Kazimierz Strzałka

Physiological responses of Lemna trisulca L. (duckweed) to cadmium and copper bioaccumulation

Abstract Aquatic plants are known to accumulate and bioconcentrate heavy metals. In this study, several physiological responses of aquatic vascular plant Lemna trisulca L. to elevated concentrations of cadmium (up to 10 mM) and copper (up to 50 μM) were investigated. It was found that Lemna fronds were able to accumulate both cadmium and copper, but Cu-treated material showed pronounced toxic symptoms at concentrations 1000-fold lower in comparison to Cd. Lemna trisulca could tolerate elevated levels of Cd, i.e. up to 10 mM, without significant changes in photosynthetic pigments concentration. On the contrary, Cu in concentrations 25 and 50 μM promoted significant pigment degradation. The main processes affected by Cd in Lemna fronds were total gas exchange and net photosynthesis. On the contrary, the inhibition of total gas exchange and net photosynthesis caused by Cu (2–50 μM) correlated with Chl a and carotenoid concentrations decrease as well as with the decay of fluorescence from PS II. Also, an increasing impact of respiration in total oxygen exchange was observed after treatment of Lemna with increasing Cd concentrations (up to 5 mM) and with Cu in concentration range between 2 and 50 μM. In Cd-treated fronds, a dose-dependent accumulation of two polypeptides with apparent molecular weights 18 and 10 kDa, respectively as well as the appearance of two smaller polypeptides (apparent molecular weights 8 and 7 kDa) was observed in SDS-PAGE. The nature of these polypeptides remains to be determined. On the contrary, in Cu-treated fronds neither accumulation of existing proteins nor appearance of any extra protein was observed.

Mechanism and regulation of the violaxanthin cycle: the role of antenna proteins and membrane lipids.

The violaxanthin cycle describes the reversible conversion of violaxanthin to zeaxanthin via the intermediate antheraxanthin. This light-dependent xanthophyll conversion is essential for the adaptation of plants and algae to different light conditions and allows a reversible switch of photosynthetic light-harvesting complexes between a light-harvesting state under low light and a dissipative state under high light. The photoprotective functions of zeaxanthin have been intensively studied during the last decade, but much less attention has been directed to the mechanism and regulation of xanthophyll conversion. In this review, an overview is given on recent progress in the understanding of the role of (i) xanthophyll binding by antenna proteins and of (ii) the lipid properties of the thylakoid membrane in the regulation of xanthophyll conversion. The consequences of these findings for the mechanism and regulation of xanthophyll conversion in the thylakoid membrane will be discussed.

Heavy metal-induced oxidative damage, defense reactions, and detoxification mechanisms in plants

Heavy metal (HMs) contamination is widespread globally due to anthropogenic, technogenic, and geogenic activities. The HMs exposure could lead to multiple toxic effects in plants by inducing reactive oxygen species (ROS), which inhibit most cellular processes at various levels of metabolism. ROS being highly unstable could play dual role (1) damaging cellular components and (2) act as an important secondary messenger for inducing plant defense system. Cells are equipped with enzymatic and non-enzymatic defense mechanisms to counteract this damage. Some are constitutive and others that are activated only when a stress-specific signal is perceived. Enzymatic scavengers of ROS include superoxide dismutase, catalase, glutathione reductase, and peroxidase, while non-enzymatic antioxidants are glutathione, ascorbic acid, α-tocopherol, flavonoids, anthocyanins, carotenoids, and organic acids. The intracellular and extracellular chelation mechanisms of HMs are associated with organic acids such as citric, malic and oxalic acid, etc. The important mechanism of detoxification includes metal complexation with glutathione, amino acids, synthesis of phytochelatins and sequestration into the vacuoles. Excessive stresses induce a cascade, MAPK (mitogen-activated protein kinase) pathway and synthesis of metal-detoxifying ligands. Metal detoxification through MAPK cascade and synthesis of metal-detoxifying ligands will be of considerable interest in the field of plant biotechnology. Further, the photoprotective roles of pigments of xanthophylls cycle under HMs stress were also discussed.

