RESUMO
Amphotericin B is a popular antifungal antibiotic, but the exact way it works is still a matter of debate. Here, we used monolayers composed of phosphatidylcholine with ergosterol as a model of fungal lipid membranes to study drug incorporation from the aqueous phase and analyze the molecular reorganization of membranes underlying the biological activity of the antibiotic. The results show that the internalization of antibiotic molecules into membranes occurs only in the presence of ergosterol in the lipid phase. Comparison of images of solid-supported monolayers obtained by atomic force microscopy and lifetime imaging fluorescence microscopy shows the formation of intramembrane clusters of various sizes in the lipid phase, consisting mainly of antibiotic dimers and relatively large membrane pores (â¼15 nm in diameter). The results reveal multiple modes of action of amphotericin B, acting simultaneously, each of which adversely affects the structural properties of the lipid membranes and their physiological functionality.
Assuntos
Anfotericina B , Fosfatidilcolinas , Anfotericina B/química , Fosfatidilcolinas/química , Ergosterol/química , Antifúngicos/química , Microscopia de Força Atômica , Antibacterianos/química , Membrana Celular/química , Microscopia de FluorescênciaRESUMO
Here, we address the problem of the antioxidant activity of carotenoids in biomembranes. The activity of lutein and zeaxanthin in the quenching of singlet oxygen generated by photosensitization was monitored in lipid vesicles using a singlet oxygen-sensitive fluorescent probe and with the application of fluorescence lifetime imaging microscopy. The antioxidant activity of xanthophylls was interpreted on the basis of electron paramagnetic resonance oximetry results showing that xanthophylls constitute a barrier to the penetration of molecular oxygen into lipid membranes: to a greater extent in the 13-cis configuration than in all-trans. These results are discussed in relation to the trans-cis photoisomerization of xanthophylls observed in the human retina. It can be concluded that photoisomerization of xanthophylls is a regulatory mechanism that is important for both the modulation of light filtration through the macula and photoprotection by quenching singlet oxygen and creating a barrier to oxygen permeation to membranes.
Assuntos
Antioxidantes , Xantofilas , Humanos , Xantofilas/metabolismo , Oxigênio Singlete , Estresse Oxidativo , Oxigênio , LipídeosRESUMO
Zeaxanthin and lutein are xanthophyll pigments present in the human retina and particularly concentrated in its center referred to as the yellow spot (macula lutea). The fact that zeaxanthin, including its isomer meso-zeaxanthin, is concentrated in the central part of the retina, in contrast to lutein also present in the peripheral regions, raises questions about the possible physiological significance of such a heterogeneous distribution of macular xanthophylls. Here, we attempt to address this problem using resonance Raman spectroscopy and confocal imaging, with different laser lines selected to effectively distinguish the spectral contribution of lutein and zeaxanthin. Additionally, fluorescence lifetime imaging microscopy (FLIM) is used to solve the problem of xanthophyll localization in the axon membranes. The obtained results allow us to conclude that one of the key advantages of a particularly high concentration of zeaxanthin in the central part of the retina is the high efficiency of this pigment in the dynamic filtration of light with excessive intensity, potentially harmful for the photoreceptors.
Assuntos
Luteína , Macula Lutea , Humanos , Luteína/química , Zeaxantinas , beta Caroteno , Retina/química , Xantofilas/análise , Macula Lutea/químicaRESUMO
Amphotericin B is a popular antifungal antibiotic, and despite decades of pharmacological application, the exact mode of its biological activity is still a matter of debate. Amphotericin B-silver hybrid nanoparticles (AmB-Ag) have been reported to be an extremely effective form of this antibiotic to combat fungi. Here, we analyze the interaction of AmB-Ag with C. albicans cells with the application of molecular spectroscopy and imaging techniques, including Raman scattering and Fluorescence Lifetime Imaging Microscopy. The results lead to the conclusion that among the main molecular mechanisms responsible for the antifungal activity of AmB is the disintegration of the cell membrane, which occurs on a timescale of minutes.
