabidopsis thaliana (L.) Heynh. cv. Landsberg erecta was grown under light regimes of differing spectral qualities, which results in differences in the stoichiometries of the two photosynthetic reaction centres. The acclimative value of these changes was investigated by assessing photosynthetic function in these plants when exposed to two spectrally distinct actinic lights. Plants grown in an environment enriched in far-red light were better able to make efficient use of non-saturating levels of actinic light enriched in long-wavelength red light. Simultaneous measurements of chlorophyll fluorescence and absorption changes at 820 nm indicated that differences between plants grown under alternative light regimes can be ascribed to imbalances in excitation of photosystems I and II (PSI, PSII). Measurements of chlorophyll fluorescence emission and excitation spectra at 77 K provided strong evidence that there was little or no difference in the composition or function of PSI or PSII between the two sets of plants, implying that changes in photosynthetic stoichiometry are primarily responsible for the observed differences in photosynthetic function.Abbreviations Chl chlorophyll - FR far-red light - HF highirradiance FR-enriched light (400 mol·m^–2·s^–1, RFR = 0.72) - HW high-irradiance white light (400 mol·m^–2 1·1 s^–1RFR = 1.40) - LHCI, LHCII light-harvesting complex of PSI, PSII - qO quenching of dark-level chlorophyll fluorescence - qN non-photochemical quenching of variable chlorophyll fluorescence - qP photochemical quenching of variable chlorophyll fluorescence - R red light - Rubisco ribulose-1,5-bisphosphate carboxylase/oxygenase We thank Dr. Sasha Ruban for assistance with the 77 K fluorescence measurements and for helpful discussions. This work was supported by Natural Environment Research Council Grant GR3/7571A.     

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.     

A quantitative determination of photochemical and non-photochemical quenching during the slow phase of the chlorophyll fluorescence induction curve of bean leaves

Estimations of the changes in the reduction-oxidation state of Photosystem II electron acceptors in Phaseolus vulgaris leaves were made during the slow decline in chlorophyll fluorescence emission from the maximal level at P to the steady-state level at T. The relative contributions of photochemical and non-photochemical processes to the fluorescence quenching were determined from these data. At a low photon flux density of 100 μmol · m^?2 · s^?1, non-photochemical quenching was the major contributor to the fluorescence decline from P to T, although large charges were observed in photochemical quenching immediately after P. On increasing the light intensity 10-fold, the contribution of photochemical processes to fluorescence quenching was markedly diminished, with nearly all the P-to-T fluorescence decline being attributable to changes in non-photochemical quenching. The possible factors responsible for changes in non-photochemical quenching within the leaves are discussed.     

Chlorophyll fluorescence in the leaves of Tradescantia species of different ecological groups: Induction events at different intensities of actinic light

Chlorophyll fluorescence analysis is one of the most convenient and widespread techniques used to monitor photosynthesis performance in plants. In this work, after a brief overview of the mechanisms of regulation of photosynthetic electron transport and protection of photosynthetic apparatus against photodamage, we describe results of our study of the effects of actinic light intensity on photosynthetic performance in Tradescantia species of different ecological groups. Using the chlorophyll fluorescence as a probe of photosynthetic activity, we have found that the shade-tolerant species Tradescantia fluminensis shows a higher sensitivity to short-term illumination (≤20 min) with low and moderate light (≤200 μE m⁻² s⁻¹) as compared with the light-resistant species Tradescantia sillamontana. In T. fluminensis, non-photochemical quenching of chlorophyll fluorescence (NPQ) and photosystem II operational efficiency (parameter Φ_PSII) saturate as soon as actinic light reaches ≈200 μE m⁻² s⁻¹. Otherwise, T. sillamontana revealed a higher capacity for NPQ at strong light (≥800 μE m⁻² s⁻¹). The post-illumination adaptation of shade-tolerant plants occurs slower than in the light-resistant species. The data obtained are discussed in terms of reactivity of photosynthetic apparatus to short-term variations of the environment light.     

