Photosystem II (PSII) uses solar energy to oxidize water and delivers electrons to fix CO2. Although the structure at atomic resolution and the basic photophysical and photochemical functions of PSII are well understood, many important questions remain. The activity of PSII in vitro and in vivo is routinely monitored by recording the induction kinetics of chlorophyll a fluorescence (ChlF). According to the 'mainstream' model, the rise from the minimum level (Fo) to the maximum (Fm) of ChlF of dark-adapted PSII reflects the closure of all functionally active reaction centers, and the Fv/Fm ratio is equated with the maximum photochemical quantum yield of PSII (where Fv=Fm-Fo). However, this model has never been free of controversies. Recent experimental data from a number of studies have confirmed that the first single-turnover saturating flash (STSF), which generates the closed state (PSIIC), produces F1

• Klíčová slova
• F v/Fm, Chlorophyll a fluorescence induction, QA-model, conformational changes, dielectric relaxation, electric field effects, light-adapted charge-separated state, photochemical quantum efficiency, photosystem II, purple bacterial reaction center,
• Publikační typ
• časopisecké články MeSH

Natural and synthetic polymers are widely explored for improving seed germination and plant resistance to environmental constraints. Here, for the first time, we explore stabilized nanomicelles composed of the biocompatible triblock co-polymer Pluronic P85 (SPM) as a priming agent for Pisum sativum (var. RAN-1) seeds. We tested a wide concentration range of 0.04-30 g(SPM) L-1. Applying several structural and functional methods we revealed that the utilized nanomicelles can positively affect root length, without any negative effects on leaf anatomy and photosynthetic efficiency at 0.2 g L-1, while strong negative effects were recorded for 10 and 30 g(SPM) L-1 concerning root length, leaf histology, and photoprotection capability. Our data strongly suggest that SPM can safely be utilized for seed priming at specific concentrations and are suitable objects for further loading with plant growth regulators.

• Klíčová slova
• chlorophyll fluorescence, garden pea, leaf anatomy, nanoparticles, plant biometry, poloxamer,
• Publikační typ
• časopisecké články MeSH

In our earlier works, we have identified rate-limiting steps in the dark-to-light transition of PSII. By measuring chlorophyll a fluorescence transients elicited by single-turnover saturating flashes (STSFs) we have shown that in diuron-treated samples an STSF generates only F1 (< Fm) fluorescence level, and to produce the maximum (Fm) level, additional excitations are required, which, however, can only be effective if sufficiently long Δτ waiting times are allowed between the excitations. Biological variations in the half-rise time (Δτ 1/2) of the fluorescence increment suggest that it may be sensitive to the physicochemical environment of PSII. Here, we investigated the influence of the lipidic environment on Δτ 1/2 of PSII core complexes of Thermosynechococcus vulcanus. We found that while non-native lipids had no noticeable effects, thylakoid membrane lipids considerably shortened the Δτ 1/2, from ~ 1 ms to ~ 0.2 ms. The importance of the presence of native lipids was confirmed by obtaining similarly short Δτ 1/2 values in the whole T. vulcanus cells and isolated pea thylakoid membranes. Minor, lipid-dependent reorganizations were also observed by steady-state and time-resolved spectroscopic measurements. These data show that the processes beyond the dark-to-light transition of PSII depend significantly on the lipid matrix of the reaction center.

• Klíčová slova
• closed state of PSII, conformational changes, dielectric relaxation, light-adapted state of PSII, light-induced changes, proteoliposomes.,
• Publikační typ
• časopisecké články MeSH

Photosystem II (PSII) uses solar energy to oxidize water and delivers electrons for life on Earth. The photochemical reaction center of PSII is known to possess two stationary states. In the open state (PSIIO), the absorption of a single photon triggers electron-transfer steps, which convert PSII into the charge-separated closed state (PSIIC). Here, by using steady-state and time-resolved spectroscopic techniques on Spinacia oleracea and Thermosynechococcus vulcanus preparations, we show that additional illumination gradually transforms PSIIC into a light-adapted charge-separated state (PSIIL). The PSIIC-to-PSIIL transition, observed at all temperatures between 80 and 308 K, is responsible for a large part of the variable chlorophyll-a fluorescence (Fv) and is associated with subtle, dark-reversible reorganizations in the core complexes, protein conformational changes at noncryogenic temperatures, and marked variations in the rates of photochemical and photophysical reactions. The build-up of PSIIL requires a series of light-induced events generating rapidly recombining primary radical pairs, spaced by sufficient waiting times between these events-pointing to the roles of local electric-field transients and dielectric relaxation processes. We show that the maximum fluorescence level, Fm, is associated with PSIIL rather than with PSIIC, and thus the Fv/Fm parameter cannot be equated with the quantum efficiency of PSII photochemistry. Our findings resolve the controversies and explain the peculiar features of chlorophyll-a fluorescence kinetics, a tool to monitor the functional activity and the structural-functional plasticity of PSII in different wild-types and mutant organisms and under stress conditions.