Assuntos
Anfotericina B , Nanopartículas , Anfotericina B/farmacologia , Anfotericina B/química , Antibacterianos/análise , Prata/química , Antifúngicos/química , Membrana Celular/metabolismo , Nanopartículas/química , Candida albicansRESUMO
Amphotericin B (AmB) is a life-saving and widely used antifungal antibiotic, but its therapeutic applicability is limited due to severe side effects. Here, we report that the formulation of the drug based on a complex with albumin (BSA) is highly effective against Candida albicans at relatively low concentrations, which implies lower toxicity to patients. This was also concluded based on the comparison with antifungal activities of other popular commercial formulations of the drug, such as Fungizone and AmBisome. Several molecular spectroscopy and imaging techniques, e.g., fluorescence lifetime imaging microscopy (FLIM), were applied to understand the phenomenon of enhanced antifungal activity of the AmB-BSA complex. The results show that the drug molecules bound to the protein remain mostly monomeric and are most likely bound in the pocket responsible for the capture of small molecules by this transport protein. The results of molecular imaging of single complex particles indicate that in most cases, the antibiotic-protein stoichiometry is 1:1. All of the analyses of the AmB-BSA system exclude the presence of the antibiotic aggregates potentially toxic to patients. Cell imaging shows that BSA-bound AmB molecules can readily bind to fungal cell membranes, unlike drug molecules present in the aqueous phase, which are effectively retained by the cell wall barrier. The advantages and prospects of pharmacological use of AmB complexed with proteins are discussed.
Assuntos
Anfotericina B , Antifúngicos , Antifúngicos/farmacologia , Antifúngicos/química , Anfotericina B/farmacologia , Anfotericina B/química , Candida albicans , Albuminas , Antibacterianos/farmacologiaRESUMO
The safe and smooth functioning of photosynthesis in plants is ensured by the operation of numerous regulatory mechanisms that adjust the density of excitation resulting from photon absorption to the capabilities of the photosynthetic apparatus. Such mechanisms include the movement of chloroplasts inside cells and the quenching of electronic excitations in the pigment-protein complexes. Here, we address the problem of a possible cause-and-effect relationship between these two mechanisms. Both the light-induced chloroplast movements and quenching of chlorophyll excitations were analyzed simultaneously with the application of fluorescence lifetime imaging microscopy of Arabidopsis thaliana leaves, wild-type and impaired in chloroplast movements or photoprotective excitation quenching. The results show that both regulatory mechanisms operate over a relatively wide range of light intensities. By contrast, impaired chloroplast translocations have no effect on photoprotection at the molecular level, indicating the direction of information flow in the coupling of these two regulatory mechanisms: from the photosynthetic apparatus to the cellular level. The results show also that the presence of the xanthophyll zeaxanthin is necessary and sufficient for the full development of photoprotective quenching of excessive chlorophyll excitations in plants.
Assuntos
Arabidopsis , Cloroplastos , Cloroplastos/metabolismo , Fotossíntese , Clorofila/metabolismo , Xantofilas/metabolismoRESUMO
In the animal kingdom, continuously erupting incisors provided an attractive model for studying the enamel matrix and mineral composition of teeth during development. Enamel, the hardest mineral tissue in the vertebrates, is a tissue sensitive to external conditions, reflecting various disturbances in its structure. The developing dental enamel was monitored in a series of incisor samples extending the first four weeks of postnatal life in the spiny mouse. The age-dependent changes in enamel surface morphology in the micrometre and nanometre-scale and a qualitative assessment of its mechanical features were examined by applying scanning electron microscopy (SEM) and atomic force microscopy (AFM). At the same time, structural studies using XRD and vibrational spectroscopy made it possible to assess crystallinity and carbonate content in enamel mineral composition. Finally, a model for predicting the maturation based on chemical composition and structural factors was constructed using artificial neural networks (ANNs). The research presented here can extend the existing knowledge by proposing a pattern of enamel development that could be used as a comparative material in environmental, nutritional, and pharmaceutical research.
RESUMO
The functioning of the human eye in the extreme range of light intensity requires a combination of the high sensitivity of photoreceptors with their photostability. Here, we identify a regulatory mechanism based on dynamic modulation of light absorption by xanthophylls in the retina, realized by reorientation of pigment molecules induced by trans-cis photoisomerization. We explore this photochemically switchable system using chromatographic analysis coupled with microimaging based on fluorescence lifetime and Raman scattering, showing it at work in both isolated human retina and model lipid membranes. The molecular mechanism underlying xanthophyll reorientation is explained in terms of hydrophobic mismatch using molecular dynamics simulations. Overall, we show that xanthophylls in the human retina act as "molecular blinds", opening and closing on a submillisecond timescale to dynamically control the intensity of light reaching the photoreceptors, thus enabling vision at a very low light intensity and protecting the retina from photodegradation when suddenly exposed to strong light.
