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Sample records for chloroplast thylakoid membrane

  1. The molecular architecture of the chloroplast thylakoid membrane

    Energy Technology Data Exchange (ETDEWEB)

    Stefansson, H.

    1996-08-01

    Non-detergent procedure for isolation of sub-thylakoid vesicle populations derived from different structural domains of the chloroplast thylakoid membrane has been developed. Sub-thylakoid vesicles representing the grana, grana core, stroma lamellae, and the grana margins have been isolated and their protein composition has been investigated. Furthermore a novel non-detergent procedure for investigating the pigment composition of photosynthetic complexes located in the different structural domains has been developed. This procedure circumvents selective extractions, an perturbing effect often combined with detergent isolations of membrane bound protein complexes. The fractionation experiments show that the NADPH dehydrogenase, suggested to operate as NADPH or ferredoxin-plastoquinone oxidoreductase in cyclic electron transport around photosystem I, is stoichiometrically depleted on photosystem I basis in the grana domain. The fractionation studies are consistent with the model of the thylakoid membrane where the photosystems in the grana are operating in a linear electron transport whereas the site of cyclic electron transport is in the stroma lamellae. It is suggested that partial destacking of grana, as a result of light-induced protein phosphorylation, may promote the exposure of the granal photosystem I centers to the chloroplast stroma and thereby enhance their participation in cyclic electron transport activity. 146 refs, 18 figs

  2. Role of membrane glycerolipids in photosynthesis, thylakoid biogenesis and chloroplast development.

    Science.gov (United States)

    Kobayashi, Koichi

    2016-07-01

    The lipid bilayer of the thylakoid membrane in plant chloroplasts and cyanobacterial cells is predominantly composed of four unique lipid classes; monogalactosyldiacylglycerol (MGDG), digalactosyldiacylglycerol (DGDG), sulfoquinovosyldiacylglycerol (SQDG) and phosphatidylglycerol (PG). MGDG and DGDG are uncharged galactolipids that constitute the bulk of thylakoid membrane lipids and provide a lipid bilayer matrix for photosynthetic complexes as the main constituents. The glycolipid SQDG and phospholipid PG are anionic lipids with a negative charge on their head groups. SQDG and PG substitute for each other to maintain the amount of total anionic lipids in the thylakoid membrane, with PG having indispensable functions in photosynthesis. In addition to biochemical studies, extensive analyses of mutants deficient in thylakoid lipids have revealed important roles of these lipids in photosynthesis and thylakoid membrane biogenesis. Moreover, recent studies of Arabidopsis thaliana suggest that thylakoid lipid biosynthesis triggers the expression of photosynthesis-associated genes in both the nucleus and plastids and activates the formation of photosynthetic machineries and chloroplast development. Meanwhile, galactolipid biosynthesis is regulated in response to chloroplast functionality and lipid metabolism at transcriptional and post-translational levels. This review summarizes the roles of thylakoid lipids with their biosynthetic pathways in plants and discusses the coordinated regulation of thylakoid lipid biosynthesis with the development of photosynthetic machinery during chloroplast biogenesis.

  3. Quantitative local photosynthetic flux measurements at isolated chloroplasts and thylakoid membranes using scanning electrochemical microscopy (SECM).

    Science.gov (United States)

    McKelvey, Kim; Martin, Sophie; Robinson, Colin; Unwin, Patrick R

    2013-07-01

    Scanning electrochemical microscopy (SECM) offers a fast and quantitative method to measure local fluxes within photosynthesis. In particular, we have measured the flux of oxygen and ferrocyanide (Fe(CN)6(4-)), from the artificial electron acceptor ferricyanide (Fe(CN)6(3-)), using a stationary ultramicroelectrode at chloroplasts and thylakoid membranes (sourced from chloroplasts). Oxygen generation at films of chloroplasts and thylakoid membranes was detected directly during photosynthesis, but in the case of thylakoid membranes, this switched to sustained oxygen consumption at longer illumination times. An initial oxygen concentration spike was detected over both chloroplast and thylakoid membrane films, and the kinetics of the oxygen generation were extracted by fitting the experimental data to a finite element method (FEM) simulation. In contrast to previous work, the oxygen generation spike was attributed to the limited size of the plastoquinone pool, a key component in the linear electron transport pathway and a contributing factor in photoinhibition. Finally, the mobile nature of the SECM probe, and its high spatial resolution, also allowed us to detect ferrocyanide produced from a single thylakoid membrane. These results further demonstrate the power of SECM for localized flux measurements in biological processes, in this case photosynthesis, and that the high time resolution, combined with FEM simulations, allows the elucidation of quantitative kinetic information.

  4. Protons, the thylakoid membrane, and the chloroplast ATP synthase.

    Science.gov (United States)

    Junge, W

    1989-01-01

    According to the chemiosmotic theory, proton pumps and ATP synthases are coupled by lateral proton flow through aqueous phases. Three long-standing challenges to this concept, all of which have been loosely subsumed under 'localized coupling' in the literature, were examined in the light of experiments carried out with thylakoids: (1) Nearest neighbor interaction between pumps and ATP synthases. Considering the large distances between photosystem II and CFoCF1, in stacked thylakoids this is a priori absent. (2) Enhanced proton diffusion along the surface of the membrane. This could not be substantiated for the outer side of the thylakoid membrane. Even for the interface between pure lipid and water, two laboratories have reported the absence of enhanced diffusion. (3) Localized proton ducts in the membrane. Intramembrane domains that can transiently trap protons do exist in thylakoid membranes, but because of their limited storage capacity for protons, they probably do not matter for photophosphorylation under continuous light. Seemingly in favor of localized proton ducts is the failure of a supposedly permeant buffer to enhance the onset lag of photophosphorylation. However, it was found that failure of some buffers and the ability of others in this respect were correlated with their failure/ability to quench pH transients in the thylakoid lumen, as predicted by the chemiosmotic theory. It was shown that the chemiosmotic concept is a fair approximation, even for narrow aqueous phases, as in stacked thylakoids. These are approximately isopotential, and protons are taken in by the ATP synthase straight from the lumen. The molecular mechanism by which F0F1 ATPases couple proton flow to ATP synthesis is still unknown. The threefold structural symmetry of the headpiece that, probably, finds a corollary in the channel portion of these enzymes appeals to the common wisdom that structural symmetry causes functional symmetry. "Rotation catalysis" has been proposed. It is

  5. Production of superoxide in chloroplast thylakoid membranes ESR study with cyclic hydroxylamines of different lipophilicity.

    Science.gov (United States)

    Kozuleva, Marina; Klenina, Irina; Proskuryakov, Ivan; Kirilyuk, Igor; Ivanov, Boris

    2011-04-06

    Accumulation of nitroxide radicals, DCP· or TMT·, under illumination of a thylakoid suspension containing either hydrophilic, DCP-H, or lipophilic, TMT-H, cyclic hydroxylamines that have high rate constants of the reaction with superoxide radicals, was measured using ESR. A slower accumulation of TMT· in contrast with DCP· accumulation was explained by re-reduction of TMT· by the carriers of the photosynthetic electron transport chain within the membrane. Superoxide dismutase suppressed TMT· accumulation to a lesser extent than DCP· accumulation. The data are interpreted as evidencing the production of intramembrane superoxide in thylakoids.

  6. Visualizing structural dynamics of thylakoid membranes

    Science.gov (United States)

    Iwai, Masakazu; Yokono, Makio; Nakano, Akihiko

    2014-01-01

    To optimize photosynthesis, light-harvesting antenna proteins regulate light energy dissipation and redistribution in chloroplast thylakoid membranes, which involve dynamic protein reorganization of photosystems I and II. However, direct evidence for such protein reorganization has not been visualized in live cells. Here we demonstrate structural dynamics of thylakoid membranes by live cell imaging in combination with deconvolution. We observed chlorophyll fluorescence in the antibiotics-induced macrochloroplast in the moss Physcomitrella patens. The three-dimensional reconstruction uncovered the fine thylakoid membrane structure in live cells. The time-lapse imaging shows that the entire thylakoid membrane network is structurally stable, but the individual thylakoid membrane structure is flexible in vivo. Our observation indicates that grana serve as a framework to maintain structural integrity of the entire thylakoid membrane network. Both the structural stability and flexibility of thylakoid membranes would be essential for dynamic protein reorganization under fluctuating light environments. PMID:24442007

  7. 植物叶绿体类囊体膜及膜蛋白研究进展%Progress in chloroplast thylakoid membrane and membrane proteins

    Institute of Scientific and Technical Information of China (English)

    胡锋; 黄俊丽; 秦峰; 岳彩黎; 王贵学

    2011-01-01

    In plants and eukaryotic algae, photosynthesis takes place in chloroplasts. Light reaction occurs in the thylakoid membranes. Thylakoid membranes contain integral and peripheral membrane protein complexes,including the pigments that absorb light energy, which form the photosystems. Much attention has been focused on the thylakoid membranes because of its significance in photosynthesis. Analysis of thylakoid membranes will benefit the study of photosynthetic mechanism. The present review summarizes the three-dimensional conformation,membrane protein composition and function of thylakoid membranes.%叶绿体是植物和真核藻类进行光合作用的场所.存在于叶绿体类囊体膜上的蛋白质复合物含有光反应所需的光合色素和电子传递链组分,在光合作用过程中,光化学反应发生在类囊体膜上.因此,类囊体膜是光能向化学能转化的主要场所,因而也一直是光合作用研究的热点.叶绿体类囊体膜的深入研究可以促进光合作用的分子机理研究.该文就叶绿体类囊体膜的三维构象及类囊体膜蛋白的组成和功能研究进行了综述.

  8. The effect of microgravity on proton permeability of thylakoid membranes and contribution of II and I photosystems in photosynthetic electron transport in pea chloroplasts.

    Science.gov (United States)

    Zolotareva, E K; Onoiko, E B; Sytnik, S K; Podorvanov, V V

    1999-07-01

    According to a number investigations microgravity conditions affect membrane apparatus of photosynthesis in cells of higher plants and alga [for review, see Kordyum et al., 1994; Kordyum, 1997]. (see for review). Chloroplasts of space-grown pea plants showed disintegration of grana, shrinkage of the membrane constituting the grana stacks and other structural perturbance of the photosynthetic membranes. However there have been no studies on the effect of microgravity on proton permeability of thylakoid membranes and closely connected with this parameter their photochemical characteristics. The aim of the study is investigation of microgravity effects on protonic permeability of photosynthetic membrane and contribution of photosystem II (PSII) and photosystem I (PSI) in electron transfer from water to potassium ferrycianide (FeCy) in isolated pea chloroplasts. Pea.

  9. Changes in antenna sizes of photosystems during state transitions in granal and stroma-exposed thylakoid membrane of intact chloroplasts in Arabidopsis mesophyll protoplasts.

    Science.gov (United States)

    Kim, Eunchul; Ahn, Tae Kyu; Kumazaki, Shigeichi

    2015-04-01

    In chloroplasts of plants and algae, state transition is an important regulatory mechanism to maintain the excitation balance between PSI and PSII in the thylakoid membrane. Light-harvesting complex II (LHCII) plays a key role as the regulated energy distributor between PSI and PSII. It is widely accepted that LHCII, which is bound to PSII localized mainly in the granal thylakoid, migrates to bind with PSI localized mainly in the stroma-exposed thylakoid under preferential excitation of PSII. The phenomena have been extensively characterized by many methods. However, the exchange of LHCII between PSII and PSI has not been directly observed in vivo at physiological temperatures. Herein we applied fluorescence spectromicroscopy to Arabidopsis mesophyll protoplasts in order to observe in vivo changes in fluorescence spectra of granal and stromal thylakoid regions during the state transition. The microscopic fluorescence spectra obtained from a few sections with different depths were decomposed into PSI and PSII spectra and self-absorption effects were removed. We were able to determine amplitude changes of PSI and PSII in fluorescence spectra solely due to state transition. Subdomain analysis of granal and stromal thylakoid regions clarified variant behaviors in the different regions.

  10. Small-angle neutron scattering study of the ultrastructure of chloroplast thylakoid membranes - Periodicity and structural flexibility of the stroma lamellae

    DEFF Research Database (Denmark)

    Posselt, Dorthe; Nagy, Gergely; Kirkensgaard, Jacob J. K.;

    2012-01-01

    The multilamellar organization of freshly isolated spinach and pea chloroplast thylakoid membranes was studied using small-angle neutron scattering. A broad peak at similar to 0.02 angstrom(-1) is ascribed to diffraction from domains of ordered, unappressed stroma lamellae, revealing a repeat...... distance of 294 angstrom +/- 7 angstrom in spinach and 345 angstrom +/- 11 angstrom in pea. The peak position and hence the repeat distance of stroma lamellae is strongly dependent on the osmolarity and the ionic strength of the suspension medium, as demonstrated by varying the sorbitol and the Mg......++-concentration in the sample. For pea thylakoid membranes, we show that the repeat distance decreases when illuminating the sample with white light, in accordance with our earlier results on spinach, also regarding the observation that addition of an uncoupler prohibits the light-induced structural changes, a strong...

  11. Proteins affecting thylakoid morphology - the key to understanding vesicle transport in chloroplasts?

    Science.gov (United States)

    Lindquist, Emelie; Aronsson, Henrik

    2014-01-01

    We recently showed that a Rab protein, CPRabA5e (CP = chloroplast localized), is located in chloroplasts of Arabidopsis thaliana where it is involved in various processes, such as thylakoid biogenesis and vesicle transport. Using a yeast two-hybrid method, CPRabA5e was shown to interact with a number of chloroplast proteins, including the CURVATURE THYLAKOID 1A (CURT1A) protein and the light-harvesting chlorophyll a/b binding protein (LHCB1.5). CURT1A has recently been shown to modify thylakoid architecture by inducing membrane curvature in grana, whereas LHCB1.5 is a protein of PSII (Photosystem II) facilitating light capture. LHCB1.5 is imported to chloroplasts and transported to thylakoid membranes using the post-translational Signal Recognition Particle (SRP) pathway. With this information as starting point, we here discuss their subsequent protein-protein interactions, given by the literature and Interactome 3D. CURT1A itself and several of the proteins interacting with CURT1A and LHCB1.5 have relations to vesicle transport and thylakoid morphology, which are also characteristics of cprabA5e mutants. This highlights the previous hypothesis of an alternative thylakoid targeting pathway for LHC proteins using vesicles, in addition to the SRP pathway.

  12. Genome-wide analysis of thylakoid-bound ribosomes in maize reveals principles of cotranslational targeting to the thylakoid membrane.

    Science.gov (United States)

    Zoschke, Reimo; Barkan, Alice

    2015-03-31

    Chloroplast genomes encode ∼ 37 proteins that integrate into the thylakoid membrane. The mechanisms that target these proteins to the membrane are largely unexplored. We used ribosome profiling to provide a comprehensive, high-resolution map of ribosome positions on chloroplast mRNAs in separated membrane and soluble fractions in maize seedlings. The results show that translation invariably initiates off the thylakoid membrane and that ribosomes synthesizing a subset of membrane proteins subsequently become attached to the membrane in a nuclease-resistant fashion. The transition from soluble to membrane-attached ribosomes occurs shortly after the first transmembrane segment in the nascent peptide has emerged from the ribosome. Membrane proteins whose translation terminates before emergence of a transmembrane segment are translated in the stroma and targeted to the membrane posttranslationally. These results indicate that the first transmembrane segment generally comprises the signal that links ribosomes to thylakoid membranes for cotranslational integration. The sole exception is cytochrome f, whose cleavable N-terminal cpSecA-dependent signal sequence engages the thylakoid membrane cotranslationally. The distinct behavior of ribosomes synthesizing the inner envelope protein CemA indicates that sorting signals for the thylakoid and envelope membranes are distinguished cotranslationally. In addition, the fractionation behavior of ribosomes in polycistronic transcription units encoding both membrane and soluble proteins adds to the evidence that the removal of upstream ORFs by RNA processing is not typically required for the translation of internal genes in polycistronic chloroplast mRNAs.

  13. Unique thylakoid membrane architecture of a unicellular N2-fixing cyanobacterium revealed by electron tomography.

    Science.gov (United States)

    Liberton, Michelle; Austin, Jotham R; Berg, R Howard; Pakrasi, Himadri B

    2011-04-01

    Cyanobacteria, descendants of the endosymbiont that gave rise to modern-day chloroplasts, are vital contributors to global biological energy conversion processes. A thorough understanding of the physiology of cyanobacteria requires detailed knowledge of these organisms at the level of cellular architecture and organization. In these prokaryotes, the large membrane protein complexes of the photosynthetic and respiratory electron transport chains function in the intracellular thylakoid membranes. Like plants, the architecture of the thylakoid membranes in cyanobacteria has direct impact on cellular bioenergetics, protein transport, and molecular trafficking. However, whole-cell thylakoid organization in cyanobacteria is not well understood. Here we present, by using electron tomography, an in-depth analysis of the architecture of the thylakoid membranes in a unicellular cyanobacterium, Cyanothece sp. ATCC 51142. Based on the results of three-dimensional tomographic reconstructions of near-entire cells, we determined that the thylakoids in Cyanothece 51142 form a dense and complex network that extends throughout the entire cell. This thylakoid membrane network is formed from the branching and splitting of membranes and encloses a single lumenal space. The entire thylakoid network spirals as a peripheral ring of membranes around the cell, an organization that has not previously been described in a cyanobacterium. Within the thylakoid membrane network are areas of quasi-helical arrangement with similarities to the thylakoid membrane system in chloroplasts. This cyanobacterial thylakoid arrangement is an efficient means of packing a large volume of membranes in the cell while optimizing intracellular transport and trafficking.

  14. Unique Thylakoid Membrane Architecture of a Unicellular N2-Fixing Cyanobacterium Revealed by Electron Tomography

    Energy Technology Data Exchange (ETDEWEB)

    Liberton, Michelle L.; Austin, Jotham R.; Berg, R. H.; Pakrasi, Himadri B.

    2011-04-01

    Cyanobacteria, descendants of the endosymbiont that gave rise to modern-day chloroplasts, are vital contributors to global biological energy conversion processes. A thorough understanding of the physiology of cyanobacteria requires detailed knowledge of these organisms at the level of cellular architecture and organization. In these prokaryotes, the large membrane protein complexes of the photosynthetic and respiratory electron transport chains function in the intracellular thylakoid membranes. Like plants, the architecture of the thylakoid membranes in cyanobacteria has direct impact on cellular bioenergetics, protein transport, and molecular trafficking. However, whole-cell thylakoid organization in cyanobacteria is not well understood. Here we present, by using electron tomography, an in-depth analysis of the architecture of the thylakoid membranes in a unicellular cyanobacterium, Cyanothece sp. ATCC 51142. Based on the results of three-dimensional tomographic reconstructions of near-entire cells, we determined that the thylakoids in Cyanothece 51142 form a dense and complex network that extends throughout the entire cell. This thylakoid membrane network is formed from the branching and splitting of membranes and encloses a single lumenal space. The entire thylakoid network spirals as a peripheral ring of membranes around the cell, an organization that has not previously been described in a cyanobacterium. Within the thylakoid membrane network are areas of quasi-helical arrangement with similarities to the thylakoid membrane system in chloroplasts. This cyanobacterial thylakoid arrangement is an efficient means of packing a large volume of membranes in the cell while optimizing intracellular transport and trafficking.

  15. Unique thylakoid membrane architecture of a unicellular N2-fixing cyanobacterium revealed by electron tomography

    Energy Technology Data Exchange (ETDEWEB)

    Liberton, Michelle; Austin II, Jotham R; Berg, R. Howard; Pakrasi, Himadri B

    2011-04-01

    Cyanobacteria, descendants of the endosymbiont that gave rise to modern-day chloroplasts, are vital contributors to global biological energy conversion processes. A thorough understanding of the physiology of cyanobacteria requires detailed knowledge of these organisms at the level of cellular architecture and organization. In these prokaryotes, the large membrane protein complexes of the photosynthetic and respiratory electron transport chains function in the intracellular thylakoid membranes. Like plants, the architecture of the thylakoid membranes in cyanobacteria has direct impact on cellular bioenergetics, protein transport, and molecular trafficking. However, whole-cell thylakoid organization in cyanobacteria is not well understood. Here we present, by using electron tomography, an in-depth analysis of the architecture of the thylakoid membranes in a unicellular cyanobacterium, Cyanothece sp. ATCC 51142. Based on the results of three-dimensional tomographic reconstructions of near-entire cells, we determined that the thylakoids in Cyanothece 51142 form a dense and complex network that extends throughout the entire cell. This thylakoid membrane network is formed from the branching and splitting of membranes and encloses a single lumenal space. The entire thylakoid network spirals as a peripheral ring of membranes around the cell, an organization that has not previously been described in a cyanobacterium. Within the thylakoid membrane network are areas of quasi-helical arrangement with similarities to the thylakoid membrane system in chloroplasts. This cyanobacterial thylakoid arrangement is an efficient means of packing a large volume of membranes in the cell while optimizing intracellular transport and trafficking.

  16. Inhibition by penem of processing peptidases from cyanobacteria and chloroplast thylakoids.

    Science.gov (United States)

    Barbrook, A C; Packer, J C; Howe, C J

    1996-12-02

    Proteins targeted to the thylakoid lumen of plants and cyanobacteria and the periplasmic space of cyanobacteria are synthesised with N-terminal presequences which are removed following translocation across the membrane. These presequences are thought to direct translocation of the preprotein by a sec-type pathway. Detergent extracts of cyanobacterial and chloroplast membranes contain enzymes which are capable of processing precursors to the mature size. We show that the processing of a range of precursors by both cyanobacterial and chloroplast enzymes is inhibited by the penem SB216357. This is the first report of an inhibitor of these enzymes and indicates that they are type 1 signal peptidases.

  17. Superoxide production in aprotic interior of chloroplast thylakoids.

    Science.gov (United States)

    Takahashi, M; Asada, K

    1988-12-01

    The site of superoxide production in spinach thylakoids was found to be the aprotic interior of the thylakoid membranes near the P700 chlorophyll a protein at the reaction center of photosystem I complexes. This conclusion was drawn from the following findings. (i) Cytochrome c reduction by illuminated thylakoids, which was confirmed to be superoxide dependent by the failure of this reaction to occur in anaerobiosis, was completely inhibited by a dibutyl catechol, but partially inhibited by a hydrophilic disulfonated derivative. (ii) P700 chlorophyll a proteins were preferentially iodinated by lactoperoxidase by the use of hydrogen peroxide that was derived from the disproportionation of superoxides in illuminated thylakoids. (iii) Hydrogen peroxide production and oxygen uptake were induced by ammonium chloride, a proton conductor that can permeate through thylakoid membranes, but whole superoxide in the bulk solution was oxidized back to molecular oxygen by cytochrome c. The effective concentration of ammonium chloride decreased to one-sixtieth of the original, when an ammonium ion ionophore, nonactin, was added. Thus, the weak acid allowed superoxide to yield hydrogen peroxide disproportionately in the thylakoid membrane interior.

  18. Vipp1 is required for basic thylakoid membrane formation but not for the assembly of thylakoid protein complexes.

    Science.gov (United States)

    Aseeva, Elena; Ossenbühl, Friederich; Sippel, Claudia; Cho, Won K; Stein, Bernhard; Eichacker, Lutz A; Meurer, Jörg; Wanner, Gerhard; Westhoff, Peter; Soll, Jürgen; Vothknecht, Ute C

    2007-02-01

    Vipp1 (vesicle inducing protein in plastids 1) is found in cyanobacteria and chloroplasts where it is essential for thylakoid formation. Arabidopsis thaliana mutant plants with a reduction of Vipp1 to about 20% of wild type content become albinotic at an early stage. We propose that this drastic phenotype results from an inability of the remaining Vipp1 protein to assemble into a homo-oligomeric complex, indicating that oligomerization is a prerequisite for Vipp1 function. A Vipp1-ProteinA fusion protein, expressed in the Deltavipp1 mutant background, is able to reinstate oligomerization and restore photoautotrophic growth. Plants containing Vipp1-ProteinA in amounts comparable to Vipp1 in the wild type exhibit a wild type phenotype. However, plants with a reduced amount of Vipp1-ProteinA protein are growth-retarded and significantly paler than the wild type. This phenotype is caused by a decrease in thylakoid membrane content and a concomitant reduction in photosynthetic activity. To the extent that thylakoid membranes are made in these plants they are properly assembled with protein-pigment complexes and are photosynthetically active. This strongly supports a function of Vipp1 in basic thylakoid membrane formation and not in the functional assembly of thylakoid protein complexes. Intriguingly, electron microscopic analysis shows that chloroplasts in the mutant plants are not equally affected by the Vipp1 shortage. Indeed, a wide range of different stages of thylakoid development ranging from wild-type-like chloroplasts to plastids nearly devoid of thylakoids can be observed in organelles of one and the same cell.

  19. Global Proteomic Analysis Reveals an Exclusive Role of Thylakoid Membranes in Bioenergetics of a Model Cyanobacterium.