• MeSH
• chlorofyl analogy a deriváty   chemie   MeSH
• diuron farmakologie   MeSH
• fluorescence MeSH
• fluorescenční spektrometrie MeSH
• fotosystém II - proteinový komplex chemie   účinky léků   metabolismus   MeSH
• konformace proteinů MeSH
• spektroskopie infračervená s Fourierovou transformací MeSH
• Spinacia oleracea chemie   MeSH
• světlo MeSH
• teplota MeSH
• Thermosynechococcus chemie   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• chlorofyl MeSH
• chlorophyll a' MeSH Prohlížeč
• diuron MeSH
• fotosystém II - proteinový komplex MeSH

The thylakoid membranes of vascular plants are differentiated into stacked granum and unstacked stroma regions. The formation of grana is triggered by the macrodomain formation of photosystem II and light-harvesting complex II (PSII-LHCII) and thus their lateral segregation from the photosystem I-light-harvesting complex I (PSI-LHCI) super-complexes and the ATP-synthase; which is then stabilized by stacking interactions of the adjacent PSII-LHCII enriched regions of the thylakoid membranes. The self-assembly and dynamics of this highly organized membrane system and the nature of forces acting between the PSII-LHCII macrodomains are not well understood. By using circular dichroism (CD) spectroscopy, small-angle neutron scattering (SANS) and transmission electron microscopy (TEM), we investigated the effects of Hofmeister salts on the organization of pigment-protein complexes and on the ultrastructure of thylakoid membranes. We found that the kosmotropic agent (NH4)2SO4 and the Hofmeister-neutral NaCl, up to 2 M concentrations, hardly affected the macro-organization of the protein complexes and the membrane ultrastructure. In contrast, chaotropic salts, NaClO4, and NaSCN destroyed the mesoscopic structures, the multilamellar organization of the thylakoid membranes and the chiral macrodomains of the protein complexes but without noticeably affecting the short-range, pigment-pigment excitonic interactions. Comparison of the concentration- and time-dependences of SANS, TEM and CD parameters revealed the main steps of the disassembly of grana in the presence of chaotropes. It begins with a rapid diminishment of the long-range periodic order of the grana membranes, apparently due to an increased stacking disorder of the thylakoid membranes, as reflected by SANS experiments. SANS measurements also allowed discrimination between the cationic and anionic effects-in stacking and disorder, respectively. This step is followed by a somewhat slower disorganization of the TEM ultrastructure, due to the gradual loss of stacked membrane pairs. Occurring last is the stepwise decrease and disappearance of the long-range chiral order of the protein complexes, the rate of which was faster in LHCII-deficient membranes. These data are interpreted in terms of a theory, from our laboratory, according to which Hofmeister salts primarily affect the hydrophylic-hydrophobic interactions of proteins, and the stroma-exposed regions of the intrinsic membrane proteins, in particular-pointing to the role of protein-water interface in the stacking interactions of granum thylakoid membranes.

• Klíčová slova
• Hofmeister effect, circular dichroism, granum, protein-water interface, small-angle neutron scattering, thylakoid membranes, ultrastructure,
• Publikační typ
• časopisecké články MeSH

The major light-harvesting system in cyanobacteria, the phycobilisome, is an essential component of the photosynthetic apparatus that regulates the utilization of the natural light source-the Sun. Earlier works revealed that the thylakoid membrane composition and its physical properties might have an important role in antennas docking. Polyunsaturated lipids and xanthophylls are among the most significant modulators of the physical properties of thylakoid membranes. In the nature, the action of these molecules is orchestrated in response to environmental stimuli among which the growth temperature is the most influential. In order to further clarify the significance of thylakoid membrane physical properties for the phycobilisomes assembly (i.e. structural integrity) and their ability to efficiently direct the excitation energy towards the photosynthetic complexes, in this work, we utilize cyanobacterial Synechocystis sp. PCC 6803 mutants deficient in polyunsaturated lipids (AD mutant) and xanthophylls (RO mutant), as well as a strain depleted of both xanthophylls and polyunsaturated lipids (ROAD multiple mutant). For the first time, we discuss the effect of those mutations on the phycobilisomes assembly, integrity and functionality at optimal (30 °C) and moderate low (25 °C) and high (35 °C) temperatures. Our results show that xanthophyll depletion exerts a much stronger effect on both phycobilisome's integrity and the response of cells to growth at suboptimal temperatures than lipid unsaturation level. The strongest effects were observed for the combined ROAD mutant, which exhibited thermally destabilized phycobilisomes and a population of energetically uncoupled phycocyanin units.