Assuntos
Retina , Protetores Solares , Humanos , Luteína , Análise Espectral Raman , XantofilasRESUMO
Safe operation of photosynthesis is vital to plants and is ensured by the activity of processes protecting chloroplasts against photo-damage. The harmless dissipation of excess excitation energy is considered to be the primary photoprotective mechanism and is most effective in the combined presence of PsbS protein and zeaxanthin, a xanthophyll accumulated in strong light as a result of the xanthophyll cycle. Here we address the problem of specific molecular mechanisms underlying the synergistic effect of zeaxanthin and PsbS. The experiments were conducted with Arabidopsis thaliana, using wild-type plants, mutants lacking PsbS (npq4), and mutants affected in the xanthophyll cycle (npq1), with the application of molecular spectroscopy and imaging techniques. The results lead to the conclusion that PsbS interferes with the formation of densely packed aggregates of thylakoid membrane proteins, thus allowing easy exchange and incorporation of xanthophyll cycle pigments into such structures. It was found that xanthophylls trapped within supramolecular structures, most likely in the interfacial protein region, determine their photophysical properties. The structures formed in the presence of violaxanthin are characterized by minimized dissipation of excitation energy. In contrast, the structures formed in the presence of zeaxanthin show enhanced excitation quenching, thus protecting the system against photo-damage.
Assuntos
Proteínas de Arabidopsis/metabolismo , Complexos de Proteínas Captadores de Luz/metabolismo , Fotossíntese , Complexo de Proteína do Fotossistema II/metabolismo , Zeaxantinas/metabolismo , Arabidopsis/metabolismo , Clorofila/metabolismo , Metabolismo Energético , Luz , Microscopia de Fluorescência , Folhas de Planta/metabolismo , Análise Espectral Raman , Tilacoides/metabolismo , Tilacoides/efeitos da radiação , Tilacoides/ultraestruturaRESUMO
Raman spectroscopic imaging and mapping were applied to characterise three-compound ceramic composite biomaterial consisting of chitosan, ß-1,3-d-glucan (curdlan) and hydroxyapatite (HA) developed as a bone tissue engineering product (TEP). In this rapidly advancing domain of medical science, the urge for quick, reliable and specific method for products evaluation and tissue-implant interaction, in this case bone formation process, is constantly present. Two types of stem cells, adipose-derived stem cells (ADSCs) and bone marrow-derived stem cells (BMDSCs), were cultured on composite surface. Raman spectroscopic imaging provided advantageous information on molecular differences and spatial distribution of compounds within and between the cell-seeded and untreated samples at a microscopic level. With the use of this, it was possible to confirm composite biocompatibility and bioactivity in vitro. Deposition of HA and changes in its crystallinity along with protein adsorption proved new bone tissue formation in both mesenchymal stem cell samples, where the cells proliferated, differentiated and produced biomineralised extracellular matrix (ECM). The usefulness of spectroscopic Raman imaging was confirmed in tissue engineering in terms of both the organic and inorganic components considering composite-cells interaction.
Assuntos
Microscopia Confocal/métodos , Análise Espectral Raman , Alicerces Teciduais/química , Tecido Adiposo/citologia , Materiais Biocompatíveis/química , Materiais Biocompatíveis/farmacologia , Células da Medula Óssea/citologia , Células Cultivadas , Quitosana/química , Durapatita/química , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/metabolismo , Engenharia TecidualRESUMO
Photosystem II (PSII) converts light into chemical energy powering almost all life on Earth. The primary photovoltaic reaction in the PSII reaction center requires energy corresponding to 680 nm, which is significantly higher than in the case of the low-energy states in the antenna complexes involved in the harvesting of excitations driving PSII. Here we show that despite seemingly insufficient energy, the low-energy excited states can power PSII because of the activity of the thermally driven up-conversion. We demonstrate the operation of this mechanism both in intact leaves and in isolated pigment-protein complex LHCII. A mechanism is proposed, according to which the effective utilization of thermal energy in the photosynthetic apparatus is possible owing to the formation of LHCII supramolecular structures, leading to the coupled energy levels corresponding to approximately 680 and 700 nm, capable of exchanging excitation energy through the spontaneous relaxation and the thermal up-conversion.