    Science.gov (United States)

    Liberton, Michelle; Saha, Rajib; Jacobs, Jon M; Nguyen, Amelia Y; Gritsenko, Marina A; Smith, Richard D; Koppenaal, David W; Pakrasi, Himadri B

    2016-06-01

    Cyanobacteria are photosynthetic microbes with highly differentiated membrane systems. These organisms contain an outer membrane, plasma membrane, and an internal system of thylakoid membranes where the photosynthetic and respiratory machinery are found. This existence of compartmentalization and differentiation of membrane systems poses a number of challenges for cyanobacterial cells in terms of organization and distribution of proteins to the correct membrane system. Proteomics studies have long sought to identify the components of the different membrane systems in cyanobacteria, and to date about 450 different proteins have been attributed to either the plasma membrane or thylakoid membrane. Given the complexity of these membranes, many more proteins remain to be identified, and a comprehensive catalogue of plasma membrane and thylakoid membrane proteins is needed. Here we describe the identification of 635 differentially localized proteins in Synechocystis sp. PCC 6803 by quantitative iTRAQ isobaric labeling; of these, 459 proteins were localized to the plasma membrane and 176 were localized to the thylakoid membrane. Surprisingly, we found over 2.5 times the number of unique proteins identified in the plasma membrane compared with the thylakoid membrane. This suggests that the protein composition of the thylakoid membrane is more homogeneous than the plasma membrane, consistent with the role of the plasma membrane in diverse cellular processes including protein trafficking and nutrient import, compared with a more specialized role for the thylakoid membrane in cellular energetics. Thus, our data clearly define the two membrane systems with distinct functions. Overall, the protein compositions of the Synechocystis 6803 plasma membrane and thylakoid membrane are quite similar to that of the plasma membrane of Escherichia coli and thylakoid membrane of Arabidopsis chloroplasts, respectively. Synechocystis 6803 can therefore be described as a Gram

  20. Correlation between spatial (3D) structure of pea and bean thylakoid membranes and arrangement of chlorophyll-protein complexes

    NARCIS (Netherlands)

    Rumak, Izabela; Mazur, Radoslaw; Gieczewska, Katarzyna; Koziol-Lipinska, Joanna; Kierdaszuk, Borys; Michalski, Wojtek P.; Shiell, Brian J.; Venema, Jan Henk; Vredenberg, Wim J.; Mostowska, Agnieszka; Garstka, Maciej

    2012-01-01

    Background: The thylakoid system in plant chloroplasts is organized into two distinct domains: grana arranged in stacks of appressed membranes and non-appressed membranes consisting of stroma thylakoids and margins of granal stacks. It is argued that the reason for the development of appressed membr

  1. Light-dependent reversal of dark-chilling induced changes in chloroplast structure and arrangement of chlorophyll-protein complexes in bean thylakoid membranes

    NARCIS (Netherlands)

    Garstka, M; Drozak, A; Rosiak, M; Venema, JH; Kierdaszuk, B; Simeonova, E; van Hasselt, PR; Dobrucki, J; Mostowska, A

    2005-01-01

    Changes in chloroplast structure and rearrangement of chlorophyll-protein (CP) complexes were investigated in detached leaves of bean (Phaseolus vulgaris L. cv. Eureka), a chilling-sensitive plant, during 5-day dark-chilling at 1 degrees C and subsequent 3-h photoactivation under white light (200 mu

  2. Chloroplast membrane transport: interplay of prokaryotic and eukaryotic traits.

    Science.gov (United States)

    Vothknecht, Ute C; Soll, Jürgen

    2005-07-18

    Chloroplasts are specific plant organelles of prokaryotic origin. They are separated from the surrounding cell by a double membrane, which represents an effective barrier for the transport of metabolites and proteins. Specific transporters in the inner envelope membrane have been described, which facilitate the exchange of metabolites. In contrast, the outer envelope has been viewed for a long time as a molecular sieve that offers a mere size constriction to the passage of molecules. This view has been challenged lately, and a number of specific and regulated pore proteins of the outer envelope (OEPs) have been identified. These pores seem to have originated by adaptation of outer membrane proteins of the cyanobacterial ancestor of the chloroplast. In a similar fashion, the transport of proteins across the two envelope membranes is achieved by two hetero-oligomeric protein complexes called Toc (translocon in the outer envelope of chloroplasts) and Tic (translocon in the inner envelope of chloroplasts). The phylogenetic provenance of the translocon components is less clear, but at least the channel protein of the Toc translocon is of cyanobacterial origin. Characteristic of cyanobacteria and chloroplasts is furthermore a specialized internal membrane system, the thylakoids, on which the components of the photosynthetic machinery are located. Despite the importance of this membrane, very little is known about its phylogenetic origin or the manner of its synthesis. Vipp1 appears to be a ubiquitous component of thylakoid formation, while in chloroplasts of land plants, additionally a vesicle transport system of eukaryotic origin might be involved in this process.

  3. Supramolecular Organization of Thylakoid Membranes in De-etioplasts Upon Exposure to Light

    Institute of Scientific and Technical Information of China (English)

    Yangyang Wang; Weitong Cui; Shihua Shen; Hui Chen

    2012-01-01

    Chloroplast is the most prominent form of plastid occurring in all green plant tissues and contains a thylakoid membrane system that carries the photosynthetic electron transport chain converting light energy into chemical energy in the forms of ATP and NADPH.Thylakoids are the dominating structure inside fully mature chloroplasts.The formation and alteration of the thylakoid membrane structure and composition are closely connected to the development of the chloroplasts from simple,undifferentiated proplastids.Despite the importance of thylakoid membranes for photosynthesis and the energy metabolism of plants,the molecular processes connected to the origin,synthesis,maintenance and adaptation of thylakoid membranes remain poorly understood.In this study,the proteome difference in developing thylakoid membranes,including integral and peripheral proteins,in the process of de-etiolated rice seedlings were analyzed by 2-DE,and the structural changes and effects of such difference on photosynthetic ability were also examined.The ultrastructure of etioplasts changed notably upon exposure to light for 1 h,5 h and 9 h.After 5 h of illumination,paracrystalline PLB transformation was completed with transverse short tubules dispersed in stroma.The first stacked thylakoid membranes were observed at time point 9 h of illumination.In the continuous illumination,the formation of intergranal thylakoid was directly from PLB material without an intervening vesicular stage.In order to investigate the function and organization of the photosynthetic membrane proteins,the low-temperature (77 K) fluorescence emission spectra of membrane factions isolated from etioplasts,de-etioplasts and mature chloroplasts were investigated.The fluorescence emission spectra from de-etioplasts and mature chloroplasts exhibited a clear red maximum centered at 681 nm and a shoulder in the far-red region near 735 nm.The 681 nm band was slightly red-shifted with the increase of illumination duration

  4. Photoactivation of electrogenic activity in chloroplasts and its relation to photoinduced swelling of thylakoids

    NARCIS (Netherlands)

    Bulychev, A.A.; Vredenberg, W.J.

    2000-01-01

    In patch-clamp experiments on isolated chloroplasts of Peperomia metallica Lind. et Rodig. (Piperaceae), the replacement of 50 mM KCl in a medium with 50 mM NH4Cl strongly influenced the parameters of photocurrent known to reflect the generation of electric potential in thylakoids. The addition of N

  5. Spinach thylakoid polyphenol oxidase isolation, activation, and properties of the native chloroplast enzyme

    Energy Technology Data Exchange (ETDEWEB)

    Golbeck, J.H.; Cammarata, K.V.

    1981-05-01

    Polyphenol oxidase activity (E.C. 1.14,18.1) has been found in two enzyme species isolated from thylakoid membranes of spinach chloroplasts. The proteins were released from the membrane by sonication and purified >900-fold by ammonium sulfate precipitation, gel filtration, and ion-exchange chromatography. The enzymes appear to be the tetramer and monomer of a subunit with a molecular weight of 42,500 as determined by lithium dodecyl sulfate gel electrophoresis. Sonication releases polyphenol oxidase from the membrane largely in the latent state. In the absence of added fatty acids, the isolated enzyme spontaneously, but slowly, activates with time. Purified polyphenol oxidase utilizes o-diphenols as substrates and shows no detectable levels of monophenol or p-diphenol oxidase activities. Suitable substrates include chlorogenic acid, catechol, caffeic acid, pyrogallol, and dopamine; however, the enzyme is substrate-inhibited by the last four at concentrations near their K/sub m/. A large seasonal variation in polyphenol oxidase activity may result from a decrease in enzyme content rather than inhibition of the enzyme present.

  6. Hydroponics on a chip: analysis of the Fe deficient Arabidopsis thylakoid membrane proteome.

    Science.gov (United States)

    Laganowsky, Arthur; Gómez, Stephen M; Whitelegge, Julian P; Nishio, John N

    2009-04-13

    The model plant Arabidopsis thaliana was used to evaluate the thylakoid membrane proteome under Fe-deficient conditions. Plants were cultivated using a novel hydroponic system, called "hydroponics on a chip", which yields highly reproducible plant tissue samples for physiological analyses, and can be easily used for in vivo stable isotope labeling. The thylakoid membrane proteome, from intact chloroplasts isolated from Fe-sufficient and Fe-deficient plants grown with hydroponics on a chip, was analyzed using liquid chromatography coupled to mass spectrometry. Intact masses of thylakoid membrane proteins were measured, many for the first time, and several proteins were identified with post-translational modifications that were altered by Fe deficiency; for example, the doubly phosphorylated form of the photosystem II oxygen evolving complex, PSBH, increased under Fe-deficiency. Increased levels of photosystem II protein subunit PSBS were detected in the Fe-deficient samples. Antioxidant enzymes, including ascorbate peroxidase and peroxiredoxin Q, were only detected in the Fe-deficient samples. We present the first biochemical evidence that the two major LHC IIb proteins (LHCB1 and LHCB2) may have significantly different functions in the thylakoid membrane. The study illustrates the utility of intact mass proteomics as an indispensable tool for functional genomics. "Hydroponics on a chip" provides the ability to grow A. thaliana under defined conditions that will be useful for systems biology.

  7. Identification of Thylakoid Membrane Protein Complexes by Using a BN-Chip/MS Approach

    Institute of Scientific and Technical Information of China (English)

    Longquan Fan; Yinghong Pan

    2012-01-01

    Thylakoid membrane protein complexes of wheat (Triticum aestivum Linn.)play crucial roles in growth and crop production.Knowledge of the composition and structure of protein complexes,as well as protein interactions,will result in a much deeper understanding of metabolic pathways and cellular processes than protein identities alone,especially if the complexes can be separated in the native forms.Whereas the analysis of membrane protein complexes is a significant challenge due to their hydrophobic properties and relatively low abundance.A rapid and efficient method of identifying membrane protein complexes will greatly facilitate the investigation of agriculture.The present work developed an BN-Chip/MS approach for exhaustive separation and identification of protein complexes,by combining using blue-native polyacrylamide gel electrophoresis (BN-PAGE) and chip-based high-performance liquid chromatography quadruple time-of-flight tandem mass spectrometry (HPLC-Chip/ESI-QT-OF-MS,Chip/MS).By using this approach,seventy-five nonredundant proteins of wheat thylakoid membrane complexes were identified from digested 13 bands of BN-gel.When the protocol of BN separation was not used,only 37 nonredundant proteins had been identified and among of them 9 proteins were uniquely identi? ed.This BN-Chip/MS approach is rapid and efficient for identifying protein complexes in wheat thylakoid membranes,and also providing reliable foundations for further functional research of wheat chloroplast and for identifying protein complexes of other species.

  8. Anisotropic Circular Dichroism Signatures of Oriented Thylakoid Membranes and Lamellar Aggregates of LHCII

    Energy Technology Data Exchange (ETDEWEB)

    Miloslavina, Y.; Hind, G.; Lambrev, P. H.; Javorfi, T.; Varkonyi, Z.; Karlicky, V.; Wall, J. S.; Garab, G.

    2011-06-12

    In photosynthesis research, circular dichroism (CD) spectroscopy is an indispensable tool to probe molecular architecture at virtually all levels of structural complexity. At the molecular level, the chirality of the molecule results in intrinsic CD; pigment-pigment interactions in protein complexes and small aggregates can give rise to excitonic CD bands, while 'psi-type' CD signals originate from large, densely packed chiral aggregates. It has been well established that anisotropic CD (ACD), measured on samples with defined non-random orientation relative to the propagation of the measuring beam, carries specific information on the architecture of molecules or molecular macroassemblies. However, ACD is usually combined with linear dichroism and can be distorted by instrumental imperfections, which given the strong anisotropic nature of photosynthetic membranes and complexes, might be the reason why ACD is rarely studied in photosynthesis research. In this study, we present ACD spectra, corrected for linear dichroism, of isolated intact thylakoid membranes of granal chloroplasts, washed unstacked thylakoid membranes, photosystem II (PSII) membranes (BBY particles), grana patches, and tightly stacked lamellar macroaggregates of the main light-harvesting complex of PSII (LHCII). We show that the ACD spectra of face- and edge-aligned stacked thylakoid membranes and LHCII lamellae exhibit profound differences in their psi-type CD bands. Marked differences are also seen in the excitonic CD of BBY and washed thylakoid membranes. Magnetic CD (MCD) spectra on random and aligned samples, and the largely invariable nature of the MCD spectra, despite dramatic variations in the measured isotropic and anisotropic CD, testify that ACD can be measured without substantial distortions and thus employed to extract detailed information on the (supra)molecular organization of photosynthetic complexes. An example is provided showing the ability of CD data to indicate such an

  9. Anisotropic Circular Dichroism Signatures of Oriented Thylakoid Membranes and Lamellar Aggregates of LHCII

    Energy Technology Data Exchange (ETDEWEB)

    Miloslavina Y.; Hind G.; Lambrev, P. H.; Javorfi, T.; Varkonyi, Z.; Karlicky, V.; Wall, J. S.; Garab, G.

    2012-03-01

    In photosynthesis research, circular dichroism (CD) spectroscopy is an indispensable tool to probe molecular architecture at virtually all levels of structural complexity. At the molecular level, the chirality of the molecule results in intrinsic CD; pigment-pigment interactions in protein complexes and small aggregates can give rise to excitonic CD bands, while 'psi-type' CD signals originate from large, densely packed chiral aggregates. It has been well established that anisotropic CD (ACD), measured on samples with defined non-random orientation relative to the propagation of the measuring beam, carries specific information on the architecture of molecules or molecular macroassemblies. However, ACD is usually combined with linear dichroism and can be distorted by instrumental imperfections, which given the strong anisotropic nature of photosynthetic membranes and complexes, might be the reason why ACD is rarely studied in photosynthesis research. In this study, we present ACD spectra, corrected for linear dichroism, of isolated intact thylakoid membranes of granal chloroplasts, washed unstacked thylakoid membranes, photosystem II (PSII) membranes (BBY particles), grana patches, and tightly stacked lamellar macroaggregates of the main light-harvesting complex of PSII (LHCII). We show that the ACD spectra of face- and edge-aligned stacked thylakoid membranes and LHCII lamellae exhibit profound differences in their psi-type CD bands. Marked differences are also seen in the excitonic CD of BBY and washed thylakoid membranes. Magnetic CD (MCD) spectra on random and aligned samples, and the largely invariable nature of the MCD spectra, despite dramatic variations in the measured isotropic and anisotropic CD, testify that ACD can be measured without substantial distortions and thus employed to extract detailed information on the (supra)molecular organization of photosynthetic complexes. An example is provided showing the ability of CD data to indicate such an

  10. The role of putrescine in the regulation of proteins and fatty acids of thylakoid membranes under salt stress.

    Science.gov (United States)

    Shu, Sheng; Yuan, Yinghui; Chen, Jie; Sun, Jin; Zhang, Wenhua; Tang, Yuanyuan; Zhong, Min; Guo, Shirong

    2015-10-05

    Polyamines can alleviate the inhibitory effects of salinity on plant growth by regulating photosynthetic efficiency. However, little information is available to explain the specific mechanisms underlying the contribution of polyamines to salt tolerance of the photosynthetic apparatus. Here, we investigated the role of putrescine (Put) on the photosynthetic apparatus of cucumber seedlings under salt stress. We found that NaCl stress resulted in severe ion toxicity and oxidative stress in cucumber chloroplasts. In addition, salinity caused a significant increase in the saturated fatty acid contents of thylakoid membranes. Put altered unsaturated fatty acid content, thereby alleviating the disintegration of thylakoid grana lamellae and reducing the number of plastoglobuli in thylakoid membranes. BN-PAGE revealed Put up-regulated the expression of ATP synthase, CP47, D1, Qb, and psbA proteins and down-regulated CP24, D2, and LHCII type III in NaCl-stressed thylakoid membranes. qRT-PCR analysis of gene expression was used to compare transcript and protein accumulation among 10 candidate proteins. For five of these proteins, induced transcript accumulation was consistent with the pattern of induced protein accumulation. Our results suggest that Put regulates protein expression at transcriptional and translational levels by increasing endogenous polyamines levels in thylakoid membranes, which may stabilise photosynthetic apparatus under salt stress.

  11. Localization of phosphatidylcholine in outer envelope membrane of spinach chloroplasts

    Science.gov (United States)

    1985-01-01

    We have examined the effects of phospholipase C from Bacillus cereus on the extent of phospholipid hydrolysis in envelope membrane vesicles and in intact chloroplasts. When isolated envelope vesicles were incubated in presence of phospholipase C, phosphatidylcholine and phosphatidylglycerol, but not phosphatidylinositol, were totally converted into diacylglycerol if they were available to the enzyme (i.e., when the vesicles were sonicated in presence of phospholipase C). These experiments demonstrate that phospholipase C can be used to probe the availability of phosphatidylcholine and phosphatidylglycerol in the cytosolic leaflet of the outer envelope membrane from spinach chloroplasts. When isolated, purified, intact chloroplasts were incubated with low amounts of phospholipase C (0.3 U/mg chlorophyll) under very mild conditions (12 degrees C for 1 min), greater than 80% of phosphatidylcholine molecules and almost none of phosphatidylglycerol molecules were hydrolyzed. Since we have also demonstrated, by using several different methods (phase-contrast and electron microscopy, immunochemical and electrophoretic analyses) that isolated spinach chloroplasts, and especially their outer envelope membrane, remained intact after mild treatment with phospholipase C, we can conclude that there is a marked asymmetric distribution of phospholipids across the outer envelope membrane of spinach chloroplasts. Phosphatidylcholine, the major polar lipid of the outer envelope membrane, is almost entirely accessible from the cytosolic side of the membrane and therefore is probably localized in the outer leaflet of the outer envelope bilayer. On the contrary, phosphatidylglycerol, the major polar lipid in the inner envelope membrane and the thylakoids, is probably not accessible to phospholipase C from the cytosol and therefore is probably localized mostly in the inner leaflet of the outer envelope membrane and in the other chloroplast membranes. PMID:3988805

  12. A bestrophin-like protein modulates the proton motive force across the thylakoid membrane in Arabidopsis

    Institute of Scientific and Technical Information of China (English)

    Zhikun Duan; Fanna Kong; Lin Zhang; Wenjing Li; Jiao Zhang; Lianwei Peng

    2016-01-01

    During photosynthesis, photosynthetic electron transport generates a proton motive force (pmf) across the thylakoid membrane, which is used for ATP biosynthesis via ATP synthase in the chloroplast. The pmf is composed of an electric potential (DC) and an osmotic component (DpH). Partitioning between these components in chloroplasts is strictly regulated in response to fluctuating environments. However, our knowledge of the molecular mechanisms that regulate pmf partitioning is limited. Here, we report a bestrophin-like protein (AtBest), which is critical for pmf partitioning. While the DpH component was slightly reduced in atbest, the DC component was much greater in this mutant than in the wild type, resulting in less efficient activation of nonphotochemical quenching (NPQ) upon both illumination and a shift from low light to high light. Although no visible phenotype was observed in the atbest mutant in the greenhouse, this mutant exhibited stronger photoinhibition than the wild type when grown in the field. AtBest belongs to the bestrophin family proteins, which are believed to function as chloride (Cl?) channels. Thus, our findings reveal an important Cl? channel required for ion transport and homeo-stasis across the thylakoid membrane in higher plants. These processes are essential for fine-tuning photosynthesis under fluctuating environmental conditions.

  13. Global Proteomic Analysis Reveals an Exclusive Role of Thylakoid Membranes in Bioenergetics of a Model Cyanobacterium

    Energy Technology Data Exchange (ETDEWEB)

    Liberton, Michelle; Saha, Rajib; Jacobs, Jon M.; Nguyen, Amelia Y.; Gritsenko, Marina A.; Smith, Richard D.; Koppenaal, David W.; Pakrasi, Himadri B.

    2016-04-07

    Cyanobacteria are photosynthetic microbes with highly differentiated membrane systems. These organisms contain an outer membrane, plasma membrane, and an internal system of thylakoid membranes where the photosynthetic and respiratory machinery are found. This existence of compartmentalization and differentiation of membrane systems poses a number of challenges for cyanobacterial cells in terms of organization and distribution of proteins to the correct membrane system. Proteomics studies have long sought to identify the components of the different membrane systems, and to date about 450 different proteins have been attributed to either the plasma membrane or thylakoid membrane. Given the complexity of these membranes, many more proteins remain to be identified in these membrane systems, and a comprehensive catalog of plasma membrane and thylakoid membrane proteins is needed. Here we describe the identification of 635 proteins in Synechocystis sp. PCC 6803 by quantitative iTRAQ isobaric labeling; of these, 459 proteins were localized to the plasma membrane and 176 were localized to the thylakoid membrane. Surprisingly, we found over 2.5 times the number of unique proteins identified in the plasma membrane compared to the thylakoid membrane. This suggests that the protein composition of the thylakoid membrane is more homogeneous than the plasma membrane, consistent with the role of the plasma membrane in diverse cellular processes including protein trafficking and nutrient import, compared to a more specialized role for the thylakoid membrane in cellular energetics. Overall, the protein composition of the Synechocystis 6803 plasma membrane and thylakoid membrane is quite similar to the E.coli plasma membrane and Arabidopsis thylakoid membrane, respectively. Synechocystis 6803 can therefore be described as a gram-negative bacterium that has an additional internal membrane system that fulfils the energetic requirements of the cell.

  14. Ultrastructural changes in the membrane system of isolated chloroplasts of spinach under the influence of carbonic anhydrase inhibitors AA and EA

    Directory of Open Access Journals (Sweden)

    Marina V. Vodka

    2013-04-01

    Full Text Available The effects of carbonic anhydrase inhibitors (АА and EA on the membrane system of isolated chloroplasts of spinach were investigated. Under the influence of AA the considerable alterations in granal structure occurred, the thickness of the granal thylakoids increased by 36% and the interspace between thylakoids by 10% comparable with the control. As a result of EA treatment, the thickness of granal thylakoids enhanced by 31% and the interspace between thylakoids increased by 8% in comparison to the control. It was shown that structure of the granal system of the chloroplast was more sensitive to AA than EA. The data obtained can indicate a decrease in the activity of the thylakoid carbonic anhydrase, inhibition of electron transport and photosynthetic process as a whole in the presence of carbonic anhydrase inhibitors (AA and EA.

  15. The changes in the chloroplast membranes of pea leaves under the influence of carbonic anhydrase inhibitors (ions of copper and zinc

    Directory of Open Access Journals (Sweden)

    M.V. Vodka

    2014-04-01

    Full Text Available Тhe effects of carbonic anhydrase inhibitors, such as ions Cu2+ and Zn2+, on the membrane system of chloroplasts in pea leaves were investigated. After treatment of pea leaves with 250 mM Cu2+ or 400 mM Zn2+ we observed changes in the granal structure and compactness of the thylakoids in granae. It was shown that the thickness of granal thylakoids and the interspace between thylakoids increased comparing to control. Changes of the size and structure of thylakoids and granae in treated leaves may be associated with the enhanced accumulation of CO2 in the membrane. It is suggested that the carbonic anhydrase may also play a structural role in chloroplast granae.

  16. Genetic Analysis of Arabidopsis Mutants Impaired in Plastid Lipid Import Reveals a Role of Membrane Lipids in Chloroplast Division

    Energy Technology Data Exchange (ETDEWEB)

    Fan, J.; Xu, C.

    2011-03-01

    The biogenesis of photosynthetic membranes in plants relies largely on lipid import from the endoplasmic reticulum (ER) and this lipid transport process is mediated by TGD proteins in Arabidopsis. Such a dependency of chloroplast biogenesis on ER-to-plastid lipid transport was recently exemplified by analyzing double mutants between tgd1-1 or tgd4-3 and fad6 mutants. The fad6 mutants are defective in the desaturation of membrane lipids in chloroplasts and therefore dependent on import of polyunsaturated lipid precursors from the ER for constructing a competent thylakoid membrane system. In support of a critical role of TGD proteins in ER-to-plastid lipid trafficking, we showed that the introduction of the tgd mutations into fad6 mutant backgrounds led to drastic reductions in relative amounts of thylakoid lipids. Moreover, the tgd1-1 fad6 and tgd4-3 fad6 double mutants were deficient in polyunsaturated fatty acids in chloroplast membrane lipids, and severely compromised in the biogenesis of photosynthetic membrane systems. Here we report that these double mutants are severely impaired in chloroplast division. The possible role of membrane lipids in chloroplast division is discussed.

  17. A novel chloroplast localized Rab GTPase protein CPRabA5e is involved in stress, development, thylakoid biogenesis and vesicle transport in Arabidopsis.

    Science.gov (United States)

    Karim, Sazzad; Alezzawi, Mohamed; Garcia-Petit, Christel; Solymosi, Katalin; Khan, Nadir Zaman; Lindquist, Emelie; Dahl, Peter; Hohmann, Stefan; Aronsson, Henrik

    2014-04-01

    A novel Rab GTPase protein in Arabidopsis thaliana, CPRabA5e (CP = chloroplast localized) is located in chloroplasts and has a role in transport. Transient expression of CPRabA5e:EGFP fusion protein in tobacco (Nicotiana tabacum) leaves, and immunoblotting using Arabidopsis showed localization of CPRabA5e in chloroplasts (stroma and thylakoids). Ypt31/32 in the yeast Saccharomyces cerevisiae are involved in regulating vesicle transport, and CPRabA5e a close homolog of Ypt31/32, restores the growth of the ypt31Δ ypt32(ts) mutant at 37 °C in yeast complementation. Knockout mutants of CPRabA5e displayed delayed seed germination and growth arrest during oxidative stress. Ultrastructural studies revealed that after preincubation at 4 °C mutant chloroplasts contained larger plastoglobules, lower grana, and more vesicles close to the envelopes compared to wild type, and vesicle formation being enhanced under oxidative stress. This indicated altered thylakoid development and organization of the mutants. A yeast-two-hybrid screen with CPRabA5e as bait revealed 13 interacting partner proteins, mainly located in thylakoids and plastoglobules. These proteins are known or predicted to be involved in development, stress responses, and photosynthesis related processes, consistent with the stress phenotypes observed. The results observed suggest a role of CPRabA5e in transport to and from thylakoids, similar to cytosolic Rab proteins involved in vesicle transport.