• Klíčová slova
• Carotenoids, Cyanobacteria, Excitation energy transfer, Lipid unsaturation, Phycobilisome, Thermal stability,
• MeSH
• fotosyntetické reakční centrum - proteinové komplexy metabolismus   MeSH
• fotosyntéza * MeSH
• fykobilizomy metabolismus   MeSH
• fykokyanin metabolismus   MeSH
• karotenoidy metabolismus   MeSH
• metabolismus lipidů MeSH
• mutace MeSH
• Synechocystis genetika   metabolismus   MeSH
• teplota MeSH
• tylakoidy metabolismus   MeSH
• xanthofyly metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• fotosyntetické reakční centrum - proteinové komplexy MeSH
• fykobilizomy MeSH
• fykokyanin MeSH
• karotenoidy MeSH
• xanthofyly MeSH

The role of non-bilayer lipids and non-lamellar lipid phases in biological membranes is an enigmatic problem of membrane biology. Non-bilayer lipids are present in large amounts in all membranes; in energy-converting membranes they constitute about half of their total lipid content-yet their functional state is a bilayer. In vitro experiments revealed that the functioning of the water-soluble violaxanthin de-epoxidase (VDE) enzyme of plant thylakoids requires the presence of a non-bilayer lipid phase. 31P-NMR spectroscopy has provided evidence on lipid polymorphism in functional thylakoid membranes. Here we reveal reversible pH- and temperature-dependent changes of the lipid-phase behaviour, particularly the flexibility of isotropic non-lamellar phases, of isolated spinach thylakoids. These reorganizations are accompanied by changes in the permeability and thermodynamic parameters of the membranes and appear to control the activity of VDE and the photoprotective mechanism of non-photochemical quenching of chlorophyll-a fluorescence. The data demonstrate, for the first time in native membranes, the modulation of the activity of a water-soluble enzyme by a non-bilayer lipid phase.

• MeSH
• diferenciální skenovací kalorimetrie MeSH
• epoxidové sloučeniny metabolismus   MeSH
• kinetika MeSH
• koncentrace vodíkových iontů MeSH
• lipidové dvojvrstvy chemie   MeSH
• lipidy chemie   MeSH
• magnetická rezonanční spektroskopie MeSH
• oxidoreduktasy metabolismus   MeSH
• rozpustnost MeSH
• Spinacia oleracea metabolismus   MeSH
• světlo MeSH
• teplota MeSH
• tylakoidy chemie   MeSH
• voda chemie   MeSH
• xanthofyly metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• epoxidové sloučeniny MeSH
• lipidové dvojvrstvy MeSH
• lipidy MeSH
• oxidoreduktasy MeSH
• violaxanthin de-epoxidase MeSH Prohlížeč
• violaxanthin MeSH Prohlížeč
• voda MeSH
• xanthofyly MeSH

Trace metal contaminations in natural waters, wetlands, and wastewaters pose serious threats to aquatic ecosystems-mainly via targeting microalgae. In this work, we investigated the effects of toxic amounts of chromium and cadmium ions on the structure and function of the photosynthetic machinery of Chlorella variabilis cells. To halt the propagation of cells, we used high concentrations of Cd and Cr, 50-50 mg L-1, in the forms of CdCl2 x 2.5 H2O and K2Cr2O7, respectively. Both treatments led to similar, about 50% gradual diminishment of the chlorophyll contents of the cells in 48 h, which was, however, accompanied by a small (~10%) but statistically significant enrichment (Cd) and loss (Cr) of ß-carotene. Both Cd and Cr inhibited the activity of photosystem II (PSII)-but with more severe inhibitions with Cr. On the contrary, the PsbA (D1) protein of PSII and the PsbO protein of the oxygen-evolving complex were retained more in Cr-treated cells than in the presence of Cd. These data and the higher susceptibility of P700 redox transients in Cr-treated cells suggest that, unlike with Cd, PSII is not the main target in the photochemical apparatus. These differences at the level of photochemistry also brought about dissimilarities at higher levels of the structural complexity of the photosynthetic apparatus. Circular dichroism (CD) spectroscopy measurements revealed moderate perturbations in the macro-organization of the protein complexes-with more pronounced decline in Cd-treated cells than in the cells with Cr. Also, as reflected by transmission electron microscopy and small-angle neutron scattering, the thylakoid membranes suffered shrinking and were largely fragmented in Cd-treated cells, whereas no changes could be discerned with Cr. The preservation of integrity of membranes in Cr-treated cells was most probably aided by high proportion of the de-epoxidized xanthophylls, which were absent with Cd. It can thus be concluded that beside strong similarities of the toxic effects of Cr and Cd, the response of the photosynthetic machinery of C. variabilis to these two trace metal ions substantially differ from each other-strongly suggesting different inhibitory and protective mechanisms following the primary toxic events.