Assuntos
Transferência de Energia , Complexos de Proteínas Captadores de Luz/metabolismo , Complexo de Proteína do Fotossistema II/metabolismo , Arabidopsis/metabolismo , Clorofila A/metabolismo , Temperatura Alta , Folhas de Planta/metabolismo , Reciclagem , Espectrometria de FluorescênciaRESUMO
Amphotericin B is an antibiotic used as the "gold standard" in the treatment of life-threatening fungal infections. Several molecular mechanisms have been proposed to explain exceptionally high effectiveness of amphotericin B in combating fungi. In the present work, we apply fluorescence lifetime imaging microscopy to track, step by step, modes of the toxic activity of amphotericin B towards a clinical strain of Candida albicans. The images recorded reveal that the antibiotic binds to cells in the form of the small aggregates characterized by a relatively short fluorescence lifetime (0.2 ns). Amphotericin B binds preferentially to the cell walls of mature cells but also to the plasma membranes of the daughter cells at the budding stage. The images recorded with the application of a scanning electron microscopy show that the antibiotic interferes with the formation of functional cell walls of such young cells. The results of imaging reveal the formation of the amphotericin B-rich extramembranous structures and also binding of the drug molecules into the cell membranes and penetration into the cells. These two modes of action of amphotericin B are observed in the time scale of minutes.
Assuntos
Anfotericina B/farmacologia , Antifúngicos/farmacologia , Candida albicans/efeitos dos fármacos , Membrana Celular/metabolismo , Parede Celular/metabolismo , Candida albicans/crescimento & desenvolvimento , Candidíase/tratamento farmacológico , Humanos , Testes de Sensibilidade Microbiana , Microscopia de Fluorescência/métodosRESUMO
Amphotericin B is an antibiotic used in pharmacotherapy of life-threatening mycotic infections. Unfortunately, the applicability of this antibiotic is associated with highly toxic side effects. In order to understand molecular mechanisms underlying toxicity of amphotericin B to patients, two cell lines, human normal colon epithelial cells (CCD 841 CoTr) and human colon adenocarcinoma cells (HT-29) were cultured in the presence of the drug and imaged with the application of fluorescence lifetime imaging microscopy and Raman scattering microscopy. The results of the cell viability assays confirm high toxicity of amphotericin B towards human cells. The images recorded demonstrate effective binding of amphotericin B to biomembranes. Analysis of the images reveals the operation of a defence mechanism based upon the elimination of molecules of the drug from living cells via formation of small amphotericin B-containing lipid vesicles. The fact that exosomes formed are devoid of cholesterol, as concluded on the basis of the results of Raman analysis, suggests that sequestration of sterols from the lipid phase of biomembranes is not a sole mechanism responsible for the toxic side effects of amphotericin B. Alternatively, the results imply that molecules of the drug present directly within the hydrophobic membrane core disturb the lipid membrane structure and affect their biological functions.
Assuntos
Anfotericina B/toxicidade , Antibacterianos/toxicidade , Antifúngicos/toxicidade , Microscopia de Fluorescência , Sobrevivência Celular/efeitos dos fármacos , Células HT29 , Humanos , Análise Espectral RamanRESUMO
Amphotericin B is a lifesaving polyene antibiotic used in the treatment of systemic mycoses. Unfortunately, the pharmacological applicability of this drug is limited because of its severe toxic side effects. At the same time, the lack of a well-defined mechanism of selectivity hampers the efforts to rationally design safer derivatives. As the drug primarily targets the biomembranes of both fungi and humans, new insights into the binding of amphotericin B to lipid membranes can be helpful in unveiling the molecular mechanisms underlying both its pharmacological activity and toxicity. We use fluorescence-lifetime-imaging microscopy combined with fluorescence-emission spectroscopy in the microscale to study the interaction of amphotericin B with single lipid bilayers, using model systems based on giant unilamellar liposomes formed with three lipids: dipalmitoylphosphatidylcholine (DPPC), dimirystoylphosphatidylcholine (DMPC), and 1-palmitoyl-2-oleoylphosphatidylcholine (POPC). The results show that amphotericin B introduced into the water phase as a DMSO solution binds to the membrane as dimers and small-molecular aggregates that we identify as tetramers and trimers. Fluorescence-detected linear-dichroism measurements revealed high orientational freedom of all the molecular-organization forms with respect to the membrane plane, which suggests that the drug partially binds to the membrane surface. The presence of sterols in the lipid phase (cholesterol but particularly ergosterol at 30 mol %) promotes the penetration of drug molecules into the lipid membrane, as concluded on the basis of the decreased orientation angle of amphotericin B molecules with respect to the axis normal to the membrane plane. Moreover, ergosterol facilitates the association of amphotericin B dimers into aggregated structures that can play a role in membrane destabilization or permeabilization. The presence of cholesterol inhibits the formation of small aggregates in the lipid phase of liposomes, making this system a promising candidate for a low-toxicity antibiotic-delivery system. Our conclusions are supported with molecular simulations that reveal the conformational properties of AmB oligomers in both aqueous solution and lipid bilayers of different compositions.