  18. Multiple sources of carbonic anhydrase activity in pea thylakoids: soluble and membrane-bound forms.

    Science.gov (United States)

    Rudenko, Natalia N; Ignatova, Lyudmila K; Ivanov, Boris N

    2007-01-01

    Carbonic anhydrase (CA) activity of pea thylakoids, thylakoid membranes enriched with photosystem I (PSI-membranes), or photosystem II (PSII-membranes) as well as both supernatant and pellet after precipitation of thylakoids treated with detergent Triton X-100 were studied. CA activity of thylakoids in the presence of varying concentrations of Triton X-100 had two maxima, at Triton/chlorophyll (triton/Chl) ratios of 0.3 and 1.0. CA activities of PSI-membranes and PSII-membranes had only one maximum each, at Triton/Chl ratio 0.3 or 1.0, respectively. Two CAs with characteristics of the membrane-bound proteins and one CA with characteristics of the soluble proteins were found in the medium after thylakoids were incubated with Triton. One of the first two CAs had mobility in PAAG after native electrophoresis the same as that of CA residing in PSI-membranes, and the other CA had mobility the same as the mobility of CA residing in PSII-membranes, but the latter was different from CA situated in PSII core-complex (Ignatova et al. 2006 Biochemistry (Moscow) 71:525-532). The properties of the "soluble" CA removed from thylakoids were different from the properties of the known soluble CAs of plant cell: apparent molecular mass was about 262 kD and it was three orders more sensitive to the specific CA inhibitor, ethoxyzolamide, than soluble stromal CA. The data are discussed as indicating the presence of, at least, four CAs in pea thylakoids.

  19. Arabidopsis ANGULATA10 is required for thylakoid biogenesis and mesophyll development.

    Science.gov (United States)

    Casanova-Sáez, Rubén; Mateo-Bonmatí, Eduardo; Kangasjärvi, Saijaliisa; Candela, Héctor; Micol, José Luis

    2014-06-01

    The chloroplasts of land plants contain internal membrane systems, the thylakoids, which are arranged in stacks called grana. Because grana have not been found in Cyanobacteria, the evolutionary origin of genes controlling the structural and functional diversification of thylakoidal membranes in land plants remains unclear. The angulata10-1 (anu10-1) mutant, which exhibits pale-green rosettes, reduced growth, and deficient leaf lateral expansion, resulting in the presence of prominent marginal teeth, was isolated. Palisade cells in anu10-1 are larger and less packed than in the wild type, giving rise to large intercellular spaces. The ANU10 gene encodes a protein of unknown function that localizes to both chloroplasts and amyloplasts. In chloroplasts, ANU10 associates with thylakoidal membranes. Mutant anu10-1 chloroplasts accumulate H2O2, and have reduced levels of chlorophyll and carotenoids. Moreover, these chloroplasts are small and abnormally shaped, thylakoidal membranes are less abundant, and their grana are absent due to impaired thylakoid stacking in the anu10-1 mutant. Because the trimeric light-harvesting complex II (LHCII) has been reported to be required for thylakoid stacking, its levels were determined in anu10-1 thylakoids and they were found to be reduced. Together, the data point to a requirement for ANU10 for chloroplast and mesophyll development.

  20. Carbonic anhydrase activity in Arabidopsis thaliana thylakoid membrane and fragments enriched with PSI or PSII.

    Science.gov (United States)

    Ignatova, Lyudmila K; Rudenko, Natalia N; Mudrik, Vilen A; Fedorchuk, Tat'yana P; Ivanov, Boris N

    2011-12-01

    The procedure of isolating the thylakoids and the thylakoid membrane fragments enriched with either photosystem I or photosystem II (PSI- and PSII-membranes) from Arabidopsis thaliana leaves was developed. It differed from the one used with pea and spinach in durations of detergent treatment and centrifugation, and in concentrations of detergent and Mg(2+) in the media. Both the thylakoid and the fragments preserved carbonic anhydrase (CA) activities. Using nondenaturing electrophoresis followed by detection of CA activity in the gel stained with bromo thymol blue, one low molecular mass carrier of CA activity was found in the PSI-membranes, and two carriers, a low molecular mass one and a high molecular mass one, were found in the PSII-membranes. The proteins in the PSII-membranes differed in their sensitivity to acetazolamide (AA), a specific CA inhibitor. AA at 5 × 10(-7) M inhibited the CA activity of the high molecular mass protein but stimulated the activity of the low molecular mass carrier in the PSII-membranes. At the same concentration, AA moderately inhibited, by 30%, the CA activity of PSI-membranes. CA activity of the PSII-membranes was almost completely suppressed by the lipophilic CA inhibitor, ethoxyzolamide at 10(-9) M, whereas CA activity of the PSI-membranes was inhibited by this inhibitor even at 5 × 10(-7) M just the same as for AA. The observed distribution of CA activity in the thylakoid membranes from A. thaliana was close to the one found in the membranes of pea, evidencing the general pattern of CA activity in the thylakoid membranes of C3-plants.

  1. Membrane heredity and early chloroplast evolution.

    Science.gov (United States)

    Cavalier-Smith, T

    2000-04-01

    Membrane heredity was central to the unique symbiogenetic origin from cyanobacteria of chloroplasts in the ancestor of Plantae (green plants, red algae, glaucophytes) and to subsequent lateral transfers of plastids to form even more complex photosynthetic chimeras. Each symbiogenesis integrated disparate genomes and several radically different genetic membranes into a more complex cell. The common ancestor of Plantae evolved transit machinery for plastid protein import. In later secondary symbiogeneses, signal sequences were added to target proteins across host perialgal membranes: independently into green algal plastids (euglenoids, chlorarachneans) and red algal plastids (alveolates, chromists). Conservatism and innovation during early plastid diversification are discussed.

  2. Distribution and dynamics of electron transport complexes in cyanobacterial thylakoid membranes.

    Science.gov (United States)

    Liu, Lu-Ning

    2016-03-01

    The cyanobacterial thylakoid membrane represents a system that can carry out both oxygenic photosynthesis and respiration simultaneously. The organization, interactions and mobility of components of these two electron transport pathways are indispensable to the biosynthesis of thylakoid membrane modules and the optimization of bioenergetic electron flow in response to environmental changes. These are of fundamental importance to the metabolic robustness and plasticity of cyanobacteria. This review summarizes our current knowledge about the distribution and dynamics of electron transport components in cyanobacterial thylakoid membranes. Global understanding of the principles that govern the dynamic regulation of electron transport pathways in nature will provide a framework for the design and synthetic engineering of new bioenergetic machinery to improve photosynthesis and biofuel production. This article is part of a Special Issue entitled: Organization and dynamics of bioenergetic systems in bacteria, edited by Conrad Mullineaux.

  3. Proteomics of the chloroplast: systematic identification and targeting analysis of lumenal and peripheral thylakoid proteins

    DEFF Research Database (Denmark)

    Peltier, J B; Friso, G; Kalume, D E;

    2000-01-01

    the twin arginine motif that is characteristic for substrates of the TAT pathway. Logoplots were used to provide a detailed analysis of the lumenal targeting signals, and all nuclear-encoded proteins identified on the two-dimensional gels were used to test predictions for chloroplast localization...... and transit peptides made by the software programs ChloroP, PSORT, and SignalP. A combination of these three programs was found to provide a useful tool for evaluating chloroplast localization and transit peptides and also could reveal possible alternative processing sites and dual targeting. The potential...

  4. Biotic stress induced demolition of thylakoid structure and loss in photoelectron transport of chloroplasts in papaya leaves.

    Science.gov (United States)

    Nanda, Rashmi Madhumita; Biswal, Basanti

    2008-04-01

    Papaya mosaic virus (PMV) causes severe mosaic symptoms in the papaya (Carica papaya L.) leaves. The PMV-induced alterations in photosystem II (PS II) structure and photochemical functions were probed. An increase in chlorophyll a (Chl a) fluorescence polarization suggests pathogen-induced transformation of thylakoid membrane to a gel phase. This transformation in physical state of thylakoid membrane may result in alteration in topology of pigments on pigment-binding proteins as reflected in pathogen-induced loss in the efficiency of energy transfer from carotenoids to chlorophylls. The fast Chl a fluorescence induction kinetics of healthy and PMV-infected plants by F(O)-F(J)-F(I)-F(P) transients revealed pathogen-induced perturbation on PS II acceptor side electron transfer equilibrium between Q(A) and Q(B) and in the pool size of electron transport acceptors. Pathogen-induced loss in photosynthetic pigments, changes in thylakoid structure and decrease in the ratio of F(V)/F(M) (photochemical potential of PS II) further correlate with the loss in photoelectron transport of PS II as probed by 2,6-dichlorophenol indophenol (DCPIP)-Hill reaction. Restoration of the loss by 1,5-diphenyl carbazide (DPC), an exogenous electron donor, that donates electron directly to reaction centre II bypassing the oxygen evolving system (OES), leads towards the conclusion that OES is one of the major targets of biotic stress. Further, the data suggest that chlorophyll fluorescence could be used as a non-invasive handy tool to assess the loss in photosynthetic efficiency and symptom severity in infected green tissues vis-a-vis the healthy ones.

  5. Chloroplast outer envelope protein CHUP1 is essential for chloroplast anchorage to the plasma membrane and chloroplast movement.

    Science.gov (United States)

    Oikawa, Kazusato; Yamasato, Akihiro; Kong, Sam-Geun; Kasahara, Masahiro; Nakai, Masato; Takahashi, Fumio; Ogura, Yasunobu; Kagawa, Takatoshi; Wada, Masamitsu

    2008-10-01

    Chloroplasts change their intracellular distribution in response to light intensity. Previously, we isolated the chloroplast unusual positioning1 (chup1) mutant of Arabidopsis (Arabidopsis thaliana). This mutant is defective in normal chloroplast relocation movement and shows aggregation of chloroplasts at the bottom of palisade mesophyll cells. The isolated gene encodes a protein with an actin-binding motif. Here, we used biochemical analyses to determine the subcellular localization of full-length CHUP1 on the chloroplast outer envelope. A CHUP1-green fluorescent protein (GFP) fusion, which was detected at the outermost part of mesophyll cell chloroplasts, complemented the chup1 phenotype, but GFP-CHUP1, which was localized mainly in the cytosol, did not. Overexpression of the N-terminal hydrophobic region (NtHR) of CHUP1 fused with GFP (NtHR-GFP) induced a chup1-like phenotype, indicating a dominant-negative effect on chloroplast relocation movement. A similar pattern was found in chloroplast OUTER ENVELOPE PROTEIN7 (OEP7)-GFP transformants, and a protein containing OEP7 in place of NtHR complemented the mutant phenotype. Physiological analyses of transgenic Arabidopsis plants expressing truncated CHUP1 in a chup1 mutant background and cytoskeletal inhibitor experiments showed that the coiled-coil region of CHUP1 anchors chloroplasts firmly on the plasma membrane, consistent with the localization of coiled-coil GFP on the plasma membrane. Thus, CHUP1 localization on chloroplasts, with the N terminus inserted into the chloroplast outer envelope and the C terminus facing the cytosol, is essential for CHUP1 function, and the coiled-coil region of CHUP1 prevents chloroplast aggregation and participates in chloroplast relocation movement.

  6. Specific interaction of IM30/Vipp1 with cyanobacterial and chloroplast membranes results in membrane remodeling and eventually in membrane fusion.

    Science.gov (United States)

    Heidrich, Jennifer; Thurotte, Adrien; Schneider, Dirk

    2017-04-01

    The photosynthetic light reaction takes place within the thylakoid membrane system in cyanobacteria and chloroplasts. Besides its global importance, the biogenesis, maintenance and dynamics of this membrane system are still a mystery. In the last two decades, strong evidence supported the idea that these processes involve IM30, the inner membrane-associated protein of 30kDa, a protein also known as the vesicle-inducing protein in plastids 1 (Vipp1). Even though we just only begin to understand the precise physiological function of this protein, it is clear that interaction of IM30 with membranes is crucial for biogenesis of thylakoid membranes. Here we summarize and discuss forces guiding IM30-membrane interactions, as the membrane properties as well as the oligomeric state of IM30 appear to affect proper interaction of IM30 with membrane surfaces. Interaction of IM30 with membranes results in an altered membrane structure and can finally trigger fusion of adjacent membranes, when Mg(2+) is present. Based on recent results, we finally present a model summarizing individual steps involved in IM30-mediated membrane fusion. This article is part of a Special Issue entitled: Lipid order/lipid defects and lipid-control of protein activity edited by Dirk Schneider.

  7. The evolutionarily conserved protein PHOTOSYNTHESIS AFFECTED MUTANT71 is required for efficient manganese uptake at the thylakoid membrane in Arabidopsis

    DEFF Research Database (Denmark)

    Schneider, Anja; Steinberger, Iris; Herdean, Andrei;

    2016-01-01

    In plants, algae, and cyanobacteria, photosystem II (PSII) catalyzes the light-driven oxidation of water. The oxygen-evolving complex of PSII is a Mn4CaO5 cluster embedded in a well-defined protein environment in the thylakoid membrane. However, transport of manganese and calcium into the thylakoid...... thylakoids relative to the wild type. The changes in Ca2+ homeostasis were accompanied by an increased contribution of the transmembrane electrical potential to the proton motive force across the thylakoid membrane. PSII activity in pam71 plants and the corresponding Chlamydomonas reinhardtii mutant cgld1...... was restored by supplementation with Mn2+, but not Ca2+. Furthermore, PAM71 suppressed the Mn2+-sensitive phenotype of the yeast mutant Δpmr1. Therefore, PAM71 presumably functions in Mn2+ uptake into thylakoids to ensure optimal PSII performance....

  8. Salt-induced redox-independent phosphorylation of light harvesting chlorophyll a/b proteins in Dunaliella salina thylakoid membranes.

    Science.gov (United States)

    Liu, Xian-De; Shen, Yun-Gang

    2005-02-17

    This study investigated the regulation of the major light harvesting chlorophyll a/b protein (LHCII) phosphorylation in Dunaliella salina thylakoid membranes. We found that both light and NaCl could induce LHCII phosphorylation in D. salina thylakoid membranes. Treatments with oxidants (ferredoxin and NADP) or photosynthetic electron flow inhibitors (DCMU, DBMIB, and stigmatellin) inhibited LHCII phosphorylation induced by light but not that induced by NaCl. Furthermore, neither addition of CuCl(2), an inhibitor of cytochrome b(6)f complex reduction, nor oxidizing treatment with ferricyanide inhibited light- or NaCl-induced LHCII phosphorylation, and both salts even induced LHCII phosphorylation in dark-adapted D. salina thylakoid membranes as other salts did. Together, these results indicate that the redox state of the cytochrome b(6)f complex is likely involved in light- but not salt-induced LHCII phosphorylation in D. salina thylakoid membranes.

  9. Flip-flop of phospholipids in proteoliposomes reconstituted from detergent extract of chloroplast membranes: kinetics and phospholipid specificity.

    Directory of Open Access Journals (Sweden)

    Archita Rajasekharan

    Full Text Available Eukaryotic cells are compartmentalized into distinct sub-cellular organelles by lipid bilayers, which are known to be involved in numerous cellular processes. The wide repertoire of lipids, synthesized in the biogenic membranes like the endoplasmic reticulum and bacterial cytoplasmic membranes are initially localized in the cytosolic leaflet and some of these lipids have to be translocated to the exoplasmic leaflet for membrane biogenesis and uniform growth. It is known that phospholipid (PL translocation in biogenic membranes is mediated by specific membrane proteins which occur in a rapid, bi-directional fashion without metabolic energy requirement and with no specificity to PL head group. A recent study reported the existence of biogenic membrane flippases in plants and that the mechanism of plant membrane biogenesis was similar to that found in animals. In this study, we demonstrate for the first time ATP independent and ATP dependent flippase activity in chloroplast membranes of plants. For this, we generated proteoliposomes from Triton X-100 extract of intact chloroplast, envelope membrane and thylakoid isolated from spinach leaves and assayed for flippase activity using fluorescent labeled phospholipids. Half-life time of flipping was found to be 6 ± 1 min. We also show that: (a intact chloroplast and envelope membrane reconstituted proteoliposomes can flip fluorescent labeled analogs of phosphatidylcholine in ATP independent manner, (b envelope membrane and thylakoid reconstituted proteoliposomes can flip phosphatidylglycerol in ATP dependent manner, (c Biogenic membrane ATP independent PC flipping activity is protein mediated and (d the kinetics of PC translocation gets affected differently upon treatment with protease and protein modifying reagents.

  10. Quantification of superoxide radical production in thylakoid membrane using cyclic hydroxylamines.

    Science.gov (United States)

    Kozuleva, Marina; Klenina, Irina; Mysin, Ivan; Kirilyuk, Igor; Opanasenko, Vera; Proskuryakov, Ivan; Ivanov, Boris

    2015-12-01

    Applicability of two lipophilic cyclic hydroxylamines (CHAs), CM-H and TMT-H, and two hydrophilic CHAs, CAT1-H and DCP-H, for detection of superoxide anion radical (O2(∙-)) produced by the thylakoid photosynthetic electron transfer chain (PETC) of higher plants under illumination has been studied. ESR spectrometry was applied for detection of the nitroxide radical originating due to CHAs oxidation by O2(∙-). CHAs and corresponding nitroxide radicals were shown to be involved in side reactions with PETC which could cause miscalculation of O2(∙-) production rate. Lipophilic CM-H was oxidized by PETC components, reducing the oxidized donor of Photosystem I, P700(+), while at the same concentration another lipophilic CHA, TMT-H, did not reduce P700(+). The nitroxide radical was able to accept electrons from components of the photosynthetic chain. Electrostatic interaction of stable cation CAT1-H with the membrane surface was suggested. Water-soluble superoxide dismutase (SOD) was added in order to suppress the reaction of CHA with O2(∙-) outside the membrane. SOD almost completely inhibited light-induced accumulation of DCP(∙), nitroxide radical derivative of hydrophilic DCP-H, in contrast to TMT(∙) accumulation. Based on the results showing that change in the thylakoid lumen pH and volume had minor effect on TMT(∙) accumulation, the reaction of TMT-H with O2(∙-) in the lumen was excluded. Addition of TMT-H to thylakoid suspension in the presence of SOD resulted in the increase in light-induced O2 uptake rate, that argued in favor of TMT-H ability to detect O2(∙-) produced within the membrane core. Thus, hydrophilic DCP-H and lipophilic TMT-H were shown to be usable for detection of O2(∙-) produced outside and within thylakoid membranes.

  11. The supramolecular architecture, function, and regulation of thylakoid membranes in red algae: an overview.

    Science.gov (United States)

    Su, Hai-Nan; Xie, Bin-Bin; Zhang, Xi-Ying; Zhou, Bai-Cheng; Zhang, Yu-Zhong

    2010-11-01

    Red algae are a group of eukaryotic photosynthetic organisms. Phycobilisomes (PBSs), which are composed of various types of phycobiliproteins and linker polypeptides, are the main light-harvesting antennae in red algae, as in cyanobacteria. Two morphological types of PBSs, hemispherical- and hemidiscoidal-shaped, are found in different red algae species. PBSs harvest solar energy and efficiently transfer it to photosystem II (PS II) and finally to photosystem I (PS I). The PS I of red algae uses light-harvesting complex of PS I (LHC I) as a light-harvesting antennae, which is phylogenetically related to the LHC I found in higher plants. PBSs, PS II, and PS I are all distributed throughout the entire thylakoid membrane, a pattern that is different from the one found in higher plants. Photosynthesis processes, especially those of the light reactions, are carried out by the supramolecular complexes located in/on the thylakoid membranes. Here, the supramolecular architecture, function and regulation of thylakoid membranes in red algal are reviewed.

  12. Biochemical Properties and Inhibition Kinetics of Phosphatase from Wheat Thylakoid Membranes

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    A phosphatase that hydrolyses phosphate monoesters has been isolated from wheat thylakoid membranes.Biochemical properties and inhibition kinetics of the phosphatase were investigated using several ions, organic solvents, and inhibitors. Wheat (Triticum aestivum L. cv. PH82-2-2) thylakoid membrane phosphatase activity was activated by Mg2+, Ca2+, and Fe2+ and was inhibited by Mn2+ and Cu2+. For example, enzyme activity was activated 34.81% by 2 mmol/L Mg2+, but was inhibited 22.3% and 8.5% by 2 and 1 mmol/L Cu2+, respectively.Methanol, ethanol and glycol were all able to activate enzyme activity. Enzyme activity was activated 58.5%, 48.2%,and 8.7% by 40% ethanol, methanol and glycol, respectively. From these results, it can be seen that the degree of activation of the phosphatase was greatest for ethanol and the type of activation was uncompetitive. Moreover,the activity of the thylakoid membrane phosphatase was inhibited by molybdate, vanadate, phosphate, and fluoride and the type of inhibition produced by these elements was uncompetitive, non-competitive, competitive and mixed, respectively.

  13. Transport of Ions Across the Inner Envelope Membrane of Chloroplasts

    Energy Technology Data Exchange (ETDEWEB)

    McCarty, R. E.

    2004-06-02

    The technical report outlines the results of nine years of research on how ions cross the inner envelope membrane of chloroplasts. The ions include protons, nitrite, calcium and ferrous iron. Bicarbonate transport was also studied.

  14. Analysis of protein interactions at native chloroplast membranes by ellipsometry.

    Directory of Open Access Journals (Sweden)

    Verena Kriechbaumer

    Full Text Available Membrane bound receptors play vital roles in cell signaling, and are the target for many drugs, yet their interactions with ligands are difficult to study by conventional techniques due to the technical difficulty of monitoring these interactions in lipid environments. In particular, the ability to analyse the behaviour of membrane proteins in their native membrane environment is limited. Here, we have developed a quantitative approach to detect specific interactions between low-abundance chaperone receptors within native chloroplast membranes and their soluble chaperone partners. Langmuir-Schaefer film deposition was used to deposit native chloroplasts onto gold-coated glass slides, and interactions between the molecular chaperones Hsp70 and Hsp90 and their receptors in the chloroplast membranes were detected and quantified by total internal reflection ellipsometry (TIRE. We show that native chloroplast membranes deposited on gold-coated glass slides using Langmuir-Schaefer films retain functional receptors capable of binding chaperones with high specificity and affinity. Taking into account the low chaperone receptor abundance in native membranes, these binding properties are consistent with data generated using soluble forms of the chloroplast chaperone receptors, OEP61 and Toc64. Therefore, we conclude that chloroplasts have the capacity to selectively bind chaperones, consistent with the notion that chaperones play an important role in protein targeting to chloroplasts. Importantly, this method of monitoring by TIRE does not require any protein labelling. This novel combination of techniques should be applicable to a wide variety of membranes and membrane protein receptors, thus presenting the opportunity to quantify protein interactions involved in fundamental cellular processes, and to screen for drugs that target membrane proteins.

  15. Change of proton motive force across thylakoid membrane in soybean leaf during state transitions

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    Change of proton gradient across thylakoid membrane in soybean leaves was studied with millisecond delayed light emission (ms-DLE) during the course of state transitions which were indicated by the chlorophyll fluorescence at room temperature and 77 K. When dark-adapted leaves were induced to stateⅠ with far-red light, Fm/Fo, F685/F735 and the intensity of fast phase of ms-DLE were affected slightly. However, during the induction to stateⅡ with red light, both Fm/Fo and F685/F735 decreased immediately and the former were quicker than the latter. In this interval, the intensity of fast phase of ms-DLE increased to a maximum and then decreased to a lower value during the transition to stateⅡ. Nigericin, an uncoupler which eliminates the proton gradient across thylakoid membrane, inhibited the increase in the intensity of fast phase of ms-DLE during the transition to stateⅡ. Another uncoupler, valinomycin, which eliminates the membrane potential, did not affect the changes of the intensity of fast phase. These results suggest that the prompt increase in the intensity of fast phase of ms-DLE at the beginning of transitions to stateⅡ is correlated mainly with the proton gradient released from water oxidation in photosystemⅡ.

  16. Nonlinear Dielectric Spectroscopy as an Indirect Probe of Metabolic Activity in Thylakoid Membrane

    Directory of Open Access Journals (Sweden)

    John H. Miller

    2011-01-01

    Full Text Available Nonlinear dielectric spectroscopy (NDS is a non-invasive probe of cellular metabolic activity with potential application in the development of whole-cell biosensors. However, the mechanism of NDS interaction with metabolic membrane proteins is poorly understood, partly due to the inherent complexity of single cell organisms. Here we use the light-activated electron transport chain of spinach thylakoid membrane as a model system to study how NDS interacts with metabolic activity. We find protein modification, as opposed to membrane pump activity, to be the dominant source of NDS signal change in this system. Potential mechanisms for such protein modifications include reactive oxygen species generation and light-activated phosphorylation.

  17. Functional Implications of Photosystem II Crystal Formation in Photosynthetic Membranes

    NARCIS (Netherlands)

    Tietz, Stefanie; Puthiyaveetil, Sujith; Enlow, Heather M; Yarbrough, Robert; Wood, Magnus; Semchonok, Dmitry A; Lowry, Troy; Li, Zhirong; Jahns, Peter; Boekema, Egbert J; Lenhert, Steven; Niyogi, Krishna K; Kirchhoff, Helmut

    2015-01-01

    The structural organization of proteins in biological membranes can affect their function. Photosynthetic thylakoid membranes in chloroplasts have the remarkable ability to change their supramolecular organization between disordered and semicrystalline states. Although the change to the semicrystall

  18. Proteomic analysis of chloroplast-to-chromoplast transition in tomato reveals metabolic shifts coupled with disrupted thylakoid biogenesis machinery and elevated energy-production components.