• Klíčová slova
• P700, cadmium, chromium, circular dichroism, electron microscopy, green alga, photosystem II, small-angle neutron scattering,
• Publikační typ
• časopisecké články MeSH

γ-Tubulin is associated with microtubule nucleation, but evidence is accumulating in eukaryotes that it also functions in nuclear processes and in cell division control independently of its canonical role. We found that in Arabidopsis thaliana, γ-tubulin interacts specifically with E2FA, E2FB, and E2FC transcription factors both in vitro and in vivo. The interaction of γ-tubulin with the E2Fs is not reduced in the presence of their dimerization partners (DPs) and, in agreement, we found that γ-tubulin interaction with E2Fs does not require the dimerization domain. γ-Tubulin associates with the promoters of E2F-regulated cell cycle genes in an E2F-dependent manner, probably in complex with the E2F-DP heterodimer. The up-regulation of E2F target genes PCNA, ORC2, CDKB1;1, and CCS52A under γ-tubulin silencing suggests a repressive function for γ-tubulin at G1/S and G2/M transitions, and the endocycle, which is consistent with an excess of cell division in some cells and enhanced endoreduplication in others in the shoot and young leaves of γ-tubulin RNAi plants. Altogether, our data show ternary interaction of γ-tubulin with the E2F-DP heterodimer and suggest a repressive role for γ-tubulin with E2Fs in controlling mitotic activity and endoreduplication during plant development.

• Klíčová slova
• Arabidopsis, E2F transcription factors, endoreduplication, gene expression, proliferation, γ-tubulin,
• MeSH
• Arabidopsis * genetika   metabolismus   MeSH
• proteiny buněčného cyklu MeSH
• proteiny huseníčku * genetika   metabolismus   MeSH
• regulace genové exprese u rostlin MeSH
• transkripční faktory E2F * genetika   metabolismus   MeSH
• tubulin * genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• E2Fb protein, Arabidopsis MeSH Prohlížeč
• proteiny buněčného cyklu MeSH
• proteiny huseníčku * MeSH
• transkripční faktory E2F * MeSH
• tubulin * MeSH

The rapidly proliferating cells in plant meristems must be protected from genome damage. Here, we show that the regulatory role of the Arabidopsis RETINOBLASTOMA RELATED (RBR) in cell proliferation can be separated from a novel function in safeguarding genome integrity. Upon DNA damage, RBR and its binding partner E2FA are recruited to heterochromatic γH2AX-labelled DNA damage foci in an ATM- and ATR-dependent manner. These γH2AX-labelled DNA lesions are more dispersedly occupied by the conserved repair protein, AtBRCA1, which can also co-localise with RBR foci. RBR and AtBRCA1 physically interact in vitro and in planta Genetic interaction between the RBR-silenced amiRBR and Atbrca1 mutants suggests that RBR and AtBRCA1 may function together in maintaining genome integrity. Together with E2FA, RBR is directly involved in the transcriptional DNA damage response as well as in the cell death pathway that is independent of SOG1, the plant functional analogue of p53. Thus, plant homologs and analogues of major mammalian tumour suppressor proteins form a regulatory network that coordinates cell proliferation with cell and genome integrity.

• Klíčová slova
• Arabidopsis, RETINOBLASTOMA RELATED, BRCA1, DNA damage response, E2FA,
• MeSH
• Arabidopsis genetika   MeSH
• ATM protein metabolismus   MeSH
• DNA rostlinná metabolismus   MeSH
• kontrolní body buněčného cyklu * MeSH
• oprava DNA * MeSH
• poškození DNA * MeSH
• proteiny huseníčku metabolismus   MeSH
• regulace genové exprese u rostlin * MeSH
• transkripční faktory E2F metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH
• Názvy látek
• ATM protein, Arabidopsis MeSH Prohlížeč
• ATM protein MeSH
• ATR protein, Arabidopsis MeSH Prohlížeč
• DNA rostlinná MeSH
• E2Fa protein, Arabidopsis MeSH Prohlížeč
• proteiny huseníčku MeSH
• RBR1 protein, Arabidopsis MeSH Prohlížeč
• transkripční faktory E2F MeSH