Phosphatidylglycerol Is Essential for Oligomerization of Photosystem I Reaction Center

Our earlier studies with the pgsA mutant of Synechocystis PCC6803 demonstrated the important role of phosphatidylglycerol (PG) in PSII dimer formation and in electron transport between the primary and secondary electron-accepting plastoquinones of PSII. Using a long-term depletion of PG from pgsA mutant cells, we could induce a decrease not only in PSII but also in PSI activity. Simultaneously with the decrease in PSI activity, dramatic structural changes of the PSI complex were detected. A 21-d PG depletion resulted in the degradation of PSI trimers and concomitant accumulation of monomer PSI. The analyses of PSI particles isolated by MonoQ chromatography showed that, following the 21-d depletion, PSI trimers were no longer detectable in the thylakoid membranes. Immunoblot analyses revealed that the PSI monomers accumulating in the PG-depleted mutant cells do not contain PsaL, the protein subunit thought to be responsible for the trimer formation. Nevertheless, the trimeric structure of PSI reaction center could be restored by readdition of PG, even in the presence of the protein synthesis inhibitor lincomycin, indicating that free PsaL was present in thylakoid membranes following the 21-d PG depletion. Our data suggest an indispensable role for PG in the PsaL-mediated assembly of the PSI reaction center.

Redox Changes of Cytochrome b 559 in the Presence of Plastoquinones

We have found that short chain plastoquinones effectively stimulated photoreduction of the low potential form of cytochrome b 559 and were also active in dark oxidation of this cytochrome under anaerobic conditions in Triton X-100-solubilized photosystem II (PSII) particles. It is also shown that molecular oxygen competes considerably with the prenylquinones in cytochrome b 559 oxidation under aerobic conditions, indicating that both molecular oxygen and plastoquinones could be electron acceptors from cytochromeb 559 in PSII preparations. α-Tocopherol quinone was not active in the stimulation of cytochrome photoreduction but efficiently oxidized it in the dark. Both the observed photoreduction and dark oxidation of the cytochrome were not sensitive to 3-(3,4-dichlorophenyl)-1,1-dimethylurea. It was concluded that both quinone-binding sites responsible for the redox changes of cytochrome b 559 are different from either the QA or QB site in PSII and represent new quinone-binding sites in PSII.

Carotenoids and Environmental Stress in Plants: Significance of Carotenoid-Mediated Modulation of Membrane Physical Properties

Carotenoids, apart of their antenna function in photosynthesis, play an important role in the mechanisms protecting the photosynthetic apparatus against various harmful environmental factors. They protect plants against overexcitation in strong light and dissipate the excess of absorbed energy, they scavenge reactive oxygen species formed during photooxidative stress and moderate the effect of extreme temperatures. One of the important factors involved in the protective action of carotenoids is their influence on the molecular dynamics of membranes. To obtain complex information about interactions between carotenoids and lipids in a membrane, different techniques were used. In this review, the data resulting from EPR–spin label spectrometry, 31P- and 13C-NMR, differential scanning calorimetry, and computer simulation of the membrane molecular dynamics are presented. The effects of selected, structurally different carotenoid species on various physical parameters of model and natural membranes are described and their relevance to protection against some environmental stresses are discussed.

Dark reoxidation of the plastoquinone-pool is mediated by the low-potential form of cytochrome b-559 in spinach thylakoids

We have found that in isolated spinach thylakoids, plastoquinone-pool (PQ-pool), after its photoreduction, undergoes dark-reoxidation with the half-time of τ1/2 = 43 ± 3 s. To explain the observed rates of PQ-pool reoxidation, a nonenzymatic plastoquinol (PQH2) autoxidation under molecular oxygen and an enzymatic oxidation by the low-potential form of cytochrome b-559 (cyt. b-559LP), as the postulated PQ-oxidase in chlororespiration, were investigated. It was found that the autoxidation rate of PQH2 in organic solvents and liposomes was too low to account for the observed oxidation rate of PQH2 in thylakoids. The rate of cyt. b-559LP autoxidation in isolated Photosystem II was found to be similar (τ1/2 = 26 ± 5 s) to that of the PQ-pool. This suggests that the LP form of cyt. b-559 is probably responsible for the PQ-oxidase activity observed during chlororespiration.