    Science.gov (United States)

    Barsan, Cristina; Zouine, Mohamed; Maza, Elie; Bian, Wanping; Egea, Isabel; Rossignol, Michel; Bouyssie, David; Pichereaux, Carole; Purgatto, Eduardo; Bouzayen, Mondher; Latché, Alain; Pech, Jean-Claude

    2012-10-01

    A comparative proteomic approach was performed to identify differentially expressed proteins in plastids at three stages of tomato (Solanum lycopersicum) fruit ripening (mature-green, breaker, red). Stringent curation and processing of the data from three independent replicates identified 1,932 proteins among which 1,529 were quantified by spectral counting. The quantification procedures have been subsequently validated by immunoblot analysis of six proteins representative of distinct metabolic or regulatory pathways. Among the main features of the chloroplast-to-chromoplast transition revealed by the study, chromoplastogenesis appears to be associated with major metabolic shifts: (1) strong decrease in abundance of proteins of light reactions (photosynthesis, Calvin cycle, photorespiration) and carbohydrate metabolism (starch synthesis/degradation), mostly between breaker and red stages and (2) increase in terpenoid biosynthesis (including carotenoids) and stress-response proteins (ascorbate-glutathione cycle, abiotic stress, redox, heat shock). These metabolic shifts are preceded by the accumulation of plastid-encoded acetyl Coenzyme A carboxylase D proteins accounting for the generation of a storage matrix that will accumulate carotenoids. Of particular note is the high abundance of proteins involved in providing energy and in metabolites import. Structural differentiation of the chromoplast is characterized by a sharp and continuous decrease of thylakoid proteins whereas envelope and stroma proteins remain remarkably stable. This is coincident with the disruption of the machinery for thylakoids and photosystem biogenesis (vesicular trafficking, provision of material for thylakoid biosynthesis, photosystems assembly) and the loss of the plastid division machinery. Altogether, the data provide new insights on the chromoplast differentiation process while enriching our knowledge of the plant plastid proteome.

  19. Mollusc-algal chloroplast endosymbiosis. Photosynthesis, thylakoid protein maintenance, and chloroplast gene expression continue for many months in the absence of the algal nucleus.

    Science.gov (United States)

    Green, B J; Li, W Y; Manhart, J R; Fox, T C; Summer, E J; Kennedy, R A; Pierce, S K; Rumpho, M E

    2000-09-01

    Early in its life cycle, the marine mollusc Elysia chlorotica Gould forms an intracellular endosymbiotic association with chloroplasts of the chromophytic alga Vaucheria litorea C. Agardh. As a result, the dark green sea slug can be sustained in culture solely by photoautotrophic CO(2) fixation for at least 9 months if provided with only light and a source of CO(2). Here we demonstrate that the sea slug symbiont chloroplasts maintain photosynthetic oxygen evolution and electron transport activity through photosystems I and II for several months in the absence of any external algal food supply. This activity is correlated to the maintenance of functional levels of chloroplast-encoded photosystem proteins, due in part at least to de novo protein synthesis of chloroplast proteins in the sea slug. Levels of at least one putative algal nuclear encoded protein, a light-harvesting complex protein homolog, were also maintained throughout the 9-month culture period. The chloroplast genome of V. litorea was found to be 119.1 kb, similar to that of other chromophytic algae. Southern analysis and polymerase chain reaction did not detect an algal nuclear genome in the slug, in agreement with earlier microscopic observations. Therefore, the maintenance of photosynthetic activity in the captured chloroplasts is regulated solely by the algal chloroplast and animal nuclear genomes.

  20. Relationships between unsaturation of thylakoid membrane lipids and xanthophyll cycle in rice under chilling and high light

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    To explore the differences of chilling induced sensitivity to photoinhibition between indica and japonica, xanthophyll cycle component, activity of violaxanthin deepoxidase (VDE), and unsaturation of thylakoid membrane lipids were measured. Varieties used were japonica 9516 and indica Shanyou 63 (SY 63).

  1. Ribosomal protein S6 kinase1 coordinates with TOR-Raptor2 to regulate thylakoid membrane biosynthesis in rice.

    Science.gov (United States)

    Sun, Linxiao; Yu, Yonghua; Hu, Weiqin; Min, Qiming; Kang, Huiling; Li, Yilu; Hong, Yue; Wang, Xuemin; Hong, Yueyun

    2016-07-01

    Ribosomal protein S6 kinase (S6K) functions as a key component in the target of rapamycin (TOR) pathway involved in multiple processes in eukaryotes. The role and regulation of TOR-S6K in lipid metabolism remained unknown in plants. Here we provide genetic and pharmacological evidence that TOR-Raptor2-S6K1 is important for thylakoid galactolipid biosynthesis and thylakoid grana modeling in rice (Oryza sativa L.). Genetic suppression of S6K1 caused pale yellow-green leaves, defective thylakoid grana architecture. S6K1 directly interacts with Raptor2, a core component in TOR signaling, and S6K1 activity is regulated by Raptor2 and TOR. Plants with suppressed Raptor2 expression or reduced TOR activity by inhibitors mimicked the S6K1-deficient phenotype. A significant reduction in galactolipid content was found in the s6k1, raptor2 mutant or TOR-inhibited plants, which was accompanied by decreased transcript levels of the set of genes such as lipid phosphate phosphatase α5 (LPPα5), MGDG synthase 1 (MGD1), and DGDG synthase 1 (DGD1) involved in galactolipid synthesis, compared to the control plants. Moreover, loss of LPPα5 exhibited a similar phenotype with pale yellow-green leaves. These results suggest that TOR-Raptor2-S6K1 is important for modulating thylakoid membrane lipid biosynthesis, homeostasis, thus enhancing thylakoid grana architecture and normal photosynthesis ability in rice.

  2. Changes in Unsaturated Levels of Fatty Acids in Thylakoid PSⅡ Membrane Lipids During Chilling-induced Resistance in Rice

    Institute of Scientific and Technical Information of China (English)

    Su-Qin Zhu; Chun-Mei Yu; Xin-Yan Liu; Ben-Hua Ji; De-Mao Jiao

    2007-01-01

    Temperature is one of the abiotic factors limiting growth and productivity of plants. In the present work, the effect of low non-freezing temperature, as an inducer of "chilling resistance", was studied in three cultivars of rice (Oryza sativa L.), japonica cv. 9516 (j-9516), the two parental lines of superhigh-yield hybrid rice between subspecies,Peiai/E32 (ji-PE), and the traditional indica hybrid rice Shanyou 63 (i-SY63). Leaves of chill-treated rice showed chilling-induced resistance, as an increase of their low-temperature tolerance was measured using chlorophyll fluorescence measurements, revealing a change in photosystem Ⅱ (PSⅡ) efficiency. After 5 d of exposure to 11℃ under low light (100 μ mol·m-2·s-1), levels of unsaturated fatty acids in PSⅡ thylakoid membrane lipids decreased during the initial 1-2 d, then increased slowly and reached 99.2%, 95.3% and 90.1% of the initial value (0 d) in j-9516,ji-PE and i-SY63, respectively, on the fifth day. However, under medium light (600 μmol·m-2·s-1), all cultivars experienced similar substantial photoinhibition, which approached steady state levels after a decline in levels of unsaturated fatty acids in PSII thylakoid membrane lipids to about 57.1%, 53.8% and 44.5% of the initial values (0 d) in j-9516,ji-PE and i-SY63 on the fifth day. Under either chilling-induced resistance (the former) or low temperature photoinhibition (the latter) conditions, the changes of other physiological parameters such as D1 protein contents,electron transport activities of PSII (ETA), Fv/Fm, xanthophyl cycle activities expressed by DES (deepoxide state)were consistent with that of levels of unsaturated fatty acids in PSⅡ thylakoid membrane lipids. So there were negative correlations between saturated levels of fatty acids (16:1(3t), 16:0, 18:0), especially the 16:1(3t) fatty acid on thylakoid membrane and other physiological parameters, such as D1 protein contents, ETA and (A+Z)/(A+V+Z). A specific role of

  3. Targeting and biogenesis of transporters and channels in chloroplast envelope membranes: Unsolved questions.

    Science.gov (United States)

    Oh, Young Jun; Hwang, Inhwan

    2015-07-01

    Chloroplasts produce carbohydrates, hormones, vitamins, amino acids, pigments, nucleotides, ATP, and secondary metabolites. Channels and transporters are required for the movement of molecules across the two chloroplast envelope membranes. These transporters and channel proteins are grouped into two different types, including β-barrel proteins and transmembrane-domain (TMD) containing proteins. Most β-barrel proteins are localized at the outer chloroplast membrane, and TMD-containing proteins are localized at the inner chloroplast membrane. Many of these transporters and channels are encoded by nuclear genes; therefore, they have to be imported into chloroplasts after translation on cytosolic ribosomes. These proteins should have specific targeting signals for their final destination in the chloroplast membrane and for assembly into specific complexes. In this review, we summarize recent progress in the identification, functional characterization, and biogenesis of transporters and channels at the chloroplast envelope membranes, and discuss outstanding questions regarding transporter and channel protein biogenesis.

  4. Thylakoids suppress appetite by increasing cholecystokinin resulting in lower food intake and body weight in high-fat fed mice

    DEFF Research Database (Denmark)

    Köhnke, Rickard; Lindqvist, Andreas; Göransson, Nathanael

    2009-01-01

    Thylakoids are membranes isolated from plant chloroplasts which have previously been shown to inhibit pancreatic lipase/colipase catalysed hydrolysis of fat in vitro and induce short-term satiety in vivo. The purpose of the present study was to examine if dietary supplementation of thylakoids could...... compared with the high-fat fed control mice. Reduced serum glucose, serum triglyceride and serum free fatty acid levels were found in the thylakoid-treated animals. The satiety hormone cholecystokinin was elevated, suggesting this hormone mediates satiety. Leptin levels were reduced, reflecting a decreased...

  5. The Prx Q protein of Arabidopsis thaliana is a member of the luminal chloroplast proteome.

    Science.gov (United States)

    Petersson, Ulrika A; Kieselbach, Thomas; García-Cerdán, José G; Schröder, Wolfgang P

    2006-11-13

    Peroxiredoxins have been discovered in many organisms ranging from eubacteria to mammals, and their known biological functions include both oxidant defense and signal transduction. The genome of Arabidopsis thaliana encodes for ten individual peroxiredoxins, of which four are located in the chloroplast. The best-characterized member of the chloroplast peroxiredoxins is 2-Cys Prx that is associated with the stroma side of the thylakoid membrane and is considered to participate in antioxidant defense and protection of photosynthesis. This study addressed the chloroplast peroxiredoxin Prx Q and showed that its subcellular location is the lumen of the thylakoid membrane. To get insight in the biological function of the Prx Q protein of Arabidopsis, the protein levels of the Prx Q protein in thylakoid membranes were studied under different light conditions and oxidative stress. A T-DNA knockout mutant of Prx Q did not show any visible phenotype and had normal photosynthetic performance with a slightly increased oxygen evolving activity.

  6. The Chloroplast Outer Envelope Membrane: The Edge of Light and Excitement

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    The chloroplast is surrounded by a double-membrane envelope at which proteins, ions, and numerous metabolites Including nucleotides, amino acids, fatty acids, and carbohydrates are exchanged between the two aqueous phases, the cytoplasm and the chloroplast stroma. The chloroplast envelope is also the location where the biosynthesis and accumulation of various lipids take place. By contrast to the inner membrane, which contains a number of specific transporters and acts as the permeability barrier, the chloroplast outer membrane has often been considered a passive compartment derived from the phagosomal membrane. However, the presence of galactoglycerolipids and β-barrel membrane proteins support the common origin of the outer membranes of the chloroplast envelope and extant cyanobacteria. Furthermore, recent progress in the field underlines that the chloroplast outer envelope plays important roles not only for translocation of various molecules, but also for regulation of metabolic activities and signaling processes. The chloroplast outer envelope membrane offers various interesting and challenging questions that are relevant to the understanding of organelle biogenesis, plant growth and development, and also membrane biology in general.

  7. Bizonoplast, a unique chloroplast in the epidermal cells of microphylls in the shade plant Selaginella erythropus (Selaginellaceae).

    Science.gov (United States)

    Sheue, Chiou-Rong; Sarafis, Vassilios; Kiew, Ruth; Liu, Ho-Yih; Salino, Alexandre; Kuo-Huang, Ling-Long; Yang, Yuen-Po; Tsai, Chi-Chu; Lin, Chun-Hung; Yong, Jean W H; Ku, Maurice S B

    2007-12-01

    Study of the unique leaf anatomy and chloroplast structure in shade-adapted plants will aid our understanding of how plants use light efficiently in low light environments. Unusual chloroplasts in terms of size and thylakoid membrane stacking have been described previously in several deep-shade plants. In this study, a single giant cup-shaped chloroplast, termed a bizonoplast, was found in the abaxial epidermal cells of the dorsal microphylls and the adaxial epidermal cells of the ventral microphylls in the deep-shade spike moss Selaginella erythropus. Bizonoplasts are dimorphic in ultrastructure: the upper zone is occupied by numerous layers of 2-4 stacked thylakoid membranes while the lower zone contains both unstacked stromal thylakoids and thylakoid lamellae stacked in normal grana structure oriented in different directions. In contrast, other cell types in the microphylls contain chloroplasts with typical structure. This unique chloroplast has not been reported from any other species. The enlargement of epidermal cells into funnel-shaped, photosynthetic cells coupled with specific localization of a large bizonoplast in the lower part of the cells and differential modification in ultrastructure within the chloroplast may allow the plant to better adapt to low light. Further experiments are required to determine whether this shade-adapted organism derives any evolutionary or ecophysiological fitness from these unique chloroplasts.

  8. Both phototropin 1 and 2 localize on the chloroplast outer membrane with distinct localization activity.

    Science.gov (United States)

    Kong, Sam-Geun; Suetsugu, Noriyuki; Kikuchi, Shingo; Nakai, Masato; Nagatani, Akira; Wada, Masamitsu

    2013-01-01

    Chloroplasts change their position to adapt cellular activities to fluctuating environmental light conditions. Phototropins (phot1 and phot2 in Arabidopsis) are plant-specific blue light photoreceptors that perceive changes in light intensity and direction, and mediate actin-based chloroplast photorelocation movements. Both phot1 and phot2 regulate the chloroplast accumulation response, while phot2 is mostly responsible for the regulation of the avoidance response. Although it has been widely accepted that distinct intracellular localizations of phototropins are implicated in the specificity, the mechanism underlying the phot2-specific avoidance response has remained elusive. In this study, we examined the relationship of the phot2 localization pattern to the chloroplast photorelocation movement. First, the fusion of a nuclear localization signal with phot2, which effectively reduced the amount of phot2 in the cytoplasm, retained the activity for both the accumulation and avoidance responses, indicating that membrane-localized phot2 but not cytoplasmic phot2 is functional to mediate the responses. Importantly, some fractions of phot2, and of phot1 to a lesser extent, were localized on the chloroplast outer membrane. Moreover, the deletion of the C-terminal region of phot2, which was previously shown to be defective in blue light-induced Golgi localization and avoidance response, affected the localization pattern on the chloroplast outer membrane. Taken together, these results suggest that dynamic phot2 trafficking from the plasma membrane to the Golgi apparatus and the chloroplast outer membrane might be involved in the avoidance response.

  9. Isolation and Suborganellar Fractionation of Arabidopsis Chloroplasts.

    Science.gov (United States)

    Flores-Pérez, Úrsula; Jarvis, Paul

    2017-01-01

    Chloroplasts are structurally complex organelles containing ~2000-3000 proteins. They are delimited by a double membrane system or envelope, have an inner aqueous compartment called the stroma, and possess a second internal membrane system called the thylakoids. Thus, determining the suborganellar location of a chloroplast protein is vital to understanding or verifying its function. One way in which protein localization can be addressed is through fractionation. Here we present two rapid and simple methods that may be applied sequentially on the same day: (a) The isolation of intact chloroplasts from Arabidopsis thaliana plants that may be used directly (e.g., for functional studies such as protein import analysis), or for further processing as follows; (b) separation of isolated chloroplasts into three suborganellar fractions (envelope membranes, a soluble fraction containing stromal proteins, and the thylakoids). These methods are routinely used in our laboratory, and they provide a good yield of isolated chloroplasts and suborganellar fractions that can be used for various downstream applications.

  10. [Fluorescence used to investigate the sensitivity of spinach chloroplast membrane to low intensity electromagnetic radiation].

    Science.gov (United States)

    Xi, Gang; Yang, Yun-Jing; Lu, Hong

    2009-07-01

    A system for studying biological effect of radio frequency electromagnetic field was developed. The system can form an area where electromagnetic wave with large frequency range is well distributed. The strength of electromagnetic wave was measured easily. Electromagnetic wave in the system did not have effect on environment. The sensitivity of spinach chloroplast membrane to low intensity electromagnetic radiation of 300 MHz under power density of 5 mW x cm(-2) was studied by the spectral analysis method of fluorescence of 8-anilino-1-naphthalene-sulfonic acid (ANS) and the changes in chlorophyll a (Chla) fluorescence parameters of spinach chloroplast membrane. The result showed that the position of spectrum of ANS fluorescence of spinach chloroplast membrane did not change, but the intensity of ANS fluorescence was obviously increased under the action of electromagnetic radiation with power density of 1-5 mW x cm(-2). There was an increase in the intensity of ANS fluorescence with the increase in electromagnetic radiation. The increase of ANS fluorescence of spinach chloroplast membrane showed that low level electromagnetic field induced the decrease in fluidity of chloroplast membrane compared with control experiment. The cause of the change in the fluidity could be related to the polarization of chloroplast membrane under the electromagnetic field. The analysis of Chla fluorescence parameters of spinach chloroplast membrane indicated that low level electromagnetic field of 300 MHz made the fluorescence parameters F0 and F(VI/)F(V) decrease, and F(V)/Fo, Fv/F(m) and deltaF(V)/T increase. It was showed that low level electromagnetic field caused the change of non-active center of photosystem II of spinach chloroplast membrane to active center and the increase in potential active and photochemical efficiency of PSII, and promoted the transmit process of electron in photosynthesis of chloroplast membrane of photosynthesis cell in spinach leaf. The study confirmed

  11. Evolution and targeting of Omp85 homologs in the chloroplast outer envelope membrane

    Directory of Open Access Journals (Sweden)

    Philip Michael Day

    2014-10-01

    Full Text Available Translocon at the outer-envelope-membrane of chloroplasts 75 (Toc75 is the core component of the chloroplast protein import machinery. It belongs to the Omp85 family whose members exist in various Gram-negative bacteria, mitochondria and chloroplasts of eukaryotes. Chloroplasts of Viridiplantae contain another Omp85 homolog called outer envelope protein 80 (OEP80, whose exact function is unknown. In addition, the Arabidopsis thaliana genome encodes truncated forms of Toc75 and OEP80. Multiple studies have shown a common origin of the Omp85 homologs of cyanobacteria and chloroplasts but their results about evolutionary relationships among cyanobacterial Omp85 (cyanoOmp85, Toc75 and OEP80 are inconsistent. The bipartite targeting sequence-dependent sorting of Toc75 has been demonstrated but the targeting mechanisms of other chloroplast Omp85 homologs remain largely unexplored. This study was aimed to address these unresolved issues in order to further our understanding of chloroplast evolution. Sequence alignments and recently determined structures of bacterial Omp85 homologs were used to predict structures of chloroplast Omp85 homologs. The results enabled us to identify amino acid residues that may indicate functional divergence of Toc75 from cyanoOmp85 and OEP80. Phylogenetic analyses using Omp85 homologs from various cyanobacteria and chloroplasts provided strong support for the grouping of Toc75 and OEP80 sister to cyanoOmp85. However, this support was diminished when the analysis included Omp85 homologs from other bacteria and mitochondria. Finally, results of import assays using isolated chloroplasts support outer membrane localization of OEP80tr and indicate that OEP80 may carry a cleavable targeting sequence.

  12. Fingerprinting the macro-organisation of pigment-protein complexes in plant thylakoid membranes in vivo by circular-dichroism spectroscopy.

    Science.gov (United States)

    Tóth, Tünde N; Rai, Neha; Solymosi, Katalin; Zsiros, Ottó; Schröder, Wolfgang P; Garab, Győző; van Amerongen, Herbert; Horton, Peter; Kovács, László

    2016-09-01

    Macro-organisation of the protein complexes in plant thylakoid membranes plays important roles in the regulation and fine-tuning of photosynthetic activity. These delicate structures might, however, undergo substantial changes during isolating the thylakoid membranes or during sample preparations, e.g., for electron microscopy. Circular-dichroism (CD) spectroscopy is a non-invasive technique which can thus be used on intact samples. Via excitonic and psi-type CD bands, respectively, it carries information on short-range excitonic pigment-pigment interactions and the macro-organisation (chiral macrodomains) of pigment-protein complexes (psi, polymer or salt-induced). In order to obtain more specific information on the origin of the major psi-type CD bands, at around (+)506, (-)674 and (+)690nm, we fingerprinted detached leaves and isolated thylakoid membranes of wild-type and mutant plants and also tested the effects of different environmental conditions in vivo. We show that (i) the chiral macrodomains disassemble upon mild detergent treatments, but not after crosslinking the protein complexes; (ii) in different wild-type leaves of dicotyledonous and monocotyledonous angiosperms the CD features are quite robust, displaying very similar excitonic and psi-type bands, suggesting similar protein composition and (macro-) organisation of photosystem II (PSII) supercomplexes in the grana; (iii) the main positive psi-type bands depend on light-harvesting protein II contents of the membranes; (iv) the (+)506nm band appears only in the presence of PSII-LHCII supercomplexes and does not depend on the xanthophyll composition of the membranes. Hence, CD spectroscopy can be used to detect different macro-domains in the thylakoid membranes with different outer antenna compositions in vivo.

  13. Supplementation by thylakoids to a high carbohydrate meal decreases feelings of hunger, elevates CCK levels and prevents postprandial hypoglycaemia in overweight women.

    Science.gov (United States)

    Stenblom, Eva-Lena; Montelius, Caroline; Östbring, Karolina; Håkansson, Maria; Nilsson, Sofia; Rehfeld, Jens F; Erlanson-Albertsson, Charlotte

    2013-09-01

    Thylakoids are chlorophyll-containing membranes in chloroplasts that have been isolated from green leaves. It has been previously shown that thylakoids supplemented with a high-fat meal can affect cholecystokinin (CCK), ghrelin, insulin and blood lipids in humans, and can act to suppress food intake and prevent body weight gain in rodents. This study investigates the addition of thylakoids to a high carbohydrate meal and its effects upon hunger motivation and fullness, and the levels of glucose, insulin, CCK, ghrelin and tumour necrosis factor (TNF)-alpha in overweight women. Twenty moderately overweight female subjects received test meals on three different occasions; two thylakoid enriched and one control, separated by 1 week. The test meals consisted of a high carbohydrate Swedish breakfast, with or without addition of thylakoids. Blood samples and VAS-questionnaires were evaluated over a 4-h period. Addition of thylakoids suppressed hunger motivation and increased secretion of CCK from 180 min, and prevented postprandial hypoglycaemia from 90 min following food intake. These effects indicate that thylakoids may intensify signals of satiety. This study therefore suggests that the dietary addition of thylakoids could aid efforts to reduce food intake and prevent compensational eating later in the day, which may help to reduce body weight over time.

  14. Biotin Carboxyl Carrier Protein in Barley Chloroplast Membranes

    DEFF Research Database (Denmark)

    Kannangara, C. G.; Jense, C J

    1975-01-01

    Biotin localized in barley chloroplast lamellae is covalently bound to a single protein with an approximate molecular weight of 21000. It contains one mole of biotin per mole of protein and functions as a carboxyl carrier in the acetyl-CoA carboxylase reaction. The protein was obtained by solubil......Biotin localized in barley chloroplast lamellae is covalently bound to a single protein with an approximate molecular weight of 21000. It contains one mole of biotin per mole of protein and functions as a carboxyl carrier in the acetyl-CoA carboxylase reaction. The protein was obtained...

  15. A fluorescence detected magnetic resonance investigation of the carotenoid triplet states associated with Photosystem II of isolated spinach thylakoid membranes

    CERN Document Server

    Santabarbara, S; Carbonera, D; Heathcote, P

    2005-01-01

    The carotenoid triplet populations associated with the fluorescence emission chlorophyll forms of Photosystem II have been investigated in isolated spinach thylakoid membranes by means of fluorescence detected magnetic resonance in zero field (FDMR). The spectra collected in the 680-690 nm emission range, have been fitted by a global analysis procedure. At least five different carotenoid triplet states coupled to the terminal emitting chlorophyll forms of PS II, peaking at 682 nm, 687 nm and 692 nm, have been characterised. The triplets associated with the outer antenna emission forms, at 682 nm, have zero field splitting parameters D = 0.0385 cm/sup -1/, E = 0.00367 cm/sup -1/; D = 0.0404 cm/sup -1/, E = 0.00379 cm/sup -1/ and D = 0.0386 cm/sup -1/, E = 0.00406 cm/sup -1/ which are very similar to those previously reported for the xanthophylls of the isolated LHC II complex. Therefore the FDMR spectra recorded in this work provide insights into the organisation of the LHC II complex in the unperturbed enviro...