Analysis of the Heat Stress Induced Quenching of Variable Chlorophyll a Fluorescence in Beet Spinach Leaves: Possible Accumulation of Oxidized Quencher P680+ under High Illumination

Effect of preheating of beet spinach leaves on chlorophyll a fluorescence yield was analyzed with the help of additional high intensity illumination pulses using a pulse modulated fluorometer. Preheating at mildly elevated temperature (35–45°C) causes a shift in the redox state of secondary donor of photosystem II, possibly due to uncoupling of phosphorylation because of thermal induced membrane disorganization and associated alkalinization of intra thylakoid space. Also, at these preheating temperatures, a rise in photosystem I catalyzed electron transfer has been shown to occur. These two effects induce rapid quenching of Chi a fluorescence, which drops even in the presence of actinic light, below the level of initial fluorescence (F_o′ monitored by the weak modulated probing light. Preheating of leaf segments induces an increase in fluorescence in the presence of dluron, which blocks electron flow between two photosystems, and thus this increases in fluorescence yield (F_o′ as monitored by weak modulated light, is not solely due to disorganization of light harvesting Chi-protein complex but also due to a shift in the redox equilibrium of the donor at the oxidizing side of photosystem II resulting in rapid reduction of Q_A the stable primary acceptor of photosystem II. In 50°C preheated DCMU treated samples, the fluorescence yield increases in weak modulated light and it approaches that of maximal steady state (F_max) level. At preheating temperature of 48°–50°C, the inactivation of enzymes in the reducing side of photosystem I, causes an impairment of the reoxidation of QA and under this condition, a strong illumination causes quenching of Chi a fluorescence. This quenching seems to arise because of accumulation of the P680⁺, the oxidized physiological donor of photosystem which is a quencher of Chi a fluorescence. This quenching depended on the pulse intensity and duration which saturates P680⁺ accumulation and is greatly manifested when water oxidation complex is damaged.     

Variable chlorophyll a fluorescence from P-700 enriched Photosystem I particles dependent on the redox state of the reaction centre

1. Photosystem I particles enriched in P-700 prepared by Triton X-100 treatment of chloroplasts show a light-induced increase in fluorescence yield of more than 100% in the presence of dithionite but not in its absence.2. Steady state light maintains the P-700, of these particles, in the oxidised state when ascorbate is present but in the presence of dithionite only a transient oxidation occurs.3. EPR data show that, in these particles, the primary electron acceptor (X) is maintained in the reduced state by light at room temperature only when the dithionite is also present. In contrast, the secondary electron acceptors are reduced in the dark by dithionite.4. Fluorescence emission and excitation spectra and fluorescence lifetime measurements for the constant and variable fluorescence indicate a heterogeneity of the chlorophyll in these particles.5. It is concluded that the variable fluorescence comes from those chlorophylls which can transfer their energy to the reaction centre and that the states PX and P⁺X are more effective quenchers of chlorophyll fluorescence than PX^?, where P is P-700.     

On the nature of the fluorescence decrease due to phosphorylation of chloroplast membrane proteins

1. Phosphorylation of chloroplast membranes by illumination in the presence of ATP results in a 15–20% increase in the rate of Photosystem I electron transfer at low light intensity. 2. Phosphorylated membranes when depleted of Mg²⁺ and resuspended in a low salt medium still show a 17% lower yield of Photosystem II fluorescence than do unphosphorylated membranes. A 31% difference is seen after restoration of the maximal yield by addition of Mg²⁺. 3. The concentration of Mg²⁺ required to induce a half-maximal increase in fluorescence is 0.9 mM for control and 1.8 mM for phosphorylated chloroplasts. Phosphorylation at 1 mM Mg²⁺ can therefore cause more than double the amount of decrease in fluorescence yield from Photosystem II compared to phosphorylation at 5 mM. 4. The above results are discussed in terms of the mechanism of the ATP-induced fluorescence changes and a suggestion is made that the apparent interaction between phosphorylation and Mg²⁺ concentration may be a physiologically important phenomenon.     