Assuntos
Anfotericina B/química , Antifúngicos/química , 1,2-Dipalmitoilfosfatidilcolina/química , Colesterol/química , Simulação de Dinâmica Molecular , Fosfatidilcolinas/químicaRESUMO
Two cell lines, human normal colon epithelial cells (CCD 841 CoTr) and human colon adenocarcinoma cells (HT-29) were cultured in the presence of exogenous carotenoids, either zeaxanthin or lutein. Both carotenoids demonstrated cytotoxicity with respect to cancer cells but not to normal cells. Cells from both the cell lines were analyzed with application of fluorescence lifetime imaging microscopy and Raman scattering microscopy. Both imaging techniques show effective incorporation of carotenoid molecules into growing cells. Comparison of the Raman scattering and fluorescence lifetime characteristics reveals different molecular organization of carotenoids in the carcinoma and normal cells. The main difference consists in a carotenoid aggregation level which is substantially lower in the carcinoma cells as compared to the normal cells. Different molecular organization of carotenoids was interpreted in terms of a different metabolism of normal and carcinoma cells and has been concluded to provide a possibility of cancer diagnosis based on spectroscopic analyses.
Assuntos
Adenocarcinoma/metabolismo , Carotenoides/química , Colo/metabolismo , Neoplasias do Colo/metabolismo , Células Epiteliais/metabolismo , Luteína/química , Zeaxantinas/química , Adenocarcinoma/patologia , Linhagem Celular Tumoral , Sobrevivência Celular , Neoplasias do Colo/patologia , Fluorescência , Humanos , Processamento de Imagem Assistida por Computador , Análise Espectral RamanRESUMO
Lensoside Aß, representing the flavonol glycosides, is a compound isolated from the aerial parts of edible lentil (Lens culinaris) cultivar Tina. This substance arouses interest because so far there is very little data about secondary metabolites isolated from the leaves and stems of this plant. Additionally, bioactive potential of flavonoids is directly coupled with the membranes as a primary target of their physiological and pharmacological activity. The aim of this study was to investigate the effect of lensoside Aß on lipid membranes. Interaction of examined compound with liposomes formed with dipalmitoylphosphatidylcholine (DPPC) was investigated with application of FTIR spectroscopy and 1H NMR technique. Molecular localization and orientation of lensoside Aß in a single lipid bilayer system represented by giant unilamellar vesicles, was also investigated with application of confocal fluorescence lifetime imaging microscopy (FLIM). FTIR analysis revealed that the tested compound incorporates into DPPC membranes via hydrogen bonding to lipid polar head groups in the PO2 group region and the COPOC segment. Furthermore 1H NMR analysis showed ordering effect in both the hydrophobic alkyl chains region and the polar heads of phospholipids. FLIM investigation has revealed roughly parallel orientation of its molecules in the membranes. This suggests that one of the possible physiological functions of this flavonol could be screening a cell against short-wavelength radiation.