  16. Computer modeling of electron and proton transport in chloroplasts.

    Science.gov (United States)

    Tikhonov, Alexander N; Vershubskii, Alexey V

    2014-07-01

    Photosynthesis is one of the most important biological processes in biosphere, which provides production of organic substances from atmospheric CO2 and water at expense of solar energy. In this review, we contemplate computer models of oxygenic photosynthesis in the context of feedback regulation of photosynthetic electron transport in chloroplasts, the energy-transducing organelles of the plant cell. We start with a brief overview of electron and proton transport processes in chloroplasts coupled to ATP synthesis and consider basic regulatory mechanisms of oxygenic photosynthesis. General approaches to computer simulation of photosynthetic processes are considered, including the random walk models of plastoquinone diffusion in thylakoid membranes and deterministic approach to modeling electron transport in chloroplasts based on the mass action law. Then we focus on a kinetic model of oxygenic photosynthesis that includes key stages of the linear electron transport, alternative pathways of electron transfer around photosystem I (PSI), transmembrane proton transport and ATP synthesis in chloroplasts. This model includes different regulatory processes: pH-dependent control of the intersystem electron transport, down-regulation of photosystem II (PSII) activity (non-photochemical quenching), the light-induced activation of the Bassham-Benson-Calvin (BBC) cycle. The model correctly describes pH-dependent feedback control of electron transport in chloroplasts and adequately reproduces a variety of experimental data on induction events observed under different experimental conditions in intact chloroplasts (variations of CO2 and O2 concentrations in atmosphere), including a complex kinetics of P700 (primary electron donor in PSI) photooxidation, CO2 consumption in the BBC cycle, and photorespiration. Finally, we describe diffusion-controlled photosynthetic processes in chloroplasts within the framework of the model that takes into account complex architecture of

  17. Immunoelectron microscopy for locating calvin cycle enzymes in the thylakoids of synechocystis 6803.

    Science.gov (United States)

    Agarwal, Rachna; Ortleb, Stefan; Sainis, Jayashree Krishna; Melzer, Michael

    2009-01-01

    Unicellular cyanobacteria Synechocystis 6803 were fixed using high-pressure freezing (HPF) and freeze substitution without any chemical cross-linkers. Immunoelectron microscopy of these cells showed that five sequential enzymes of the Calvin cycle (phosphoriboisomerase, phosphoribulokinase, ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO), 3-phosphoglyceratekinase and glyceraldehyde-3-phosphate dehydrogenase) and the catalytic portion of the chloroplast H+-ATP synthase (CF1) are located adjacent to the thylakoid membranes. Cell-free extracts of Synechocystis were processed by ultracentrifugation to isolate thylakoid fractions sedimenting at 40,000, 90,000, and 150,000 g. Among these, the 150,000-g fraction showed the highest linked activity of the above five sequential Calvin cycle enzymes and also the highest coordinated activity of light and dark reactions as assessed by ribose-5-phosphate (R-5-P) +ADP dependent CO2 fixation. Immunogold labeling of this membrane fraction confirmed the presence of the above five enzymes as well as the catalytic portion of the CF1 ATP synthase. Notably, the protein A-gold labeling of the thylakoids was observed without use of chemical cross-linkers and in spite of the normal washing steps used during standard immunolabeling. The results showed that soluble Calvin cycle enzymes might be organized along the thylakoid membranes.

  18. Immunoelectron Microscopy for Locating Calvin Cycle Enzymes in the Thylakoids of Synechocystis 6803

    Institute of Scientific and Technical Information of China (English)

    Rachna Agarwal; Stefan Ortleb; Jayashree Krishna Saini; Michael Melzer

    2009-01-01

    Unicellular cyanobacteria Synechocystis 6803 were fixed using high-pressure freezing (HPF) and freeze substitution without any chemical cross-linkers. Immunoelectron microscopy of these cells showed that five sequential enzymes of the Calvin cycle (phosphoriboisomerase, phosphoribulokinase, ribulose-l,5-bisphosphate carboxylase/oxygenase (RuBisCO), 3-phosphoglyceratekinase and glyceraldehyde-3-phosphate dehydrogenase) and the catalytic portion of the chloroplast H+-ATP synthase (CF1) are located adjacent to the thylakoid membranes. Cell-free extracts of Synechocystis were processed by ultracentrifugation to isolate thylakoid fractions sedimenting at 40 000, 90 000, and 150 000 g.Among these, the 150 000-g fraction showed the highest linked activity of the above five sequential Calvin cycle enzymes and also the highest coordinated activity of light and dark reactions as assessed by ribose-5-phosphate (R-5-P) +ADP dependent CO2 fixation. Immunogold labeling of this membrane fraction confirmed the presence of the above five enzymes as well as the catalytic portion of the CF1 ATP synthase. Notably, the protein A-gold labeling of the thylakoids was observed without use of chemical cross-linkers and in spite of the normal washing steps used during standard immunolabeling. The results showed that soluble Calvin cycle enzymes might be organized along the thylakoid membranes.

  19. Research on the ultrafast fluorescence property of thylakoid membranes of the wild-type and mutant rice

    Institute of Scientific and Technical Information of China (English)

    任兆玉; 许晓明; 王水才; 辛越勇; 贺俊芳; 侯洵

    2003-01-01

    A high yielding rice variety mutant (Oryza sativa L., Zhenhui 249) with low chlorophyll b (Chl b) has been discovered in natural fields. It has a quality character controlled by a pair of recessive genes (nuclear gene). The partial loss of Chl b in content affects the efficiency of light harvest in a light harvest complex (LHC), thus producing the difference of the exciting energy transfer and the efficiency of photochemistry conversion between the mutant and wild-type rice in photosynthetic unit. The efficiency of utilizing light energy is higher in the mutant than that in the wildtype rice relatively. For further discussion of the above-mentioned difference and learning about the mechanism of the increase in the photochemical efficiency of the mutant, the pico-second resolution fluorescence spectrum measurement with delay-frame-scanning single photon counting technique is adopted. Thylakoid membranes of the mutant and the wild-type rice are excited by an Ar+ laser with a pulse width of 120ps, repetition rate of 4MHz and wavelength of 514nm. Compared with the time and spectrum property of exciting fluorescence, conclusions of those ultrafast dynamic experiments are: 1) The speeds of the exciting energy transferred in photo-system I are faster than that in photo-system II in both samples. 2) The speeds of the exciting energy transfer of mutant sample are faster than those of the wildtype. This might be one of the major reasons why the efficiency of photosynthesis is higher in mutant than that in the wild-type rice.

  20. The synthesis of NPQ-effective zeaxanthin depends on the presence of a transmembrane proton gradient and a slightly basic stromal side of the thylakoid membrane.

    Science.gov (United States)

    Goss, Reimund; Opitz, Christian; Lepetit, Bernard; Wilhelm, Christian

    2008-11-01

    In the present study we address the question which factors during the synthesis of zeaxanthin determine its capacity to act as a non-photochemical quencher of chlorophyll fluorescence. Our results show that zeaxanthin has to be synthesized in the presence of a transmembrane proton gradient. However, it is not essential that the proton gradient is generated by the light-driven electron transport. NPQ-effective zeaxanthin can also be formed by an artificial proton gradient in the dark due to ATP hydrolysis. Zeaxanthin that is synthesized in the dark in the absence of a proton gradient by the low pH-dependent activation of violaxanthin de-epoxidase is not able to induce NPQ. The second important factor during the synthesis of zeaxanthin is the pH-value of the stromal side of the thylakoid membrane. Here we show that the stromal side has to be neutral or slightly basic in order to generate zeaxanthin which is able to induce NPQ. Thylakoid membranes in reaction medium pH 5.2, which experience low pH-values on both sides of the membrane, are unable to generate NPQ-effective zeaxanthin, even in the presence of an additional light-driven proton gradient. Analysing the pigment contents of purified photosystem II light-harvesting complexes we are further able to show that the NPQ ineffectiveness of zeaxanthin formed in the absence of a proton gradient is not caused by changes in its rebinding to the light-harvesting proteins. Purified monomeric and trimeric light-harvesting complexes contain comparable amounts of zeaxanthin when they are isolated from thylakoid membranes enriched in either NPQ-effective or ineffective zeaxanthin.

  1. Is chloroplast import of photosynthesis proteins facilitated by an actin-TOC-TIC-VIPP1 complex?

    Science.gov (United States)

    Jouhet, Juliette; Gray, John C

    2009-10-01

    Actin filaments are major components of the cytoskeleton that interact with chloroplast envelope membranes to allow chloroplast positioning and movement, stromule mobility and gravitropism perception. We recently reported that Toc159, a component of the TOC complex of the chloroplast protein import apparatus, interacts directly with actin. The interaction of Toc159 and actin was identified by co-immunoprecipitation and co-sedimentation experiments with detergent-solubilised pea chloroplast envelope membranes. In addition, many of the components of the TOC-TIC protein import apparatus and VIPP1 (vesicle-inducing protein in plastids 1) were identified by mass spectroscopy in the material co-immunoprecipitated with antibodies to actin. Toc159 is the receptor for the import of photosynthesis proteins and VIPP1 is involved in thylakoid membrane formation by inducing vesicle formation from the chloroplast inner envelope membrane, suggesting we may have identified an actin-TOC-TIC-VIPP1 complex that may provide a means of channeling cytosolic preproteins to the thylakoid membrane. The interaction of Toc159 with actin may facilitate exchange between the putative soluble and membrane forms of Toc159 and promote the interaction of cytosolic preproteins with the TOC complex.

  2. The puzzle of chloroplast vesicle transport – involvement of GTPases

    Directory of Open Access Journals (Sweden)

    Sazzad eKarim

    2014-09-01

    Full Text Available In the cytosol of plant cells vesicle transport occurs via secretory pathways among the endoplasmic reticulum (ER network, Golgi bodies, secretory granules, endosome and plasma membrane. Three systems transfer lipids, proteins and other important molecules through aqueous spaces to membrane-enclosed compartments, via vesicles that bud from donor membranes, being coated and uncoated before tethered and fused with acceptor membranes. In addition, molecular, biochemical and ultrastructural evidence indicates presence of a vesicle transport system in chloroplasts. Little is known about the protein components of this system. However, as chloroplasts harbour the photosynthetic apparatus that ultimately supports most organisms on the planet, close attention to their pathways is warranted. This may also reveal novel diversification and/or distinct solutions to the problems posed by the targeted intra-cellular trafficking of important molecules. To date two homologues to well-known yeast cytosolic vesicle transport proteins, CPSAR1 and CPRabA5e, have been shown to have roles in chloroplast vesicle transport, both being GTPases. Bioinformatic data indicate that several homologues of cytosolic vesicle transport system components are putatively chloroplast-localized and in addition other proteins have been implicated to participate in chloroplast vesicle transport, including vesicle-inducing protein in plastids 1 (VIPP1, thylakoid formation 1 (THF1, snowy cotyledon 2/cotyledon chloroplast biogenesis factor (SCO2/CYO1, curvature thylakoid 1 (CURT1 proteins, and a dynamin like GTPase FZO-like (FZL protein. Several putative potential cargo proteins have also been identified, including building blocks of the photosynthetic apparatus. Here we discuss details of the largely unknown putative chloroplast vesicle transport system, focusing on GTPase-related components.

  3. Quality control of Photosystem II: the mechanisms for avoidance and tolerance of light and heat stresses are closely linked to membrane fluidity of the thylakoids

    Directory of Open Access Journals (Sweden)

    Yasusi Yamamoto

    2016-08-01

    Full Text Available When oxygenic photosynthetic organisms are exposed to excessive light and/or heat, Photosystem II is damaged and electron transport is blocked. In these events, reactive oxygen species, endogenous radicals and lipid peroxidation products generated by photochemical reaction and/or heat cause the damage. Regarding light stress, plants first dissipate excessive light energy captured by light-harvesting chlorophyll protein complexes as heat to avoid the hazards, but once light stress is unavoidable, they tolerate the stress by concentrating damage in a particular protein in photosystem II, i.e. the reaction-center binding D1 protein of Photosystem II. The damaged D1 is removed by specific proteases and replaced with a new copy produced through de novo synthesis (reversible photoinhibition. When light intensity becomes extremely high, irreversible aggregation of D1 occurs and thereby D1 turnover is prevented. Once the aggregated products accumulate in Photosystem II complexes, removal of them by proteases is difficult, and irreversible inhibition of Photosystem II takes place (irreversible photoinhibition. Important is that various aspects of both the reversible and irreversible photoinhibition are highly dependent on the membrane fluidity of the thylakoids. Heat stress-induced inactivation of photosystem II is an irreversible process, which may be also affected by the fluidity of the thylakoid membranes. Here I describe why the membrane fluidity is a key to regulate the avoidance and tolerance of Photosystem II on environmental stresses.

  4. RECOVERY ACT - Thylakoid Assembly and Folded Protein Transport by the Tat Pathway

    Energy Technology Data Exchange (ETDEWEB)

    Dabney-Smith, Carole [Miami Univ., Oxford, OH (United States)

    2016-07-18

    Assembly of functional photosystems complete with necessary intrinsic (membrane-bound) and extrinsic proteins requires the function of at least 3 protein transport pathways in thylakoid membranes. Our research focuses on one of those pathways, a unique and essential protein transport pathway found in the chloroplasts of plants, bacteria, and some archaebacteria, the Twin arginine translocation (Tat) system. The chloroplast Tat (cpTat) system is thought to be responsible for the proper location of ~50% of thylakoid lumen proteins, several of which are necessary for proper photosystem assembly, maintenance, and function. Specifically, cpTat systems are unique because they transport fully folded and assembled proteins across ion tight membranes using only three membrane components, Tha4, Hcf106, and cpTatC, and the protonmotive force generated by photosynthesis. Despite the importance of the cpTat system in plants, the mechanism of transport of a folded precursor is not well known. Our long-term goal is to investigate the role protein transport systems have on organelle biogenesis, particularly the assembly of membrane protein complexes in thylakoids of chloroplasts. The objective of this proposal is to correlate structural changes in the membrane-bound cpTat component, Tha4, to the mechanism of translocation of folded-precursor substrates across the membrane bilayer by using a cysteine accessibility and crosslinking approach. Our central hypothesis is that the precursor passes through a proteinaceous pore of assembled Tha4 protomers that have undergone a conformational or topological change in response to transport. This research is predicated upon the observations that Tha4 exists in molar excess in the membrane relative to the other cpTat components; its regulated assembly to the precursor-bound receptor; and our data showing oligomerization of Tha4 into very large complexes in response to transport. Our rationale for these studies is that understanding cp

  5. Comparative proteomics of chloroplasts envelopes from bundle sheath and mesophyll chloroplasts reveals novel membrane proteins with a possible role in C4-related metabolite fluxes and development.

    Directory of Open Access Journals (Sweden)

    Kalpana eManandhar-Shrestha

    2013-03-01

    Full Text Available As the world population grows, our need for food increases drastically. Limited amounts of arable land lead to a competition between food and fuel crops, while changes in the global climate may impact future crop yields. Thus, a second green revolution will need a better understanding of the processes essential for plant growth and development. One approach toward the solution of this problem is to better understand regulatory and transport processes in C4 plants. C4 plants display an up to 10-fold higher apparent CO2 assimilation and higher yields while maintaining high water use efficiency. This requires differential regulation of mesophyll (M and bundle sheath (BS chloroplast development as well as higher metabolic fluxes of photosynthetic intermediates between cells and across chloroplast envelopes. While previous analyses of overall chloroplast membranes have yielded significant insight, our comparative proteomics approach using enriched BS and M chloroplast envelopes of Zea mays allowed us to identify 37 proteins of unknown function that have not been seen in these earlier studies. We identified 280 proteins, 84% of which are known/predicted to be present in chloroplasts (cp. 74% have a known or predicted membrane association. 21 membrane proteins were 2-15 times more abundant in BS cells, while 36 proteins were more abundant in M cp envelopes. These proteins could represent additional candidates of proteins essential for development or metabolite transport processes in C4 plants. RT-PCR confirmed differential expression of thirteen candidate genes. Cp association was confirmed using GFP labeling. Genes for a PIC-like protein and an ER-AP-like protein show an early transient increase in gene expression during the transition to light. In addition, PIC gene expression is increased in the immature part of the leaf and was lower in the fully developed parts of the leaf, suggesting a need for/incorporation of the protein during chloroplast

  6. Specific and efficient targeting of cyanobacterial bicarbonate transporters to the inner envelope membrane of chloroplasts in Arabidopsis

    Directory of Open Access Journals (Sweden)

    Susumu eUehara

    2016-02-01

    Full Text Available Installation of cyanobacterial bicarbonate transporters to the inner envelope membrane (IEM of chloroplasts in C3 plants has been thought to improve photosynthetic performance. However, the method to deliver cyanobacterial bicarbonate transporters to the chloroplast IEM remains to be established. In this study, we provide evidence that the cyanobacterial bicarbonate transporters, BicA and SbtA, can be specifically installed into the chloroplast IEM using the chloroplast IEM targeting signal in conjunction with the transit peptide. We fused the transit peptide and the mature portion of Cor413im1, whose targeting mechanism to the IEM has been characterized in detail, to either BicA or SbtA isolated from Synechocystis sp. PCC6803. Among the seven chimeric constructs tested, we confirmed that four chimeric bicarbonate transporters, designated as BicAI, BicAII, SbtAII, and SbtAIII, were expressed in Arabidopsis. Furthermore, these chimeric transporters were specifically targeted to the chloroplast IEM. They were also resistant to alkaline extraction but can be solubilized by Triton X-100, indicating that they are integral membrane proteins in the chloroplast IEM. One of the transporters, BicA, could reside in the chloroplast IEM even after removal of the IEM targeting signal. Taken together, our results indicate that the addition of IEM targeting signal, as well as the transit peptide, to bicarbonate transporters allows us to efficiently target nuclear-encoded chimeric bicarbonate transporters to the chloroplast IEM.

  7. Contrasting effect of dark-chilling on chloroplast structure and arrangement of chlorophyll-protein complexes in pea and tomato : Plants with a different susceptibility to non-freezing temperature

    NARCIS (Netherlands)

    Garstka, Maciej; Venema, Jan Henk; Rumak, Izabela; Gieczewska, Katarzyna; Rosiak, Malgorzata; Koziol-Lipinska, Joanna; Kierdaszuk, Borys; Vredenberg, Wim J.; Mostowska, Agnieszka

    2007-01-01

    The effect of dark-chilling and subsequent photoactivation on chloroplast structure and arrangements of chlorophyll-protein complexes in thylakoid membranes was studied in chilling-tolerant (CT) pea and in chilling-sensitive (CS) tomato. Dark-chilling did not influence chlorophyll content and Chl a/

  8. Characterization of chloroplast phosphoproteins controlling manganese use efficiency using quantitative proteomics

    DEFF Research Database (Denmark)

    Petersen, Jørgen; Sprenger, Richard Remko; Rogowska-Wrzesinska, Adelina

    Manganese is important for molecular functions in plants, i.e. as a co-factor in enzymes and in the oxygen evolving complex of photosystem II, located like most of the photosynthetic machinery, in the thylakoid membranes of chloroplasts. Soils that lack plant available micronutrients such as mang...... better protein solubilization. Phosphopeptides will be purified using titaniumdioxide beads. Peptides will be analyzed and label free quantified using a 1D-LC UPLC system coupled to an Orbitrap Velos mass spectrometer....

  9. A C-terminal membrane association domain of phototropin 2 is necessary for chloroplast movement.

    Science.gov (United States)

    Kong, Sam-Geun; Kagawa, Takatoshi; Wada, Masamitsu; Nagatani, Akira

    2013-01-01

    Phototropins (phot1 and phot2), plant-specific blue light receptor kinases, mediate a range of physiological responses in Arabidopsis, including phototropism, chloroplast photorelocation movement, stomatal opening and leaf flattening. Phototropins consist of two photoreceptive domains at their N-terminus, LOV1 (light, oxygen or voltage 1) and LOV2, and a serine/threonine kinase domain at their C-terminus. Here, we determined the molecular moiety for the membrane association of phototropins using the yeast CytoTrap and Arabidopsis protoplast systems. We then examined the physiological significance of the membrane association of phototropins. This detailed study with serial deletions narrowed down the association domain to a relatively small part of the C-terminal domain of phototropin. The functional analysis of phot2 deletion mutants in the phot2-deficient Adiantum and Arabidopsis mutants revealed that the ability to mediate the chloroplast avoidance response correlated well with phot2's membrane association, especially with the Golgi apparatus. Taken together, our data suggest that a small part of the C-terminal domain of phototropins is necessary not only for membrane association but also for the physiological activities that elicit phototropin-specific responses.

  10. The KAC family of kinesin-like proteins is essential for the association of chloroplasts with the plasma membrane in land plants.

    Science.gov (United States)

    Suetsugu, Noriyuki; Sato, Yoshikatsu; Tsuboi, Hidenori; Kasahara, Masahiro; Imaizumi, Takato; Kagawa, Takatoshi; Hiwatashi, Yuji; Hasebe, Mitsuyasu; Wada, Masamitsu

    2012-11-01

    Chloroplasts require association with the plasma membrane for movement in response to light and for appropriate positioning within the cell to capture photosynthetic light efficiently. In Arabidopsis, CHLOROPLAST UNUSUAL POSITIONING 1 (CHUP1), KINESIN-LIKE PROTEIN FOR ACTIN-BASED CHLOROPLAST MOVEMENT 1 (KAC1) and KAC2 are required for both the proper movement of chloroplasts and the association of chloroplasts with the plasma membrane, through the reorganization of short actin filaments located on the periphery of the chloroplasts. Here, we show that KAC and CHUP1 orthologs (AcKAC1, AcCHUP1A and AcCHUP1B, and PpKAC1 and PpKAC2) play important roles in chloroplast positioning in the fern Adiantum capillus-veneris and the moss Physcomitrella patens. The knockdown of AcKAC1 and two AcCHUP1 genes induced the aggregation of chloroplasts around the nucleus. Analyses of A. capillus-veneris mutants containing perinuclear-aggregated chloroplasts confirmed that AcKAC1 is required for chloroplast-plasma membrane association. In addition, P. patens lines in which two KAC genes had been knocked out showed an aggregated chloroplast phenotype similar to that of the fern kac1 mutants. These results indicate that chloroplast positioning and movement are mediated through the activities of KAC and CHUP1 proteins, which are conserved in land plants.

  11. Spectroelectrochemistry of P700 in native photosystem I particles and diethyl ether-treated thylakoid membranes from spinach and Thermosynechococcus elongatus.

    Science.gov (United States)

    Zhang, Yanrong; Nakamura, Akimasa; Kuroiwa, Yoshinori; Kato, Yuki; Watanabe, Tadashi

    2008-04-02

    The redox potentials (E(composite function')) of P700 in intact and diethyl ether-treated thylakoid membranes as well as native photosystem (PS) I particles from spinach and Thermosynechococcus elongatus have been measured by a spectroelectrochemistry with an error range of +/-2-3 mV. Stepwise removal of antenna pigments by ether treatment caused distinct shifts of the E( composite function') value with increasing degree of water saturation in ether; negatively from +471 to +428 mV for spinach, but positively from +423 to +436 mV for T. elongatus. Such a contrasting behavior is discussed by invoking the mode of action of ether on the microenvironments around P700.

  12. Expression of fungal cutinase and swollenin in tobacco chloroplasts reveals novel enzyme functions and/or substrates.

    Directory of Open Access Journals (Sweden)

    Dheeraj Verma

    Full Text Available In order to produce low-cost biomass hydrolyzing enzymes, transplastomic lines were generated that expressed cutinase or swollenin within chloroplasts. While swollenin expressing plants were homoplasmic, cutinase transplastomic lines remained heteroplasmic. Both transplastomic lines showed interesting modifications in their phenotype, chloroplast structure, and functions. Ultrastructural analysis of chloroplasts from cutinase- and swollenin-expressing plants did not show typical lens shape and granal stacks. But, their thylakoid membranes showed unique scroll like structures and chloroplast envelope displayed protrusions, stretching into the cytoplasm. Unusual honeycomb structures typically observed in etioplasts were observed in mature chloroplasts expressing swollenin. Treatment of cotton fiber with chloroplast-derived swollenin showed enlarged segments and the intertwined inner fibers were irreversibly unwound and fully opened up due to expansin activity of swollenin, causing disruption of hydrogen bonds in cellulose fibers. Cutinase transplastomic plants showed esterase and lipase activity, while swollenin transplastomic lines lacked such enzyme activities. Higher plants contain two major galactolipids, monogalactosyldiacylglycerol (MGDG and digalactosyldiacylglycerol (DGDG, in their chloroplast thylakoid membranes that play distinct roles in their structural organization. Surprisingly, purified cutinase effectively hydrolyzed DGDG to MGDG, showing alpha galactosidase activity. Such hydrolysis resulted in unstacking of granal thylakoids in chloroplasts and other structural changes. These results demonstrate DGDG as novel substrate and function for cutinase. Both MGDG and DGDG were reduced up to 47.7% and 39.7% in cutinase and 68.5% and 67.5% in swollenin expressing plants. Novel properties and functions of both enzymes reported here for the first time should lead to better understanding and enhanced biomass hydrolysis.

  13. Ultrastructural changes in chloroplasts of mesophyll cells of chlorotic and prematurely yellowed leaves of Betula pendula Rothr

    Directory of Open Access Journals (Sweden)

    Krystyna Przybył

    2011-04-01

    Full Text Available The ultrastructure of chloroplasts was studied in mesophyll cells of the leaves of silver birch (Betula pendula showing interveinal chlorosis or premature yellowing, in comparison with leaves without symptoms or exhibiting symptoms of natural senescence. The leaves were collected between May 26 to June 7 and additionally in the September 10-12 from the upper part of the crown, from increments of the past four years. No major difference in ultrastructure of chloroplasts was found between spongy and palisade mesophyll cells. The following senescencerelated changes were observed in chloroplasts of prematurely yellowed leaves and showing inteveinal chlorosis: reduced chloroplast size, degeneration of the membrane systems of thylakoids and increased electron density of plastoglobuli. The most electron dark globules (lipid droplets were found together with starch grains in cells of spongy mesophyll of leaves showing interveinal chlorosis. Abnormal, spherical and rounded chloroplasts with electron-dark inside of thylakoids or the electron-dark stroma between thylakoids were found only in yellowed and chlorotic leaves in spring.