Effect of β-carotene on structural and dynamic properties of model phosphatidylcholine membranes. I. An EPR spin label study

The influence of beta-carotene on structural and dynamic properties of model membranes (multilamellar liposomes) prepared of dipalmitoylphosphatidylcholine was investigated. It was found that beta-carotene: (1) decreases order within crystalline state of the membrane; the effect of beta-carotene was more pronounced than in the case of the polar carotenoid, lutein, as revealed by means of spin label EPR; (2) increases penetration, stronger than lutein, of apolar molecules into the membrane as indicated by greater partition coefficient of 5-doxyldecane; (3) increases correlation times tau B tau C stronger than lutein. In all cases the effect of beta-carotene on a membrane was more pronounced at crystalline state than at fluid state. On this basis a hypothesis is proposed that beta-carotene plays a physiological function in the fluidization of chloroplast membranes in a chilling stress to the photosynthetic apparatus.

The xanthophyll cycle - molecular mechanism and physiological significance

The light-dependent, cyclic changes of xanthophyll pigments: violaxanthin, antheraxanthin and zeaxanthin, called the xanthophyll cycle, have been known for about fifty years. This process was characterised for higher plants, several fern and moss species and in some algal groups. Two enzymes, violaxanthin de-epoxidase (VDE) and zeaxanthin epoxidase (ZE), belonging to the lipocalin protein family, are engaged in the xanthophyll cycle. VDE requires for its activity ascorbic acid and reversed hexagonal structure formed by monogalactosyldiacylglycerol. ZE, postulated to be a flavoprotein, has not been purified yet and it is known from its gene sequence only. Zeaxanthin epoxidation is dependent on the reducing power of NADPH and presence of additional proteins.The xanthophyll cycle is postulated to play a role in many important physiological processes. Zeaxanthin, formed from violaxanthin under high light conditions, is thought to be a main photoprotector in autotrophic cells due to its ability to dissipate excess of absorbed light energy that can be measured as a non-photochemical quenching. In addition the zeaxanthin formation is important in protection of the thylakoid membranes against lipid peroxidation. Other postulated functions of the xanthophyll cycle, which include regulation of membrane physical properties, blue light reception and regulation of abscisic acid synthesis, are also discussed.

Plastoquinol and α-tocopherol quinol are more active than ubiquinol and α-tocopherol in inhibition of lipid peroxidation

Abstract Comparative studies of antioxidant activities of such natural prenyllipids as plastoquinol-9 (PQH2-9), α-tocopherol quinol (α-TQH2), ubiquinol-10 (UQH2-10) and α-tocopherol (α-T) in egg yolk lecithin liposomes have been performed. The investigated compounds showed oxidation under molecular oxygen in the order UQH2-10>α-TQH2>PQH2-9>>α-T. The corresponding second order rate constants have been determined in Tris buffer (pH=6.5) and were 0.413, 0.268, 0.154 and 0.022 M−1/s, respectively. The inhibition order of Fe2+-H2O2 -induced lipid peroxidation, corrected for the amount of prenyllipids oxidized during the initiation period, was α-TQH2>PQH2-9>α-T>UQH2-10 for 5 mol% of the antioxidants content in liposomes. The radicals formed in the initiation phase of the reaction caused oxidation of 27.5–33% α-T, 40–64% UQH2-10, 42–85% PQH2-9 and 43–80% α-TQH2, depending on the antioxidant concentration in liposomes (5–1 mol%, respectively) which reflects approximately their reactivity against radicals derived from the Fenton reaction. The antioxidant activity of the investigated prenylquinols, in relation to the activity of α-T, in natural membranes is discussed.