Assuntos
Bicamadas Lipídicas/metabolismo , Lipossomos/metabolismo , Quercetina/metabolismo , Lipossomas Unilamelares/metabolismo , 1,2-Dipalmitoilfosfatidilcolina/química , 1,2-Dipalmitoilfosfatidilcolina/metabolismo , Ligação de Hidrogênio , Bicamadas Lipídicas/química , Lipossomos/química , Lipídeos de Membrana/química , Lipídeos de Membrana/metabolismo , Estrutura Molecular , Espectroscopia de Prótons por Ressonância Magnética , Quercetina/química , Espectroscopia de Infravermelho com Transformada de Fourier , Lipossomas Unilamelares/químicaRESUMO
Studies focused on GPCRs, particularly on the ß2-adrenergic receptor (ß2-AR), have demonstrated the relationship between ligand structure, receptor conformational changes and the corresponding pharmacological outcomes. Herein, we studied the molecular details of the rotameric flip of the W2866.48 sidechain, i.e. a presumed action switch that has not been reported in native ß2-AR thus far. It is believed that although both the 'active' and 'inactive' conformers of ß2-AR exhibit similar conformations of this switch, it may still play a substantial role in the ligand-induced activation of the receptor. By using both experimental methods (time-resolved fluorescence spectroscopy) and molecular modeling techniques (enhanced-sampling molecular dynamics), we characterized the conformational rearrangements of W2866.48 in relation to the type of ligand present in the binding cavity and to the conformation of the receptor ('active' vs. 'inactive' ß2-AR). We found that the conformational behaviour of W2866.48 is correlated with the pharmacological character of the ligand present in the binding cavity but not with the instantaneous conformation of the receptor. Namely, agonists promote the W2866.48 conformations that facilitate the increase of the solvation within the inner receptor channel. In contrast, antagonists and inverse agonists act toward the decrease of the solvation in the inner channel. This creates an opportunity for using computational methodologies in determining the pharmacological properties of various ligands. The combination of the time-resolved fluorescence spectroscopy technique with the enhanced-sampling molecular dynamics simulations is shown to be a powerful tool for studying the ligand-induced conformational rearrangements in GPCRs.
RESUMO
Xanthophylls (polar carotenoids) play diverse biological roles, among which are modulation of the physical properties of lipid membranes and protection of biomembranes against oxidative damage. Molecular mechanisms underlying these functions are intimately related to the localization and orientation of xanthophyll molecules in lipid membranes. In the present work, we address the problem of localization and orientation of two xanthophylls present in the photosynthetic apparatus of plants and in the retina of the human eye, zeaxanthin and lutein, in a single lipid bilayer membrane formed with dimyristoylphosphatidylcholine. By using fluorescence microscopic analysis and Raman imaging of giant unilamellar vesicles, as well as molecular dynamics simulations, we show that lutein and zeaxanthin adopt a very similar transmembrane orientation within a lipid membrane. In experimental and computational approach, the average tilt angle of xanthophylls relative to the membrane normal is independently found to be ~40 deg, and results from hydrophobic mismatch between the membrane thickness and the distance between the terminal hydroxyl groups of the xanthophylls. Consequences of such a localization and orientation for biological activity of xanthophylls are discussed.
RESUMO
It emerges from numerous experiments that LHCII, the major photosynthetic antenna complex of plants, can appear not only in the trimeric or monomeric states but also as a dimer. We address the problem whether the dimeric form of the complex is just a simple intermediate element of the trimer-monomer transformation or if it can also be a physiologically relevant molecular organization form? Dimers of LHCII were analyzed with application of native electrophoresis, time-resolved fluorescence spectroscopy, and fluorescence correlation spectroscopy. The results reveal the appearance of two types of LHCII dimers: one formed by the dissociation of one monomer from the trimeric structure and the other formed by association of monomers into a distinctively different molecular organizational form, characterized by a high rate of chlorophyll excitation quenching. The hypothetical structure of such an energy quencher is proposed. The high light-induced LHCII dimerization is discussed as a potential element of the photoprotective response in plants.
Assuntos
Complexo de Proteína do Fotossistema II/metabolismo , Tilacoides/metabolismo , Clorofila/metabolismo , Luz , Complexos de Proteínas Captadores de Luz/química , Complexos de Proteínas Captadores de Luz/metabolismo , Fotossíntese/efeitos da radiação , Complexo de Proteína do Fotossistema II/química , Estrutura Secundária de Proteína , Espectrometria de Fluorescência , Spinacia oleracea/metabolismo , Spinacia oleracea/efeitos da radiaçãoRESUMO
The efficient and fluent operation of photosynthesis in plants relies on activity of pigment-protein complexes called antenna, absorbing light and transferring excitations toward the reaction centers. Here we show, based on the results of the fluorescence lifetime imaging analyses of single chloroplasts, that pigment-protein complexes, in dark-adapted plants, are not able to act effectively as photosynthetic antennas, due to pronounced, adverse excitation quenching. It appeared that the antenna function could be activated by a short (on a minute timescale) illumination with light of relatively low intensity, substantially below the photosynthesis saturation threshold. The low-light-induced activation of the antenna function was attributed to phosphorylation of the major accessory light-harvesting complex LHCII, based on the fact that such a mechanism was not observed in the stn7 Arabidopsis thaliana mutant, with impaired LHCII phosphorylation. It is proposed that the protein phosphorylation-controlled change in the LHCII clustering ability provides mechanistic background for this regulatory process.