Modulation of the fluorescence yield in heliobacterial cells by induction of charge recombination in the photosynthetic reaction center

Heliobacteria contain a very simple photosynthetic apparatus, consisting of a homodimeric type I reaction center (RC) without a peripheral antenna system and using the unique pigment bacteriochlorophyll (BChl) g. They are thought to use a light-driven cyclic electron transport pathway to pump protons, and thereby phosphorylate ADP, although some of the details of this cycle are yet to be worked out. We previously reported that the fluorescence emission from the heliobacterial RC in vivo was increased by exposure to actinic light, although this variable fluorescence phenomenon exhibited very different characteristics to that in oxygenic phototrophs (Collins et al. 2010). Here, we describe the underlying mechanism behind the variable fluorescence in heliobacterial cells. We find that the ability to stably photobleach P₈₀₀, the primary donor of the RC, using brief flashes is inversely correlated to the variable fluorescence. Using pump-probe spectroscopy in the nanosecond timescale, we found that illumination of cells with bright light for a few seconds put them in a state in which a significant fraction of the RCs underwent charge recombination from P₈₀₀ ⁺A₀ ^? with a time constant of ~20 ns. The fraction of RCs in the rapidly back-reacting state correlated very well with the variable fluorescence, indicating that nearly all of the increase in fluorescence could be explained by charge recombination of P₈₀₀ ⁺A₀ ^?, some of which regenerated the singlet excited state. This hypothesis was tested directly by time-resolved fluorescence studies in the ps and ns timescales. The major decay component in whole cells had a 20-ps decay time, representing trapping by the RC. Treatment of cells with dithionite resulted in the appearance of a ~18-ns decay component, which accounted for ~0.6 % of the decay, but was almost undetectable in the untreated cells. We conclude that strong illumination of heliobacterial cells can result in saturation of the electron acceptor pool, leading to reduction of the acceptor side of the RC and the creation of a back-reacting RC state that gives rise to delayed fluorescence.     

Analysis of chlorophyll fluorescence quenching by DBMIB as a means of investigating the consequences of thylakoid membrane phosphorylation

The effect of Mg²⁺ concentration and phosphorylation of the light harvesting chlorophyll $\text{a}\text{b}$ protein on the ability of DBMIB to quench chlorophyll fluorescence of isolated pea thylakoids has been studied. Over a wide range of Mg²⁺ concentrations (5?0.33 mM), the observed changes in fluorescence yield are mirrored by similar changes in the quenching ability of DBMIB, indicating that the cation-induced phenomenon involves alterations in radiative lifetimes. In contrast, phosphorylation at 10 mM Mg²⁺ brings about a lowering of the chlorophyll fluorescence yield, while having no effect on the quenching capacity of DBMIB. This result can be interpreted as a phosphorylation-induced decrease in PS II absorption cross-section. At Mg²⁺ levels between 5 and 1 mM, phosphorylation leads to a change in the quenching of fluorescence by DBMIB, when compared with non-phosphorylated thylakoids. At these cation levels, the degree of DBMIB-induced quenching cannot wholly account for the observed changes in chlorophyll fluorescence due to phosphorylation. It is concluded that the phosphorylation- and Mg²⁺-induced changes in fluorescence yield are independent but inter-related processes which involve surface charge screening as emphasised by the change in cation sensitivity of the DBMIB quenching before and after phosphorylation.     

Effects of Cations and Abscisic Acid on Chlorophyll a Fluorescence in Guard Cells of Vicia faba

The effects of cations and abscisic acid on chloroplast activity in guard cells of Vicia faba were investigated by analysis of the transient of chlorophyll a fluorescence. When epidermal strips containing guard cells as the only living cells were incubated in water and illuminated with strong light, chlorophyll a fluorescence rose rapidly to a high intensity and then declined slowly to a stationary level. The rate of this decline was enhanced by K⁺ or Na⁺, and the effect of these cations was greater when added with phosphate than with chloride as the anion. Ca²⁺ suppressed the enhancement by Na⁺ and, to a lesser extent, that by K⁺. Abscisic acid also suppressed the enhancement by K⁺ and Na⁺. Since the fluorescence decline reflects the increase of intrathylakoid H⁺ concentration necessary for photophosphorylation, the acceleration of the decline by K⁺ (or Na⁺ in the absence of Ca²⁺) implicates chloroplast activity in ion accumulation by guard cells in the light. The differential effects of phosphate and chloride suggest that chloroplast activity may be involved in malate formation in guard cells in the light.     