  14. Chloroplast movement.

    Science.gov (United States)

    Wada, Masamitsu

    2013-09-01

    Chloroplast movement is important for plant survival under high light and for efficient photosynthesis under low light. This review introduces recent knowledge on chloroplast movement and shows how to analyze the responses and the moving mechanisms, potentially inspiring research in this field. Avoidance from the strong light is mediated by blue light receptor phototropin 2 (phot2) plausibly localized on the chloroplast envelop and accumulation at the week light-irradiated area is mediated by phot1 and phot2 localized on the plasma membrane. Chloroplasts move by chloroplast actin (cp-actin) filaments that must be polymerized by Chloroplast Unusual Positioning1 (CHUP1) at the front side of moving chloroplast. To understand the signal transduction pathways and the mechanism of chloroplast movement, that is, from light capture to motive force-generating mechanism, various methods should be employed based on the various aspects. Observation of chloroplast distribution pattern under different light condition by fixed cell sectioning is somewhat an old-fashioned technique but the most basic and important way. However, most importantly, precise chloroplast behavior during and just after the induction of chloroplast movement by partial cell irradiation using an irradiator with either low light or strong light microbeam should be recorded by time lapse photographs under infrared light and analyzed. Recently various factors involved in chloroplast movement, such as cp-actin filaments and CHUP1, could be traced in Arabidopsis transgenic lines with fluorescent protein tags under a confocal laser scanning microscope (CLSM) and/or a total internal reflection fluorescence microscope (TIRFM). These methods are listed and their advantages and disadvantages are evaluated.

  15. A difference gel electrophoresis study on thylakoids isolated from poplar leaves reveals a negative impact of ozone exposure on membrane proteins.

    Science.gov (United States)

    Bohler, Sacha; Sergeant, Kjell; Hoffmann, Lucien; Dizengremel, Pierre; Hausman, Jean-Francois; Renaut, Jenny; Jolivet, Yves

    2011-07-01

    Populus tremula L. x P. alba L. (Populus x canescens (Aiton) Smith), clone INRA 717-1-B4, saplings were subjected to 120 ppb ozone exposure for 28 days. Chloroplasts were isolated, and the membrane proteins, solubilized using the detergent 1,2-diheptanoyl-sn-glycero-3-phosphocholine (DHPC), were analyzed in a difference gel electrophoresis (DiGE) experiment comparing control versus ozone-exposed plants. Extrinsic photosystem (PS) proteins and adenosine triphosphatase (ATPase) subunits were detected to vary in abundance. The general trend was a decrease in abundance, except for ferredoxin-NADP(+) oxidoreductase (FNR), which increased after the first 7 days of exposure. The up-regulation of FNR would increase NAPDH production for reducing power and detoxification inside and outside of the chloroplast. Later on, FNR and a number of PS and ATPase subunits decrease in abundance. This could be the result of oxidative processes on chloroplast proteins but could also be a way to down-regulate photochemical reactions in response to an inhibition in Calvin cycle activity.

  16. A novel chloroplast-localized protein EMB1303 is required for chloroplast development in Arabidopsis

    Institute of Scientific and Technical Information of China (English)

    Xiaozhen Huang; Xiaoyan Zhang; Shuhua Yang

    2009-01-01

    To understand the molecular mechanisms underlying chloroplast development, we isolated and characterized the albino mutant emb1303-1 in Arabidopsis. The mutant displayed a severe dwarf phenotype with small albino rosette leaves and short roots on a synthetic medium containing sucrose. It is pigment-deficient and seedling lethal when grown in soil. Embryo development was delayed in the mutant, although seed germination was not significantly im-paired. The plastids of emb1303-1 were arrested in early developmental stages without the classical stack of thylakoid membrane. Genetic and molecular analyses uncovered that the EMB1303 gene encodes a novel chloroplast-localized protein. Mieroarray and RT-PCR analyses revealed that a number of nuclear-and plastid-encoded genes involved in photosynthesis and chloroplast biogenesis were substantially downregulated in the mutant. Moreover, the accu-mulation of several major chloroplast proteins was severely compromised in emb1303-1. These results suggest that EMBI303 is essential for chloroplast development.

  17. The toxic dinoflagellate Dinophysis acuminata harbors permanent chloroplasts of cryptomomad prigin, not kleptochloroplasts

    DEFF Research Database (Denmark)

    Garcia, Lydia; Moestrup, Øjvind; Hansen, Per Juel;

    2010-01-01

    Most species belonging to the toxigenic genus Dinophysis have chloroplasts of cryptophyte origin. Whether these chloroplasts are temporarily sequestered from the prey, or permanently established under the control of the dinoflagellate is currently disputed. To investigate this, a culture...... of Dinophysis acuminata was established by feeding it the phototrophic ciliate Mesodinium rubrum (= Myrionecta rubra), which again was fed the cryptophyte Teleaulax amphioxeia. Molecular analysis comprising the nucleomorph LSU and two chloroplast markers (tufA gene and a fragment from the end of 16S r......DNA to the beginning of 23S rDNA) resulted in identical sequences for the three organisms. Yet, transmission electron microscopy of the three organisms revealed that several chloroplast features separated D. acuminata from both T. amphioxeia and M. rubrum. The thylakoid arrangement, the number of membranes around...

  18. Chloroplast ultrastructure in leaves of Cucumis sativus chlorophyll mutant

    Directory of Open Access Journals (Sweden)

    Irena Palczewska

    2014-02-01

    Full Text Available The developing and young leaves of Cucumis sativus chlorophyll mutants are yellow, when mature they become green and do not differ in their colour from those of control plants. The mesophyll of yellow leaves contains a diversiform plastid population with a varying degree of defectiveness, which is mainly manifested in the reduction or disorganization of the typical thylakoid system. DNA areas, ribosome-like particles and aggregates of electron-dense material are preserved in the stroma of mutated plastids. Starch grains are deficient. Apart from mutated plastids, chloroplasts with a normal structure, as in control plants, were also observed.The leaf greening process is accompanied by a reconstruction and rearrangement of the inner chloroplast lamellar system and an ability to accumulate starch. However, in the mutant chloroplasts as compared with control-plant ones, an irregular arrangement of grana and reduced number of inter-grana thylakoids can be seen. An osmiophilic substance stored in the stroma of mutated plastids and the vesicles formed from an internal plastid membrane take part in restoration of the membrane system.

  19. SUPPRESSOR OF VARIEGATION4,a New var2 Suppressor Locus,Encodes a Pioneer Protein that Is Required for Chloroplast Biogenesis

    Institute of Scientific and Technical Information of China (English)

    Fei Yu; Gordon R.Gray; Steven R.Rodermel; Sung-Soon Park; Xiayan Liu; Andrew Foudree; Aigen Fu; Marta Powikrowska; Anastassia Khrouchtchova; Poul Erik Jensen; Jillian N.Kriger

    2011-01-01

    VAR2 is an integral thylakoid membrane protein and a member of the versatile FtsH class of metalloproteases in prokaryotes and eukaryotes. Recessive mutations in the VAR2 locus give rise to variegated plants (var2) that contain white sectors with abnormal plastids and green sectors with normal-appearing chloroplasts. In a continuing effort to isolate second-site suppressors of var2 variegation,we characterize in this report ems2505,a suppressor strain that has a vi-rescent phenotype due to a missense mutation in At4g28590,the gene for a pioneer protein. We designated this gene SVR4 (for SUPPRESSOR OF VARIEGATI0N4) and the mutant allele in ems2505 as svr4-1. We demonstrate that SVR4 is located in chloroplasts and that svr4-1 single mutants are normal with respect to chloroplast anatomy and thylakoid membrane protein accumulation. However,they are modestly impaired in several aspects of photochemistry and have enhanced non-photochemical quenching (NPQ) capacity. A T-DNA insertion allele of SVR4,svr4-2,is seedling-lethal due to an early blockage of chloroplast development. We conclude that SVR4 is essential for chloroplast biogenesis,and hypothesize that SVR4 mediates some aspect of thylakoid structure or function that controls NPQ. We propose that in the suppressor strain,photoinhibitory pressure caused by a lack of VAR2 is ameliorated early in chloroplast development by enhanced NPQ capacity caused by reduced SVR4 activity. This would result in an increase in the number of chloroplasts that are able to surmount a threshold necessary to avoid photo-damage and thereby develop into functional chloroplasts.

  20. The small molecule fenpropimorph rapidly converts chloroplast membrane lipids to triacylglycerols in Chlamydomonas reinhardtii

    Science.gov (United States)

    Kim, Hanul; Jang, Sunghoon; Kim, Sangwoo; Yamaoka, Yasuyo; Hong, Daewoong; Song, Won-Yong; Nishida, Ikuo; Li-Beisson, Yonghua; Lee, Youngsook

    2015-01-01

    Concern about global warming has prompted an intense interest in developing economical methods of producing biofuels. Microalgae provide a promising platform for biofuel production, because they accumulate high levels of lipids, and do not compete with food or feed sources. However, current methods of producing algal oil involve subjecting the microalgae to stress conditions, such as nitrogen deprivation, and are prohibitively expensive. Here, we report that the fungicide fenpropimorph rapidly causes high levels of neutral lipids to accumulate in Chlamydomonas reinhardtii cells. When treated with fenpropimorph (10 μg mL-1) for 1 h, Chlamydomonas cells accumulated at least fourfold the amount of triacylglycerols (TAGs) present in the untreated control cells. Furthermore, the quantity of TAGs present after 1 h of fenpropimorph treatment was over twofold higher than that formed after 9 days of nitrogen starvation in medium with no acetate supplement. Biochemical analysis of lipids revealed that the accumulated TAGs were derived mainly from chloroplast polar membrane lipids. Such a conversion of chloroplast polar lipids to TAGs is desirable for biodiesel production, because polar lipids are usually removed during the biodiesel production process. Thus, our data exemplified that a cost and time effective method of producing TAGs is possible using fenpropimorph or similar drugs. PMID:25759683

  1. Transfer of the cytochrome P450-dependent dhurrin pathway from Sorghum bicolor into Nicotiana tabacum chloroplasts for light-driven synthesis

    DEFF Research Database (Denmark)

    Gnanasekaran, Thiyagarajan; Karcher, Daniel; Nielsen, Agnieszka Janina Zygadlo;

    2016-01-01

    . For this purpose, we stably engineered the dhurrin pathway from Sorghum bicolor into the chloroplasts of Nicotiana tabacum (tobacco). Dhurrin is a cyanogenic glucoside and its synthesis from the amino acid tyrosine is catalysed by two membrane-bound cytochrome P450 enzymes (CYP79A1 and CYP71E1) and a soluble...... compared to 6% in sorghum. The results obtained pave the way for plant P450s involved in the synthesis of economically important compounds to be engineered into the thylakoid membrane of chloroplasts, and demonstrate that their full catalytic cycle can be driven directly by photosynthesis-derived electrons....

  2. MET1 is a thylakoid-associated TPR protein involved in photosystem II supercomplex formation and repair in Arabidopsis.

    Science.gov (United States)

    Bhuiyan, Nazmul H; Friso, Giulia; Poliakov, Anton; Ponnala, Lalit; van Wijk, Klaas J

    2015-01-01

    Photosystem II (PSII) requires constant disassembly and reassembly to accommodate replacement of the D1 protein. Here, we characterize Arabidopsis thaliana MET1, a PSII assembly factor with PDZ and TPR domains. The maize (Zea mays) MET1 homolog is enriched in mesophyll chloroplasts compared with bundle sheath chloroplasts, and MET1 mRNA and protein levels increase during leaf development concomitant with the thylakoid machinery. MET1 is conserved in C3 and C4 plants and green algae but is not found in prokaryotes. Arabidopsis MET1 is a peripheral thylakoid protein enriched in stroma lamellae and is also present in grana. Split-ubiquitin assays and coimmunoprecipitations showed interaction of MET1 with stromal loops of PSII core components CP43 and CP47. From native gels, we inferred that MET1 associates with PSII subcomplexes formed during the PSII repair cycle. When grown under fluctuating light intensities, the Arabidopsis MET1 null mutant (met1) showed conditional reduced growth, near complete blockage in PSII supercomplex formation, and concomitant increase of unassembled CP43. Growth of met1 in high light resulted in loss of PSII supercomplexes and accelerated D1 degradation. We propose that MET1 functions as a CP43/CP47 chaperone on the stromal side of the membrane during PSII assembly and repair. This function is consistent with the observed differential MET1 accumulation across dimorphic maize chloroplasts.

  3. A Nucleus-Encoded Chloroplast Protein YL1 Is Involved in Chloroplast Development and Efficient Biogenesis of Chloroplast ATP Synthase in Rice

    Science.gov (United States)

    Chen, Fei; Dong, Guojun; Wu, Limin; Wang, Fang; Yang, Xingzheng; Ma, Xiaohui; Wang, Haili; Wu, Jiahuan; Zhang, Yanli; Wang, Huizhong; Qian, Qian; Yu, Yanchun

    2016-01-01

    Chloroplast ATP synthase (cpATPase) is an importance thylakoid membrane-associated photosynthetic complex involved in the light-dependent reactions of photosynthesis. In this study, we isolated and characterized a rice (Oryza sativa) mutant yellow leaf 1 (yl1), which exhibits chlorotic leaves throughout developmental stages. The YL1 mutation showed reduced chlorophyll contents, abnormal chloroplast morphology, and decreased photochemical efficiency. Moreover, YL1 deficiency disrupts the expression of genes associated with chloroplast development and photosynthesis. Molecular and genetic analyses revealed that YL1 is a nucleus-encoded protein with a predicted transmembrane domain in its carboxyl-terminus that is conserved in the higher plant kingdom. YL1 localizes to chloroplasts and is preferentially expressed in green tissues containing chloroplasts. Immunoblot analyses showed that inactivation of YL1 leads to drastically reduced accumulation of AtpA (α) and AtpB (β), two core subunits of CF1αβ subcomplex of cpATPase, meanwhile, a severe decrease (ca. 41.7%) in cpATPase activity was observed in the yl1-1 mutant compared with the wild type. Furthermore, yeast two-hybrid and bimolecular fluorescence complementation assays revealed a specific interaction between YL1 and AtpB subunit of cpATPase. Taken together, our results suggest that YL1 is a plant lineage-specific auxiliary factor involved in the biogenesis of the cpATPase complex, possibly via interacting with the β-subunit. PMID:27585744

  4. Light-dependent degradation of the QB-protein in isolated pea thylakoids

    Science.gov (United States)

    Ohad, I.; Kyle, D. J.; Hirschberg, J.

    1985-01-01

    The 32 000-dalton QB-protein of photosystem II (PS II) is rapidly damaged and removed from isolated pea thylakoids during incubation in the light resulting in a loss of photosynthetic electron flow through PS II. This in vitro photoinhibition is similar to that previously reported with intact Chlamydomonas cells. The damage occurs at a faster rate in vitro, however, due to the inability of isolated thylakoids to synthesize replacement QB-protein. The removal of the damaged QB-protein does not require any soluble components of the chloroplast stroma and is unaffected by the protease inhibitors phenyl-methylsulfonylfluoride or antipain. Unlike the effect of trypsin, no low mol. wt. membrane-bound or soluble fragments of the labelled QB-protein could be identified either by autoradiography or immunologically using polyclonal antibodies specific for the QB-protein. The lightinduced damage to the QB-protein (indicated by a loss of QB functional activity), preceded the removal of the protein from the membrane. We conclude that photodamage of the QB-protein generates a conformational change which renders the protein susceptible to attack by a highly efficient, intrinsic membrane protease. ImagesFig. 1.Fig. 3.Fig. 4.Fig. 5. PMID:16453621

  5. Signal integration by chloroplast phosphorylation networks: An update

    Directory of Open Access Journals (Sweden)

    Anna eSchoenberg

    2012-11-01

    Full Text Available Forty years after the initial discovery of light-dependent protein phosphorylation at the thylakoid membrane system, we are now beginning to understand the roles of chloroplast phosphorylation networks in their function to decode and mediate information on the metabolic status of the organelle to long-term adaptations in plastid and nuclear gene expression. With the help of genetics and functional genomics tools, chloroplast kinases and several hundred phosphoproteins were identified that now await detailed functional characterization. The regulation and the target protein spectrum of some kinases are understood, but this information is fragmentary with respect to kinase and target protein crosstalk in a changing environment. In this review we will highlight the most recent advances in the field and discuss approaches that might lead to a comprehensive understanding of plastid signal integration by protein phosphorylation.

  6. Combined effects of simulated acid rain and lanthanum chloride on chloroplast structure and functional elements in rice.

    Science.gov (United States)

    Hu, Huiqing; Wang, Lihong; Zhou, Qing; Huang, Xiaohua

    2016-05-01

    Acid rain and rare earth element (REE) pollution exist simultaneously in many agricultural regions. However, how REE pollution and acid rain affect plant growth in combination remains largely unknown. In this study, the combined effects of simulated acid rain and lanthanum chloride (LaCl3) on chloroplast morphology, chloroplast ultrastructure, functional element contents, chlorophyll content, and the net photosynthetic rate (P n) in rice (Oryza sativa) were investigated by simulating acid rain and rare earth pollution. Under the combined treatment of simulated acid rain at pH 4.5 and 0.08 mM LaCl3, the chloroplast membrane was smooth, proteins on this membrane were uniform, chloroplast structure was integrated, and the thylakoids were orderly arranged, and simulated acid rain and LaCl3 exhibited a mild antagonistic effect; the Mg, Ca, Mn contents, the chlorophyll content, and the P n increased under this combined treatment, with a synergistic effect of simulated acid rain and LaCl3. Under other combined treatments of simulated acid rain and LaCl3, the chloroplast membrane surface was uneven, a clear "hole" was observed on the surface of chloroplasts, and the thylakoids were dissolved and loose; and the P n and contents of functional elements (P, Mg, K, Ca, Mn, Fe, Ni, Cu, Zn and Mo) and chlorophyll decreased. Under these combined treatments, simulated acid rain and LaCl3 exhibited a synergistic effect. Based on the above results, a model of the combined effects of simulated acid rain and LaCl3 on plant photosynthesis was established in order to reveal the combined effects on plant photosynthesis, especially on the photosynthetic organelle-chloroplast. Our results would provide some references for further understanding the mechanism of the combined effects of simulated acid rain and LaCl3 on plant photosynthesis.

  7. The DnaJ-like zinc finger domain protein PSA2 affects light acclimation and chloroplast development in Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Yan-Wen eWang

    2016-03-01

    Full Text Available The biosynthesis of chlorophylls and carotenoids and the assembly of thylakoid membranes are critical for the photoautotrophic growth of plants. Different factors are involved in these two processes. In recent years, members of the DnaJ-like zinc finger domain proteins have been found to take part in the biogenesis and/or the maintenance of plastids. One member of this family of proteins, PSA2, was recently found to localize to the thylakoid lumen and regulate the accumulation of photosystem I. In this study, we report that the silencing of PSA2 in Arabidopsis thaliana resulted in variegated leaves and retarded growth. Although both chlorophylls and total carotenoids decreased in the psa2 mutant, violaxanthin and zeaxanthin accumulated in the mutant seedlings grown under growth condition. Lower levels of non-photochemical quenching and electron transport rate were also found in the psa2 mutant seedlings under growth condition compared with those of the wild-type plants, indicating an impaired capability to acclimate to normal light irradiance when PSA2 was silenced. Moreover, we also observed an abnormal assembly of grana thylakoids and poorly developed stroma thylakoids in psa2 chloroplasts. Taken together, our results demonstrate that PSA2 is a member of the DnaJ-like zinc finger domain protein family that affects light acclimation and chloroplast development.

  8. Fatty acid phytyl ester synthesis in chloroplasts of Arabidopsis.

    Science.gov (United States)

    Lippold, Felix; vom Dorp, Katharina; Abraham, Marion; Hölzl, Georg; Wewer, Vera; Yilmaz, Jenny Lindberg; Lager, Ida; Montandon, Cyrille; Besagni, Céline; Kessler, Felix; Stymne, Sten; Dörmann, Peter

    2012-05-01

    During stress or senescence, thylakoid membranes in chloroplasts are disintegrated, and chlorophyll and galactolipid are broken down, resulting in the accumulation of toxic intermediates, i.e., tetrapyrroles, free phytol, and free fatty acids. Chlorophyll degradation has been studied in detail, but the catabolic pathways for phytol and fatty acids remain unclear. A large proportion of phytol and fatty acids is converted into fatty acid phytyl esters and triacylglycerol during stress or senescence in chloroplasts. We isolated two genes (PHYTYL ESTER SYNTHASE1 [PES1] and PES2) of the esterase/lipase/thioesterase family of acyltransferases from Arabidopsis thaliana that are involved in fatty acid phytyl ester synthesis in chloroplasts. The two proteins are highly expressed during senescence and nitrogen deprivation. Heterologous expression in yeast revealed that PES1 and PES2 have phytyl ester synthesis and diacylglycerol acyltransferase activities. The enzymes show broad substrate specificities and can employ acyl-CoAs, acyl carrier proteins, and galactolipids as acyl donors. Double mutant plants (pes1 pes2) grow normally but show reduced phytyl ester and triacylglycerol accumulation. These results demonstrate that PES1 and PES2 are involved in the deposition of free phytol and free fatty acids in the form of phytyl esters in chloroplasts, a process involved in maintaining the integrity of the photosynthetic membrane during abiotic stress and senescence.

  9. Sycamore amyloplasts can import and process precursors of nuclear encoded chloroplast proteins.

    Science.gov (United States)

    Strzalka, K; Ngernprasirtsiri, J; Watanabe, A; Akazawa, T

    1987-12-16

    Amyloplasts isolated from white-wild suspension-cultured cells of sycamore (Acer pseudoplatanus L.) are found to import and process the precursor of the small subunit (pS) of ribulose-1,5-bisphosphate carboxylase/oxygenase of spinach, but they lack the ability to form its holoenzyme due to the absence of both the large subunit and its binding-protein. They also import the precursor of the 33-kDa extrinsic protein (p33-kDa) of the O2-evolving complex of Photosystem II from spinach, but process is only to an intermediate form (i33-kDa). Chloroplasts from green-mutant cells of sycamore process p33-kDa to its mature form in this heterologous system. These results suggest that the thylakoid-associated protease responsible for the second processing step of p33-kDa is missing in amyloplasts, possibly due to the absence of thylakoid-membranes. In contrast, the apparent import of the precursor of the light-harvesting chlorophyll a/b-binding apoprotein (pLHCP) from spinach was not detected. Sycamore amyloplasts may lack the ability to import this particular thylakoid-protein, or rapidly degrade the imported molecules in the absence of thylakoid-membranes for their proper insertion.

  10. Organization into Higher Ordered Ring Structures Counteracts Membrane Binding of IM30, a Protein Associated with Inner Membranes in Chloroplasts and Cyanobacteria.

    Science.gov (United States)

    Heidrich, Jennifer; Wulf, Verena; Hennig, Raoul; Saur, Michael; Markl, Jürgen; Sönnichsen, Carsten; Schneider, Dirk

    2016-07-15

    The IM30 (inner membrane-associated protein of 30 kDa), also known as the Vipp1 (vesicle-inducing protein in plastids 1), has a crucial role in thylakoid membrane biogenesis and maintenance. Recent results suggest that the protein binds peripherally to membranes containing negatively charged lipids. However, although IM30 monomers interact and assemble into large oligomeric ring complexes with different numbers of monomers, it is still an open question whether ring formation is crucial for membrane interaction. Here we show that binding of IM30 rings to negatively charged phosphatidylglycerol membrane surfaces results in a higher ordered membrane state, both in the head group and in the inner core region of the lipid bilayer. Furthermore, by using gold nanorods covered with phosphatidylglycerol layers and single particle spectroscopy, we show that not only IM30 rings but also lower oligomeric IM30 structures interact with membranes, although with higher affinity. Thus, ring formation is not crucial for, and even counteracts, membrane interaction of IM30.

  11. Photonic multilayer structure of Begonia chloroplasts enhances photosynthetic efficiency.

    Science.gov (United States)

    Jacobs, Matthew; Lopez-Garcia, Martin; Phrathep, O-Phart; Lawson, Tracy; Oulton, Ruth; Whitney, Heather M

    2016-10-24

    Enhanced light harvesting is an area of interest for optimizing both natural photosynthesis and artificial solar energy capture(1,2). Iridescence has been shown to exist widely and in diverse forms in plants and other photosynthetic organisms and symbioses(3,4), but there has yet to be any direct link demonstrated between iridescence and photosynthesis. Here we show that epidermal chloroplasts, also known as iridoplasts, in shade-dwelling species of Begonia(5), notable for their brilliant blue iridescence, have a photonic crystal structure formed from a periodic arrangement of the light-absorbing thylakoid tissue itself. This structure enhances photosynthesis in two ways: by increasing light capture at the predominantly green wavelengths available in shade conditions, and by directly enhancing quantum yield by 5-10% under low-light conditions. These findings together imply that the iridoplast is a highly modified chloroplast structure adapted to make best use of the extremely low-light conditions in the tropical forest understorey in which it is found(5,6). A phylogenetically diverse range of shade-dwelling plant species has been found to produce similarly structured chloroplasts(7-9), suggesting that the ability to produce chloroplasts whose membranes are organized as a multilayer with photonic properties may be widespread. In fact, given the well-established diversity and plasticity of chloroplasts(10,11), our results imply that photonic effects may be important even in plants that do not show any obvious signs of iridescence to the naked eye but where a highly ordered chloroplast structure may present a clear blue reflectance at the microscale. Chloroplasts are generally thought of as purely photochemical; we suggest that one should also think of them as a photonic structure with a complex interplay between control of light propagation, light capture and photochemistry.

  12. Interdependence between oscillations and transients of delayed fluorescence induction processes in the thylakoid membrane of the intact maize leaf: Responses to effects of increased temperatures and drought

    Directory of Open Access Journals (Sweden)

    Radenović Čedomir N.

    2010-01-01

    Full Text Available Standard induction processes of delayed fluorescence (DF of chlorophyll (induction signals occur when an intact leaf segment of maize inbreds and hybrids is kept in the phosphoroscope darkroom for more than 15 minutes (t > 15 min, and at the same time the leaf is illuminated with the intermittent white light. Resolved induction processes of DF chlorophyll into transients: A, B, C, D and E occur when the intact leaf segment of maize inbreds and hybrids is kept in the phosphoroscope darkroom for a significantly shorter period (500 s > t > 30 s, with the time rate t of 30 s, prior to its illumination with the intermittent white light. Induction transients: A, B, C, D and E are characterized with the time of their generation: tA = 31±6 ms (A, tB = 5 ± 0,5 s (B, tC = 15±5 s (C, tD = 360±20 s (D and tE = 670±35 s (E, dynamics of changes in transients intensities (IA, IB, IC, ID and IE and mechanisms of their generation. The induction processes of chlorophyll DF of the intact leaf of maize inbreds and hybrids resolved into transients: A, B, C, D and E are accompanied by the occurrence and different levels of activation energy (Ea, kJ mol-1 that correspond to critical temperatures ranging from 28 to 33°C. The generation mechanisms of induction transients A, B, C, D and E classify them into two groups. Transients A and B are of a physical character, while the transients: C, D, and E are of a chemical character. It is shown that the generation of the induction transients: B, C, D and E simultaneously follows establishing of the oscillations of induction processes of the DF chlorophyll. Oscillating of induction processes of DF chlorophyll is explained by the ion (K+, Na+, H+, Cl- transport mechanism across the thylakoid membrane of the intact leaf of maize inbreds and hybrids grown under conditions of air drought, increased temperatures and water deficiency in the medium.

  13. The chloroplast outer membrane protein CHUP1 interacts with actin and profilin.

    Science.gov (United States)

    Schmidt von Braun, Serena; Schleiff, Enrico

    2008-04-01

    Chloroplasts accumulate in response to low light, whereas high light induces an actin-dependent avoidance movement. This is a long known process, but its molecular base is barely understood. Only recently first components of the blue light perceiving signal cascade initiating this process were described. Among these, a protein was identified by the analysis of a deletion mutant in the corresponding gene resulting in a chloroplast unusual positioning phenotype. The protein was termed CHUP1 and initial results suggested chloroplast localization. We demonstrate that the protein is indeed exclusively and directly targeted to the chloroplast surface. The analysis of the deletion mutant of CHUP1 using microarray analysis shows an influence on the expression of genes found to be up-regulated, but not on genes found to be down-regulated upon high light exposure in wild-type. Analyzing a putative role of CHUP1 as a linker between chloroplasts and the cytoskeleton, we demonstrate an interaction with actin, which is independent on the filamentation status of actin. Moreover, binding of CHUP1 to profilin -- an actin modifying protein -- could be shown and an enhancing effect of CHUP1 on the interaction of profilin to actin is demonstrated. Therefore, a role of CHUP1 in bridging chloroplasts to actin filaments and a regulatory function in actin polymerization can be discussed.

  14. Targeting and assembly of components of the TOC protein import complex at the chloroplast outer envelope membrane.

    Science.gov (United States)

    Richardson, Lynn G L; Paila, Yamuna D; Siman, Steven R; Chen, Yi; Smith, Matthew D; Schnell, Danny J

    2014-01-01

    The translocon at the outer envelope membrane of chloroplasts (TOC) initiates the import of thousands of nuclear encoded preproteins required for chloroplast biogenesis and function. The multimeric TOC complex contains two GTP-regulated receptors, Toc34 and Toc159, which recognize the transit peptides of preproteins and initiate protein import through a β-barrel membrane channel, Toc75. Different isoforms of Toc34 and Toc159 assemble with Toc75 to form structurally and functionally diverse translocons, and the composition and levels of TOC translocons is required for the import of specific subsets of coordinately expressed proteins during plant growth and development. Consequently, the proper assembly of the TOC complexes is key to ensuring organelle homeostasis. This review will focus on our current knowledge of the targeting and assembly of TOC components to form functional translocons at the outer membrane. Our analyses reveal that the targeting of TOC components involves elements common to the targeting of other outer membrane proteins, but also include unique features that appear to have evolved to specifically facilitate assembly of the import apparatus.

  15. Targeting and Assembly of Components of the TOC Protein Import Complex at the Chloroplast Outer Envelope Membrane

    Directory of Open Access Journals (Sweden)

    Lynn G.L. Richardson

    2014-06-01

    Full Text Available The translocon at the outer envelope membrane of chloroplasts (TOC initiates the import of thousands of nuclear encoded preproteins required for chloroplast biogenesis and function. The multimeric TOC complex contains two GTP-regulated receptors, Toc34 and Toc159, which recognize the transit peptides of preproteins and initiate protein import through a β–barrel membrane channel, Toc75. Different isoforms of Toc34 and Toc159 assemble with Toc75 to form structurally and functionally diverse translocons, and the composition and levels of TOC translocons is required for the import of specific subsets of coordinately expressed proteins during plant growth and development. Consequently, the proper assembly of the TOC complexes is key to ensuring organelle homeostasis. This review will focus on our current knowledge of the targeting and assembly of TOC components to form functional translocons at the outer membrane. Our analyses reveal that the targeting of TOC components involves elements common to the targeting of other outer membrane proteins, but also include unique features that appear to have evolved to specifically facilitate assembly of the import apparatus.

  16. Essential role of VIPP1 in chloroplast envelope maintenance in Arabidopsis.

    Science.gov (United States)

    Zhang, Lingang; Kato, Yusuke; Otters, Stephanie; Vothknecht, Ute C; Sakamoto, Wataru

    2012-09-01

    VESICLE-INDUCING PROTEIN IN PLASTIDS1 (VIPP1), proposed to play a role in thylakoid biogenesis, is conserved in photosynthetic organisms and is closely related to Phage Shock Protein A (PspA), which is involved in plasma membrane integrity in Escherichia coli. This study showed that chloroplasts/plastids in Arabidopsis thaliana vipp1 knockdown and knockout mutants exhibit a unique morphology, forming balloon-like structures. This altered morphology, as well as lethality of vipp1, was complemented by expression of VIPP1 fused to green fluorescent protein (VIPP1-GFP). Several lines of evidence show that the balloon chloroplasts result from chloroplast swelling related to osmotic stress, implicating VIPP1 in the maintenance of plastid envelopes. In support of this, Arabidopsis VIPP1 rescued defective proton leakage in an E. coli pspA mutant. Microscopy observation of VIPP1-GFP in transgenic Arabidopsis revealed that VIPP1 forms large macrostructures that are integrated into various morphologies along the envelopes. Furthermore, live imaging revealed that VIPP1-GFP is highly mobile when chloroplasts are subjected to osmotic stress. VIPP1-GFP showed dynamic movement in the transparent area of spherical chloroplasts, as the fluorescent molecules formed filament-like structures likely derived from disassembly of the large VIPP1 complex. Collectively, our data demonstrate that VIPP1 is a multifunctional protein in chloroplasts that is critically important for envelope maintenance.

  17. Role of plastid transglutaminase in LHCII polyamination and thylakoid electron and proton flow.

    Directory of Open Access Journals (Sweden)

    Nikolaos E Ioannidis

    Full Text Available Transglutaminases function as biological glues in animal cells, plant cells and microbes. In energy producing organelles such as chloroplasts the presence of transglutaminases was recently confirmed. Furthermore, a plastidial transglutaminase has been cloned from maize and the first plants overexpressing tgz are available (Nicotiana tabacum TGZ OE. Our hypothesis is that the overexpression of plastidal transglutaminase will alter photosynthesis via increased polyamination of the antenna of photosystem II. We have used standard analytical tools to separate the antenna from photosystem II in wild type and modified plants, 6 specific antibodies against LHCbs to confirm their presence and sensitive HPLC method to quantify the polyamination level of these proteins. We report that bound spermidine and spermine were significantly increased (∼80% in overexpressors. Moreover, we used recent advances in in vivo probing to study simultaneously the proton and electron circuit of thylakoids. Under physiological conditions overexpressors show a 3-fold higher sensitivity of the antenna down regulation loop (qE to the elicitor (luminal protons which is estimated as the ΔpH component of thylakoidal proton motive force. In addition, photosystem (hyper-PSIIα with an exceptionally high antenna (large absorption cross section, accumulate in transglutaminase over expressers doubling the rate constant of light energy utilization (Kα and promoting thylakoid membrane stacking. Polyamination of antenna proteins is a previously unrecognized mechanism for the modulation of the size (antenna absorption cross section and sensitivity of photosystem II to down regulation. Future research will reveal which peptides and which residues of the antenna are responsible for such effects.

  18. High Yield Non-detergent Isolation of Photosystem I-Light-harvesting Chlorophyll II Membranes from Spinach Thylakoids: IMPLICATIONS FOR THE ORGANIZATION OF THE PS I ANTENNAE IN HIGHER PLANTS.

    Science.gov (United States)

    Bell, Adam J; Frankel, Laurie K; Bricker, Terry M

    2015-07-24

    Styrene-maleic acid copolymer was used to effect a non-detergent partial solubilization of thylakoids from spinach. A high density membrane fraction, which was not solubilized by the copolymer, was isolated and was highly enriched in the Photosystem (PS) I-light-harvesting chlorophyll (LHC) II supercomplex and depleted of PS II, the cytochrome b6/f complex, and ATP synthase. The LHC II associated with the supercomplex appeared to be energetically coupled to PS I based on 77 K fluorescence, P700 photooxidation, and PS I electron transport light saturation experiments. The chlorophyll (Chl) a/b ratio of the PS I-LHC II membranes was 3.2 ± 0.9, indicating that on average, three LHC II trimers may associate with each PS I. The implication of these findings within the context of higher plant PS I antenna organization is discussed.

  19. Translocation of a phycoerythrin alpha subunit across five biological membranes.

    Science.gov (United States)

    Gould, Sven B; Fan, Enguo; Hempel, Franziska; Maier, Uwe-G; Klösgen, Ralf Bernd

    2007-10-12

    Cryptophytes, unicellular algae, evolved by secondary endosymbiosis and contain plastids surrounded by four membranes. In contrast to cyanobacteria and red algae, their phycobiliproteins do not assemble into phycobilisomes and are located within the thylakoid lumen instead of the stroma. We identified two gene families encoding phycoerythrin alpha and light-harvesting complex proteins from an expressed sequence tag library of the cryptophyte Guillardia theta. The proteins bear a bipartite topogenic signal responsible for the transport of nuclear encoded proteins via the ER into the plastid. Analysis of the phycoerythrin alpha sequences revealed that more than half of them carry an additional, third topogenic signal comprising a twin arginine motif, which is indicative of Tat (twin arginine transport)-specific targeting signals. We performed import studies with several derivatives of one member using a diatom transformation system, as well as intact chloroplasts and thylakoid vesicles isolated from pea. We demonstrated the different targeting properties of each individual part of the tripartite leader and show that phycoerythrin alpha is transported across the thylakoid membrane into the thylakoid lumen and protease-protected. Furthermore, we showed that thylakoid transport of phycoerythrin alpha takes place by the Tat pathway even if the 36 amino acid long bipartite topogenic signal precedes the actual twin arginine signal. This is the first experimental evidence of a protein being targeted across five biological membranes.

  20. The photoelectrical properties of thylakoid membrane from two plants fabricated on Nano-ZnO%两种植物类囊体膜在纳米ZnO上的组装和光电性质

    Institute of Scientific and Technical Information of China (English)

    刘双; 张攀辉; 邱宇; 张建; 于道永

    2012-01-01

    ZnO nanowires were synthesized on the FTO glass by hydrothermal method and characterized by atomic force microscopy(AFM) and scanning electron microscopy(SEM).Thylakoid membranes isolated from higher plant spinach and marine green alga Ulva lactuca were fabricated on the ZnO nanowires by using Langmuir Blodgett(LB) technique.The photoelectrical properties of these two resulting systems were assessed and compared by solar cell test system.Photocurrents were both observed the solar cells containing the LB films of the thylakoid membranes of these two plants.The photoelectric conversion efficiencies of solar cells were increased significantly with the numbers of LB film layer.In addition,the photoelectric conversion efficiency of cell containing Ulva thylakoid membrane was much higher than that of spinach.%用水热合成法在掺氟氧化锡(SnO2:F,FTO)导电玻璃上合成了ZnO纳米线并用原子力显微镜(AFM)和扫描电镜(SEM)对其进行了表征。利用LB膜技术将高等植物菠菜(Spinacia oleracea)和海洋绿藻石莼(Ulva lactuca)的类囊体膜分别固定在纳米ZnO上组装成光电池,用太阳能测试系统检测比较了其光电性质。研究表明,由两种植物的类囊体膜LB膜组成的光电池都能产生光生电流;蛋白LB膜的层数显著影响了光电池的光电转化效率,随着层数的增加,光电转化效率大大增加。此外,石莼类囊体膜组装的光电池光电转化效率明显高于菠菜类囊体膜。

  1. Maintenance of Chloroplast Components during Chromoplast Differentiation in the Tomato Mutant Green Flesh.

    Science.gov (United States)

    Cheung, A. Y.; McNellis, T.; Piekos, B.

    1993-04-01

    During ripening of tomato (Lycopersicon esculentum) fruit, chloroplasts develop into chromoplasts. The chloroplast-chromoplast transition is marked by the accumulation of carotenoids and the disappearance of chlorophyll, the degradation of the highly structured thylakoid membrane system, and a reduction in the levels of proteins and mRNAs associated with photosynthesis. In the tomato mutant green flesh (gf), detectable amounts of chlorophyll remain in the ripe, mutant fruit, giving rise to a rusty red fruit color and suggesting that at least chlorophyll degradation is defective in the mutant. We show here that the ultrastructure of the plastids in the ripe gf fruit maintained significant amouonts of the chloroplast thylakoid grana along with structures characteristic of tomato chromoplasts. The maintenance of chloroplast structure in the gf ripe fruit was paralleled on the molecular level by the retention of plastid photosynthetic components that normally decline significantly in ripening tomato fruits. These included the light-harvesting chlorophyll a/b-binding proteins of photosystem II, the second electron accepting plastoquinone of photosystem II binding protein, the large and small subunits of ribulose bisphosphate carboxylase/oxygenase, the 33-kD oxygen evolution protein, and cytochrome b559. Similarly, photosynthetic transcripts, cab, psbA, rbcL, rbcS, and psbE mRNAs, also accumulated to higher levels in ripening gf fruit than wild type. It is interesting that the levels of some of these transcripts, especially cab mRNA, were noticeably higher in the mature gf green fruit than in the corresponding wild-type fruit. This suggests that the onset of the effect from the gf mutation might be earlier than fruit ripening. We also observed that when chloroplast formation was blocked during the development and ripening of gf fruit, these mutant fruits were bright red and their chromoplasts were indistinguishable from those found in wild-type ripe fruits grown and

  2. Discrete redox signaling pathways regulate photosynthetic light-harvesting and chloroplast gene transcription.

    Directory of Open Access Journals (Sweden)

    John F Allen

    Full Text Available In photosynthesis in chloroplasts, two related regulatory processes balance the actions of photosystems I and II. These processes are short-term, post-translational redistribution of light-harvesting capacity, and long-term adjustment of photosystem stoichiometry initiated by control of chloroplast DNA transcription. Both responses are initiated by changes in the redox state of the electron carrier, plastoquinone, which connects the two photosystems. Chloroplast Sensor Kinase (CSK is a regulator of transcription of chloroplast genes for reaction centres of the two photosystems, and a sensor of plastoquinone redox state. We asked whether CSK is also involved in regulation of absorbed light energy distribution by phosphorylation of light-harvesting complex II (LHC II. Chloroplast thylakoid membranes isolated from a CSK T-DNA insertion mutant and from wild-type Arabidopsis thaliana exhibit similar light- and redox-induced (32P-labelling of LHC II and changes in 77 K chlorophyll fluorescence emission spectra, while room-temperature chlorophyll fluorescence emission transients from Arabidopsis leaves are perturbed by inactivation of CSK. The results indicate indirect, pleiotropic effects of reaction centre gene transcription on regulation of photosynthetic light-harvesting in vivo. A single, direct redox signal is transmitted separately to discrete transcriptional and post-translational branches of an integrated cytoplasmic regulatory system.

  3. 盐胁迫对水稻幼苗类囊体膜脂肪酸组分的影响%Effects of Salt Stress on Fatty Acid Composition of Thylakoid Membrane of Two Rice Cultivars Differing in Salt Tolerance

    Institute of Scientific and Technical Information of China (English)

    王仁雷; 华春; 周峰; 周泉澄; 周斌伟

    2008-01-01

    [Objective] The aim of the study is to understand the changes of fatty acid composition of rice thylakoid membrane under salt stress. [Method] Under salt stress of different concentrations of NaCl, rice seedlings of Pokkali and Peta with six leaves and one central leaf were used as experimental materials to extract the fatty acid from their thylakoid membranes, and gas chromatograph(1890) was used to analyze fatty acid composition. [Result] Fatty acid component 14∶ 0, 18∶ 0, 16∶ 1(3t), 18∶ 1 in both the two experimental materials showed little variations in the first four days of salt stress, whereafter they increased slightly; while the fatty acid component 16∶ 0 and level of saturation of fatty acid (LSFA) showed the similar variation trend in the first four days of treatment compared to those of the fatty acid components mentioned above, whereafter they rose in Pokkali and presented an opposite variation trend in Peta; fatty acid component 18∶ 3 and level of unsaturation of fatty acid (LUFA) reduced all the time under stress condition, and the reducing amplitude in 100 mmol/L NaCl treatment group was smaller than that of 100 mmol/L NaCl treatment group, and in Pokkali was smaller than that in Peta under specific conditions. Meanwhile, level of saturation of fatty acid in both experimental materials increased, and the rising amplitude in Peta was smaller than that of Pokkali. [Conclusion] With regard to LUFA, Pokkali is endowed with more salt tolerance than Peta.

  4. AtDeg2 – a chloroplast protein with dual protease/chaperone activity

    Directory of Open Access Journals (Sweden)

    Przemysław Jagodzik

    2014-07-01

    Full Text Available Chloroplast protease AtDeg2 (an ATP-independent serine endopeptidase is cytosolically synthesized as a precursor, which is imported into the chloroplast stroma and deprived of its transit peptide. Then the mature protein undergoes routing to its functional location at the stromal side of thylakoid membrane. In its linear structure AtDeg2 molecule contains the protease domain with catalytic triad (HDS and two PDZ domains (PDZ1 and PDZ2. In vivo AtDeg2 most probably exists as a supposedly inactive haxamer, which may change its oligomeric stage to form active 12-mer, or 24-mer. AtDeg2 has recently been demonstrated to exhibit dual protease/chaperone function. This review is focused on the current awareness with regard to AtDeg2 structure and functional significance.

  5. Lipid composition determines the effects of arbutin on the stability of membranes.

    OpenAIRE

    Hincha, D K; Oliver, A E; Crowe, J H

    1999-01-01

    Arbutin (hydroquinone-beta-D-glucopyranoside) is an abundant solute in the leaves of many freezing- or desiccation-tolerant plants. Its physiological role in plants, however, is not known. Here we show that arbutin protects isolated spinach (Spinacia oleracea L.) thylakoid membranes from freeze-thaw damage. During freezing of liposomes, the presence of only 20 mM arbutin led to complete leakage of a soluble marker from egg PC (EPC) liposomes. When the nonbilayer-forming chloroplast lipid mono...

  6. Biogenesis of cytochrome b6 in photosynthetic membranes.

    Science.gov (United States)

    Saint-Marcoux, Denis; Wollman, Francis-André; de Vitry, Catherine

    2009-06-29

    In chloroplasts, binding of a c'-heme to cytochrome b(6) on the stromal side of the thylakoid membranes requires a specific mechanism distinct from the one at work for c-heme binding to cytochromes f and c(6) on the lumenal side of membranes. Here, we show that the major protein components of this pathway, the CCBs, are bona fide transmembrane proteins. We demonstrate their association in a series of hetero-oligomeric complexes, some of which interact transiently with cytochrome b(6) in the process of heme delivery to the apoprotein. In addition, we provide preliminary evidence for functional assembly of cytochrome b(6)f complexes even in the absence of c'-heme binding to cytochrome b(6). Finally, we present a sequential model for apo- to holo-cytochrome b(6) maturation integrated within the assembly pathway of b(6)f complexes in the thylakoid membranes.

  7. The TIC complex uncovered: The alternative view on the molecular mechanism of protein translocation across the inner envelope membrane of chloroplasts.

    Science.gov (United States)

    Nakai, Masato

    2015-09-01

    Chloroplasts must import thousands of nuclear-encoded preproteins synthesized in the cytosol through two successive protein translocons at the outer and inner envelope membranes, termed TOC and TIC, respectively, to fulfill their complex physiological roles. The molecular identity of the TIC translocon had long remained controversial; two proteins, namely Tic20 and Tic110, had been proposed to be central to protein translocation across the inner envelope membrane. Tic40 also had long been considered to be another central player in this process. However, recently, a novel 1-megadalton complex consisting of Tic20, Tic56, Tic100, and Tic214 was identified at the chloroplast inner membrane of Arabidopsis and was demonstrated to constitute a general TIC translocon which functions in concert with the well-characterized TOC translocon. On the other hand, direct interaction between this novel TIC transport system and Tic110 or Tic40 was hardly observed. Consequently, the molecular model for protein translocation across the inner envelope membrane of chloroplasts might need to be extensively revised. In this review article, I intend to propose such alternative view regarding the TIC transport system in contradistinction to the classical view. I also would emphasize importance of reevaluation of previous works in terms of with what methods these classical Tic proteins such as Tic110 or Tic40 were picked up as TIC constituents at the very beginning as well as what actual evidence there were to support their direct and specific involvement in chloroplast protein import. This article is part of a Special Issue entitled: Chloroplast Biogenesis.

  8. Structure and dynamics of thylakoids in land plants

    DEFF Research Database (Denmark)

    Pribil, Mathias; Labs, Mathias; Leister, Dario

    2014-01-01

    of the granum. Stacking of grana is thought to be due to adhesion between Lhcb proteins (LHCII or CP26) located in opposed thylakoid membranes. The grana margins contain oligomers of CURT1 proteins, which appear to control the size and number of grana discs in a dosage- and phosphorylation-dependent manner...

  9. Chloroplastic responses of ponderosa pine (Pinus ponderosa) seedlings to ozone exposure.

    Science.gov (United States)

    Anderson, Paul D; Palmer, Brent; Houpis, James L J; Smith, Mary K; Pushnik, James C

    2003-06-01

    Integrity of chloroplast membranes is essential to photosynthesis. Loss of thylakoid membrane integrity has been proposed as a consequence of ozone (O(3)) exposure and therefore may be a mechanistic basis for decreased photosynthetic rates commonly associated with ozone exposure. To investigate this hypothesis, Pinus ponderosa seedlings were exposed to ambient air or ozone concentrations maintained at 0.15 or 0.30 microliter l(-1) for 10 h day(-1) for 51 days during their second growing season. Over the course of the study, foliage samples were periodically collected for thylakoid membrane, chlorophyll and protein analyses. Additionally, gas-exchange measurements were made in conjunction with foliage sampling to verify that observed chloroplastic responses were associated with ozone-induced changes in photosynthesis. Needles exposed to elevated ozone exhibited decreases in chlorophyll a and b content. The decreases were dependent on the duration and intensity of ozone exposure. When based on equal amounts of chlorophyll, ozone-exposed sample tissue exhibited an increase in total protein. When based on equal amounts of protein, ozone-exposed samples exhibited an increase in 37 kDa proteins, possibly consisting of breakdown products, and a possible decrease in 68 kDa proteins, Rubisco small subunit. There was also a change in the ratio of Photosystem I protein complexes CPI and CPII that may have contributed to decreased photosynthesis. Net photosynthetic rates were decreased in the high ozone treatment suggesting that observed structural and biochemical changes in the chloroplast were associated with alterations of the photosynthetic process.

  10. Mediatorless solar energy conversion by covalently bonded thylakoid monolayer on the glassy carbon electrode.

    Science.gov (United States)

    Lee, Jinhwan; Im, Jaekyun; Kim, Sunghyun

    2016-04-01

    Light reactions of photosynthesis that take place in thylakoid membranes found in plants or cyanobacteria are among the most effective ways of utilizing light. Unlike most researches that use photosystem I or photosystem II as conversion units for converting light to electricity, we have developed a simple method in which the thylakoid monolayer was covalently immobilized on the glassy carbon electrode surface. The activity of isolated thylakoid membrane was confirmed by measuring evolving oxygen under illumination. Glassy carbon surfaces were first modified with partial or full monolayers of carboxyphenyl groups by reductive C-C coupling using 4-aminobenzoic acid and aniline and then thylakoid membrane was bioconjugated through the peptide bond between amine residues of thylakoid and carboxyl groups on the surface. Surface properties of modified surfaces were characterized by cyclic voltammetry, contact angle measurements, and electrochemical impedance spectroscopy. Photocurrent of 230 nA cm(-2) was observed when the thylakoid monolayer was formed on the mixed monolayer of 4-carboxylpheny and benzene at applied potential of 0.4V vs. Ag/AgCl. A small photocurrent resulted when the 4-carboxyphenyl full monolayer was used. This work shows the possibility of solar energy conversion by directly employing the whole thylakoid membrane through simple surface modification.