Fixation of the plasma membrane/cytoplasm complex: A mechanism of toxic interaction of tributyltin with the cell

Flow cytometric and light/fluorescence microscopic analysis of murine erythroleukemic cells (MELC) and electron microscopic investigation of porcine microsomal membrane preparations suggest that tributyltin (TBT) toxicity is mediated through fixation processes (protein denaturation, crosslinking, and so on) within the plasma membrane/cytoplasm complex. This hypothesis was derived from the following observations:

1. Exposure of the MELC to micromolar concentrations of TBT results in increased resistance to detergent-mediated cytolysis;
2. Exposure of porcine renal microsomal membrane preparations to similar concentrations results in inhibition of vanadate-mediated crystallization of Na⁺,K⁺-ATPase, a process requiring protein mobility within the membrane;
3. Flow cytometric and fluorescence microscopic analyses indicate that MELC exposed to submicromolar concentrations of TBT exhibit increased cellular carboxyfluorescein retention; and
4. Nuclei prepared from TBT-treated cells by detergent-mediated cytolysis exhibit increased axial light loss, 90° light scatter, fluorescein isothiocyanate fluorescence, and the presence of adherent protein-aceous tags. The DNA distribution histogram of such nuclei also is perturbed.

     

Intravital Fluorescence Excitation in Whole-Animal Optical Imaging

Whole-animal fluorescence imaging with recombinant or fluorescently-tagged pathogens or cells enables real-time analysis of disease progression and treatment response in live animals. Tissue absorption limits penetration of fluorescence excitation light, particularly in the visible wavelength range, resulting in reduced sensitivity to deep targets. Here, we demonstrate the use of an optical fiber bundle to deliver light into the mouse lung to excite fluorescent bacteria, circumventing tissue absorption of excitation light in whole-animal imaging. We present the use of this technology to improve detection of recombinant reporter strains of tdTomato-expressing Mycobacterium bovis BCG (Bacillus Calmette Guerin) bacteria in the mouse lung. A microendoscope was integrated into a whole-animal fluorescence imager to enable intravital excitation in the mouse lung with whole-animal detection. Using this technique, the threshold of detection was measured as 10³ colony forming units (CFU) during pulmonary infection. In comparison, the threshold of detection for whole-animal fluorescence imaging using standard epi-illumination was greater than 10⁶ CFU.     

A comparison between cation and protein phosphorylation effects on the fluorescence induction curve in chloroplasts treated with 3-(3,4-dichlorophenyl)-1,1-dimethylurea

Fluorescence induction curves in chloroplasts phosphorylated by the thylakoid protein kinase activated at low light intensity and high chlorophyll concentration have been measured. At 5 mM Mg²⁺, phosphorylation did not preferentially quench variable fluorescence. At 1 mM, preferential quenching of variable fluorescence was observed, indicating a second effect of phosphorylation at low Mg²⁺ (Horton, P. and Black, M.T. (1982) Biochim. Biophys. Acta 680, 22–27). Comparison of the extent of fluorescence decrease and the resulting ratio of variable to maximum fluorescence after phosphorylation and after lowering Mg²⁺ concentration demonstrated a difference between these two mechanisms of lowering of fluorescence. The significance of these results in terms of how phosphorylation may alter membrane organization is discussed.     

Alterations of pigment composition and their interactions in response to different light conditions in the diatom Chaetoceros gracilis probed by time-resolved fluorescence spectroscopy

Maintenance of energy balance under changeable light conditions is an essential function of photosynthetic organisms to achieve efficient photochemical reactions. Among the photosynthetic organisms, diatoms possess light-harvesting fucoxanthin chlorophyll (Chl) a/c-binding protein (FCP) as peripheral antennas. However, how diatoms regulate excitation-energy distribution between FCP and the two photosystem cores during light adaptation is poorly understood. In this study, we examined spectroscopic properties of a marine diatom Chaetoceros gracilis adapted in the dark and at photosynthetic photon flux density at 30 and 300?μmol?photons?m^?2?s^?1. Absorption spectra at 77?K showed significant changes in the Soret region, and 77-K steady-state fluorescence spectra showed significant differences in the spectral shape and relative fluorescence intensity originating from both PSII and PSI, among the cells grown under different light conditions. These results suggest alterations of pigment composition and their interactions under the different light conditions. These alterations affected the excitation-energy dynamics monitored by picosecond time-resolved fluorescence analyses at 77?K significantly. The contributions of Chls having lower energy levels than the reaction center Chls in the two photosystems to the energy dynamics were clearly identified in the three cells but with presumably different roles. These findings provide insights into the regulatory mechanism of excitation-energy balance in diatoms under various light conditions.