  11. Expression and membrane-targeting of an active plant cytochrome P450 in the chloroplast of the green alga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Gangl, Doris; Zedler, Julie A Z; Włodarczyk, Artur; Jensen, Poul Erik; Purton, Saul; Robinson, Colin

    2015-02-01

    The unicellular green alga Chlamydomonas reinhardtii has potential as a cell factory for the production of recombinant proteins and other compounds, but mainstream adoption has been hindered by a scarcity of genetic tools and a need to identify products that can be generated in a cost-effective manner. A promising strategy is to use algal chloroplasts as a site for synthesis of high value bioactive compounds such as diterpenoids since these are derived from metabolic building blocks that occur naturally within the organelle. However, synthesis of these complex plant metabolites requires the introduction of membrane-associated enzymes including cytochrome P450 enzymes (P450s). Here, we show that a gene (CYP79A1) encoding a model P450 can be introduced into the C. reinhardtii chloroplast genome using a simple transformation system. The gene is stably expressed and the P450 is efficiently targeted into chloroplast membranes by means of its endogenous N-terminal anchor domain, where it is active and accounts for 0.4% of total cell protein. These results provide proof of concept for the introduction of diterpenoid synthesis pathways into the chloroplast of C. reinhardtii.

  12. 菊花黄绿叶突变体的光合与类囊体膜光谱%Characteristics of Photosynthesis and Spectra Properties of Thylakoid Membranes in the Yellow-Green Leaf Mutant of Chrysanthemum

    Institute of Scientific and Technical Information of China (English)

    常青山; 张利霞; 陈煜; 陈素梅; 刘兆磊; 房伟民; 陈发棣

    2013-01-01

    In order to study photosynthetic mechanism of the yellow-green leaf mutant- 'Jinglingguozi ' of chrysanthemum, the characteristics of photosynthesis and the spectra properties of thylakoid membrane in the green and yellow leaf tissue of the mutant were studied. We measured the photosynthesis, stomatal characteristics, room temperature absorption spectra, chlorophyll emission fluorescence spectra of thylakoid membrane. The results that compared to the green leaf tissue of the mutant, the yellow leaf tissue had lower net photosynthetic rate ( Pn ) , light saturation point (LSP) , dark respiration rate (Rd) , apparent quantum yield (AQY) , but higher light compensation point (LCP). The yellow leaf tissue had lower stomata limit value ( Ls ) , but higher non-stomata limit value. There was no significant difference in the characteristics of stomatal microstructure between the green leaf and yellow leaf tissue. The absorption spectra and fluorescence spectra significantly decreased in the yellow leaf tissue. The yellow leaf tissue had significantly lower capacity of the capture and excitation of light energy and lower the photosynthetic capacity than the green leaf tissue did, which was caused by non-stomata factors, such as a decrease of the function of thylakoid membrane.%以菊花‘金陵国紫’黄绿叶突变体为试验材料,研究该突变体黄绿叶的绿叶与黄叶组织光合与类囊体膜光谱特性.对突变体黄绿叶中绿叶与黄叶组织的光合速率、光响应曲线、气孔特征与类囊体膜光谱特性进行测定与分析.结果表明:与绿叶组织相比,突变体黄叶组织的净光合速率、光饱和点、暗呼吸速率、表观量子效率均显著低于绿叶组织,而光补偿点则显著高于绿叶组织;黄叶组织的气孔限制值低于绿叶组织,非气孔限制值则显著高于绿叶组织,而黄叶与绿叶组织的气孔特征并无显著性差异.突变体黄叶组织类囊体膜叶绿素捕光能力与受激

  13. Artemisinin inhibits chloroplast electron transport activity: mode of action.

    Directory of Open Access Journals (Sweden)

    Adyasha Bharati

    Full Text Available Artemisinin, a secondary metabolite produced in Artemisia plant species, besides having antimalarial properties is also phytotoxic. Although, the phytotoxic activity of the compound has been long recognized, no information is available on the mechanism of action of the compound on photosynthetic activity of the plant. In this report, we have evaluated the effect of artemisinin on photoelectron transport activity of chloroplast thylakoid membrane. The inhibitory effect of the compound, under in vitro condition, was pronounced in loosely and fully coupled thylakoids; being strong in the former. The extent of inhibition was drastically reduced in the presence of uncouplers like ammonium chloride or gramicidin; a characteristic feature described for energy transfer inhibitors. The compound, on the other hand, when applied to plants (in vivo, behaved as a potent inhibitor of photosynthetic electron transport. The major site of its action was identified to be the Q(B; the secondary quinone moiety of photosystemII complex. Analysis of photoreduction kinetics of para-benzoquinone and duroquinone suggest that the inhibition leads to formation of low pool of plastoquinol, which becomes limiting for electron flow through photosystemI. Further it was ascertained that the in vivo inhibitory effect appeared as a consequence of the formation of an unidentified artemisinin-metabolite rather than by the interaction of the compound per se. The putative metabolite of artemisinin is highly reactive in instituting the inhibition of photosynthetic electron flow eventually reducing the plant growth.

  14. Effects of exogenous spermine on chlorophyll fluorescence, antioxidant system and ultrastructure of chloroplasts in Cucumis sativus L. under salt stress.

    Science.gov (United States)

    Shu, Sheng; Yuan, Ling-Yun; Guo, Shi-Rong; Sun, Jin; Yuan, Ying-Hui

    2013-02-01

    The effects of exogenous spermine (Spm) on plant growth, chlorophyll fluorescence, ultrastructure and anti-oxidative metabolism of chloroplasts were investigated in Cucumis sativus L. under NaCl stress. Salt stress significantly reduced plant growth, chlorophylls content and F(v)/F(m). These changes could be alleviated by foliar spraying with Spm. Salt stress caused an increase in malondialdehyde (MDA) content and superoxide anion [Formula: see text] generation rate in chloroplasts. Application of Spm significantly increased activities of superoxidase dismutase (SOD, EC 1.15.1.1), peroxidase (POD, EC 1.11.1.7), and ascorbate peroxidase (APX, EC 1.11.1.11) which decreased the levels of [Formula: see text] and MDA in the salt-stressed chloroplasts. Salt stress decreased the activities of dehydroascorbate reductase (DHAR, EC 1.8.5.1) and glutathione reductase (GR, EC 1.6.4.2) in the chloroplasts and reduced the contents of dehydroascorbate (DAsA) and glutathione (GSH), but increased monodehydroascorbate reductase (MDAR, EC 1.6.5.4) activity. On the other hand, Spm significantly increased the activities of antioxidant enzymes and levels of antioxidants in the salt-stressed chloroplasts. Further analysis of the ultrastructure of chloroplasts indicated that salinity induced destruction of the chloroplast envelope and increased the number of plastoglobuli with aberrations in thylakoid membranes. However, Spm application to salt-stressed plant leaves counteracted the adverse effects of salinity on the structure of the photosynthetic apparatus. These results suggest that Spm alleviates salt-induced oxidative stress through regulating antioxidant systems in chloroplasts of cucumber seedlings, which is associated with an improvement of the photochemical efficiency of PSII.

  15. Changes in chloroplast lipid contents and chloroplast ultrastructure in Sulla carnosa and Sulla coronaria leaves under salt stress.

    Science.gov (United States)

    Bejaoui, Fatma; Salas, Joaquín J; Nouairi, Issam; Smaoui, Abderrazak; Abdelly, Chedly; Martínez-Force, Enrique; Youssef, Nabil Ben

    2016-07-01

    The possible involvement of chloroplast lipids in the mechanisms of NaCl tolerance was studied in leaves of two varieties of Fabaceae: Sulla carnosa and Sulla coronaria, which were subjected to 200mM NaCl over 20days. Changes in membrane lipid peroxidation, chloroplast lipids content, fatty acids (FA) composition and the ultrastructure of chloroplasts under salt stress were investigated. Chloroplast lipids were separated and quantified by high performance liquid chromatography coupled to evaporative light scattering detection (HPLC/ELSD). The results showed that salinity induced a significant decrease in digalactosyldiacylglycerol (DGDG), phosphatidylglycerol (PG) and sulfoquinovosylglycerol (SQDG) content in both S. carnosa and S. coronaria leaves, whereas monogalactosyldiacylglycerol (MGDG) content did not change significantly in S. carnosa leaves. The MGDG/DGDG ratio remained stable in S. coronaria leaves but increased in those of S. carnosa. In addition, the unsaturated-to-saturated fatty acids ratio (UFAs:SFAs) did not change under salt stress in S. coronaria leaves, while it decreased significantly in S. carnosa leaves. Moreover, salinity did not induce significant changes in MGDG and DGDG unsaturation level in S. carnosa leaves, in contrast to S. coronaria, in which salinity seems to enhance the unsaturation level in MGDG, DGDG and PG. Furthermore, the level of membrane lipid peroxidation, as expressed by malondialdehyde (MDA) levels, increased at 200mM in S. carnosa leaves, while it did not change significantly in those of S. coronaria. With respect to the ultrastructure of chloroplasts at 200mM NaCl, investigated by transmission electron microscopy (TEM), salt-stress caused the swelling of thylakoids in S. carnosa mesophyll. These ultrastructural changes were observed especially in the spongy tissue in S. coronaria. Taken together, these findings suggest that the stability of MGDG/DGDG ratio, the unchanged unsaturation level, and increasing unsaturation

  16. Maize ZmFDR3 localized in chloroplasts is involved in iron transport

    Institute of Scientific and Technical Information of China (English)

    HAN JianHui; SONG XiuFang; LI Peng; YANG HuiJun; YIN LiPing

    2009-01-01

    Iron is an essential nutrient for plant metabolism such that Fe-limited plants display chlorosis and suffer from reduced photosynthetic efficiency. Differential display previously identified genes whose expression was elevated in Fe-deficient maize roots. Here, we describe the functional characterization of one of the genes identified in the screen, ZmFDR3 (Zea maize Fe-deficiency-related). Heterologous functional complementation assays using a yeast iron uptake mutant showed that ZmFDR3 functions in iron transport. ZmFDR3 contains a domain found in FliN-proteins of the type Ⅲ secretion system and is predicted to localize to the thylakoid of plastids. Fluorescence immunocytochemistry showed that ZmFDR3 is localized in the plastids of roots, stems and leaves, with high expression found in guard cell chloroplasts. Transgenic tobacco expressing a 355-ZmFDR3 construct contains elevated iron content, displays well arranged thylakoid membranes and has photosynthetic indices that are higher than those of the wild type. Together, these results suggest that ZmFDR3 functions in chloroplast iron transport.

  17. Maize ZmFDR3 localized in chloroplasts is involved in iron transport

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    Iron is an essential nutrient for plant metabolism such that Fe-limited plants display chlorosis and suffer from reduced photosynthetic efficiency. Differential display previously identified genes whose expression was elevated in Fe-deficient maize roots. Here,we describe the functional characterization of one of the genes identified in the screen,ZmFDR3 (Zea maize Fe-deficiency-related). Heterologous functional complementation assays using a yeast iron uptake mutant showed that ZmFDR3 functions in iron transport. ZmFDR3 contains a domain found in FliN-proteins of the type III secretion system and is predicted to localize to the thylakoid of plastids. Fluorescence immunocytochemistry showed that ZmFDR3 is localized in the plastids of roots,stems and leaves,with high expression found in guard cell chloroplasts. Transgenic tobacco expressing a 35S-ZmFDR3 construct contains elevated iron content,displays well arranged thylakoid membranes and has photosynthetic indices that are higher than those of the wild type. Together,these results suggest that ZmFDR3 functions in chloroplast iron transport.

  18. Subunit movements in single membrane-bound H+-ATP synthases from chloroplasts during ATP synthesis.

    Science.gov (United States)

    Bienert, Roland; Rombach-Riegraf, Verena; Diez, Manuel; Gräber, Peter

    2009-12-25

    Subunit movements within the H(+)-ATP synthase from chloroplasts (CF(0)F(1)) are investigated during ATP synthesis. The gamma-subunit (gammaCys-322) is covalently labeled with a fluorescence donor (ATTO532). A fluorescence acceptor (adenosine 5'-(beta,gamma-imino)triphosphate (AMPPNP)-ATTO665) is noncovalently bound to a noncatalytic site at one alpha-subunit. The labeled CF(0)F(1) is integrated into liposomes, and a transmembrane pH difference is generated by an acid base transition. Single-pair fluorescence resonance energy transfer is measured in freely diffusing proteoliposomes with a confocal two-channel microscope. The fluorescence time traces reveal a repetitive three-step rotation of the gamma-subunit relative to the alpha-subunit during ATP synthesis. Some traces show splitting into sublevels with fluctuations between the sublevels. During catalysis the central stalk interacts, with equal probability, with each alphabeta-pair. Without catalysis the central stalk interacts with only one specific alphabeta-pair, and no stepping between FRET levels is observed. Two inactive states of the enzyme are identified: one in the presence of AMPPNP and one in the presence of ADP.

  19. Suborganellar localisation and effect of light on Helianthus tuberosus chloroplast transglutaminases and their substrates.

    Science.gov (United States)

    Dondini, L; Del Duca, S; Dall'Agata, L; Bassi, R; Gastaldelli, M; Della Mea, M; Di Sandro, A; Claparols, I; Serafini-Fracassini, D

    2003-05-01

    The light stimulation of transglutaminase (TGase EC 2.3.2.13) activity was verified by incubating isolated chloroplasts of Helianthus tuberosus L. continuously or for alternate periods of light or dark (light/dark and dark/light). The first 10 min of incubation always represented the critical period. Light-harvesting complexes of photosystem II (LHCII) were more intensely labelled by (14)C-polyamines under light and light/dark than under dark and dark/light conditions. Chloroplasts were fractionated into thylakoid- and stroma-enriched fractions in which multiple TGase forms and substrates were found. Antibodies against TGase recognised 58- and 24-kDa bands in thylakoids and a 150-kDa band in the stroma. The latter, and its 150-kDa fraction, catalysed the conjugation of 14C-polyamines to Rubisco. In both fractions (thylakoid-pre and stroma-pre) the analysis of polyamine glutamyl derivatives showed a significant light-affected conjugation of polyamines to endogenous proteins. Alternatively, entire chloroplasts were incubated and afterwards their sub-fractions were isolated (thylakoid-post and stroma-post). The PSII and LHCII complexes were more intensely immunodetected in thylakoid-post than in thylakoid-pre, especially under dark conditions. Conversely, the conjugation of polyamines to thylakoid proteins was clearly light-stimulated in thylakoid-post, and much less in thylakoid-pre. Stroma-pre proteins were poorly polyamine-conjugated and not light-affected; on the contrary, stroma-post proteins were much more polyamine-modified and strongly light-stimulated. Thus, the light-activated conjugation depends mainly on the presence of the thylakoid fraction during the assay. The protective effect on chloroplasts under photo-damage, stress or senescence conditions attributed in the literature to free polyamines is discussed with regard to the occurrence of polyamine conjugates catalysed by TGases.

  20. Coat protein mutations in an attenuated Cucumber mosaic virus encoding mutant 2b protein that lacks RNA silencing suppressor activity induces chlorosis with photosynthesis gene repression and chloroplast abnormalities in infected tobacco plants.

    Science.gov (United States)

    Mochizuki, Tomofumi; Yamazaki, Ryota; Wada, Tomoya; Ohki, Satoshi T

    2014-05-01

    In tobacco plants, the Cucumber mosaic virus (CMV) pepo strain induces mosaic symptoms, including pale green chlorosis and malformed tissues. Here, we characterized the involvement of 2b protein and coat protein (CP) in the development of mosaic symptoms. A 2b mutant (R46C) that lacks viral suppressor of RNA silencing (VSR) activity showed an asymptomatic phenotype with low levels of virus accumulation. Tomato spotted wilt virus NSs protein did not complement the virulence of the R46C, although it did restore high-level virus accumulation. However, R46C mutants expressing mutated CP in which the amino acid P129 was mutated to A, E, C, Q, or S induced chlorosis that was associated with reduced expression of chloroplast and photosynthesis related genes (CPRGs) and abnormal chloroplasts with fewer thylakoid membranes. These results suggest that the CP of the CMV pepo strain acquires virulence by amino acid mutations, which causes CPRG repression and chloroplast abnormalities.

  1. The selective biotin tagging and thermolysin proteolysis of chloroplast outer envelope proteins reveals information on protein topology and association into complexes

    Directory of Open Access Journals (Sweden)

    Hélène eHardré

    2014-05-01

    Full Text Available The understanding of chloroplast function requires the precise localization of proteins in each of its sub-compartments. High-sensitivity mass spectrometry has allowed the inventory of proteins in thylakoid, stroma and envelope fractions. Concerning membrane association, proteins can be either integral or peripheral or even soluble proteins bound transiently to a membrane complex. We sought a method providing information at the surface of the outer envelope membrane (OEM, based on specific tagging with biotin or proteolysis using thermolysin, a non-membrane permeable protease. To evaluate this method, envelope, thylakoid and stroma proteins were separated by two-dimensional electrophoresis and analyzed by immunostaining and mass spectrometry. A short selection of proteins associated to the chloroplast envelope fraction was checked after superficial treatments of intact chloroplasts. We showed that this method could allow the characterization of OEM embedded proteins facing the cytosol, as well as peripheral and soluble proteins associated via tight or lose interactions. Some stromal proteins were associated with biotinylated spots and analyzes are still needed to determine whether polypeptides were tagged prior import or if they co-migrated with OEM proteins. This method also suggests that some proteins associated with the inner envelope membrane (IEM might need the integrity of a trans-envelope (IEM-OEM protein complex (e.g. division ring-forming components or at least an intact OEM partner. Following this evaluation, proteomic analyzes should be refined and the putative role of inter-membrane space components stabilizing trans-envelope complexes demonstrated. For future comprehensive studies, perspectives include the dynamic analyses of OEM proteins and IEM-OEM complexes in various physiological contexts and using virtually any other purified membrane organelle.

  2. Monogalactosyldiacylglycerol synthesis in the outer envelope membrane of chloroplasts is required for enhanced growth under sucrose supplementation

    Directory of Open Access Journals (Sweden)

    Masato eMurakawa

    2014-06-01

    Full Text Available Plant galactolipid synthesis on the outer envelope membranes of chloroplasts is an important biosynthetic pathway for sustained growth under conditions of phosphate (Pi depletion. During Pi starvation, the amount of digalactosyldiacylglycerol (DGDG is increased to substitute for the phospholipids that are degraded for supplying Pi. An increase in DGDG concentration depends on an adequate supply of monogalactosyldiacylglycerol (MGDG, which is a substrate for DGDG synthesis and is synthesized by a type-B MGDG synthase, MGD3. Recently, sucrose was suggested to be a global regulator of plant responses to Pi starvation. Thus, we analyzed expression levels of several genes involved in lipid remodeling during Pi starvation in Arabidopsis thaliana and found that the abundance of MGD3 mRNA increased when sucrose was exogenously supplied to the growth medium. Sucrose supplementation retarded the growth of the Arabidopsis MGD3 knockout mutant mgd3 but enhanced the growth of transgenic Arabidopsis plants overexpressing MGD3 compared with wild type, indicating the involvement of MGD3 in plant growth under sucrose-replete conditions. Although most features such as chlorophyll content, photosynthetic activity, and Pi content were comparable between wild-type and the transgenic plants overexpressing MGD3, sucrose content in shoot tissues decreased and incorporation of exogenously supplied carbon to DGDG was enhanced in the MGD3-overexpressing plants compared with wild type. Our results suggest that MGD3 plays an important role in supplying DGDG as a component of extraplastidial membranes to support enhanced plant growth under conditions of carbon excess.

  3. CHLOROPLAST STRUCTURAL AND FUNCTIONAL CHANGES AS BIOMARKERS OF HEAVY METAL CONTAMINATION

    Directory of Open Access Journals (Sweden)

    M. V.

    2016-02-01

    Full Text Available The aim was to confirm the hypothesis of possibility to use the chloroplast structural and functional changes in higher plants as biomarkers to assess heavy metal contamination. Chloroplast ultra-structural changes of Pisum sativum L were detected using the transmission electron microscopy. This work deals with studies of chloroplast structure responses to a high content of copper (250 μmМ and zinc (400 μmМ. Data on changes in the structure of chloroplasts in particular, heterogeneity in the grain thylakoid packing, increase of interthylakoid gaps and thickness of chloroplast grain thylakoids in comparison with controls were obtained. The results of studies on structural and functional chloroplasts changes offer challenges for their use as markers for an early diagnostics of abiotic stress effects and in biotechnological studies to produce novel advanced varieties of crops resistant to stress.

  4. Dietary thylakoids suppress blood glucose and modulate appetite-regulating hormones in pigs exposed to oral glucose tolerance test

    DEFF Research Database (Denmark)

    Montelius, Caroline; Szwiec, Katarzyna; Kardas, Marek

    2014-01-01

    BACKGROUND & AIMS: Dietary chloroplast thylakoids have previously been found to reduce food intake and body weight in animal models, and to change metabolic profiles in humans in mixed-food meal studies. The aim of this study was to investigate the modulatory effects of thylakoids on glucose...... metabolism and appetite-regulating hormones during an oral glucose tolerance test in pigs fed a high fat diet. METHODS: Six pigs were fed a high fat diet (36 energy% fat) for one month before oral glucose tolerance test (1 g/kg d-glucose) was performed. The experiment was designed as a cross-over study......, and decreased late postprandial secretion of ghrelin. CONCLUSION: Dietary thylakoids may be a novel agent in reducing the glycaemic responses to high carbohydrate and high glycaemic index foods. Thylakoids may in the future be promising for treatment and prevention of diabetes, overweight and obesity....

  5. Coherent X-Ray Diffraction Imaging of Chloroplasts from Cyanidioschyzon merolae by Using X-Ray Free Electron Laser.

    Science.gov (United States)

    Takayama, Yuki; Inui, Yayoi; Sekiguchi, Yuki; Kobayashi, Amane; Oroguchi, Tomotaka; Yamamoto, Masaki; Matsunaga, Sachihiro; Nakasako, Masayoshi

    2015-07-01

    Coherent X-ray diffraction imaging (CXDI) is a lens-less technique for visualizing the structures of non-crystalline particles with the dimensions of submicrometer to micrometer at a resolution of several tens of nanometers. We conducted cryogenic CXDI experiments at 66 K to visualize the internal structures of frozen-hydrated chloroplasts of Cyanidioschyzon merolae using X-ray free electron laser (XFEL) as a coherent X-ray source. Chloroplast dispersed specimen disks at a number density of 7/(10×10 µm(2)) were flash-cooled with liquid ethane without staining, sectioning or chemical labeling. Chloroplasts are destroyed at atomic level immediately after the diffraction by XFEL pulses. Thus, diffraction patterns with a good signal-to-noise ratio from single chloroplasts were selected from many diffraction patterns collected through scanning specimen disks to provide fresh specimens into the irradiation area. The electron density maps of single chloroplasts projected along the direction of the incident X-ray beam were reconstructed by using the iterative phase-retrieval method and multivariate analyses. The electron density map at a resolution of 70 nm appeared as a C-shape. In addition, the fluorescence image of proteins stained with Flamingo™ dye also appeared as a C-shape as did the autofluorescence from Chl. The similar images suggest that the thylakoid membranes with an abundance of proteins distribute along the outer membranes of chloroplasts. To confirm the present results statistically, a number of projection structures must be accumulated through high-throughput data collection in the near future. Based on the results, we discuss the feasibility of XFEL-CXDI experiments in the structural analyses of cellular organelles.

  6. Acute Effects of a Spinach Extract Rich in Thylakoids on Satiety: A Randomized Controlled Crossover Trial

    OpenAIRE

    Rebello, Candida J.; Chu, Jessica; Beyl, Robbie; Edwall, Dan; Erlanson-Albertsson, Charlotte; Frank L. Greenway

    2015-01-01

    Objective: By retarding fat digestion, thylakoids, the internal photosynthetic membrane system of green plants, promote the release of satiety hormones. This study examined the effect of consuming a single dose of concentrated extract of thylakoids from spinach on satiety, food intake, lipids, and glucose compared to a placebo. Design: Sixty overweight and obese individuals enrolled in a double-blind randomized crossover study consumed the spinach extract or placebo in random order at least a...

  7. Phenol homeostasis is ensured in vanilla fruit by storage under solid form in a new chloroplast-derived organelle, the phenyloplast.

    Science.gov (United States)

    Brillouet, Jean-Marc; Verdeil, Jean-Luc; Odoux, Eric; Lartaud, Marc; Grisoni, Michel; Conéjéro, Geneviève

    2014-06-01

    A multiple cell imaging approach combining immunofluorescence by confocal microscopy, fluorescence spectral analysis by multiphotonic microscopy, and transmission electron microscopy identified the site of accumulation of 4-O-(3-methoxybenzaldehyde) β-d-glucoside, a phenol glucoside massively stockpiled by vanilla fruit. The glucoside is sufficiently abundant to be detected by spectral analysis of its autofluorescence. The convergent results obtained by these different techniques demonstrated that the phenol glucoside accumulates in the inner volume of redifferentiating chloroplasts as solid amorphous deposits, thus ensuring phenylglucoside cell homeostasis. Redifferentiation starts with the generation of loculi between thylakoid membranes which are progressively filled with the glucoside until a fully matured organelle is obtained. This peculiar mode of storage of a phenolic secondary metabolite is suspected to occur in other plants and its generalization in the Plantae could be considered. This new chloroplast-derived organelle is referred to as a 'phenyloplast'.

  8. Increase in electron transfer activity in photosystem Ⅱ of spinach thylakoids caused by conversion of phosphatidyl-glycerol to phosphatidic acid molecules

    Institute of Scientific and Technical Information of China (English)

    WU Feng; YANG Zhenle; LI Liangbi; KUANG Tingyun

    2003-01-01

    The techniques of oxygen electrode polarography, sodium dodecyl sulfate-polyacryamide gel electrophoresis (SDS-PAGE) and thin layer chromatography (TLC) were employed to investigate the effect of phospholipase D treatment on physiological function of spinach thylakoids. It was shown that the phospholipase D treatment on thylakoid resulted in the degradation of phosphatidylglycerol (PG) and occurrence of phosphatidic acid (PA). The changes of PG to PA molecules caused an increase in oxygen evolution in photosystem Ⅱ (PSⅡ), which was accompanied by an uncoupling effect on thylakoid membrane. It was revealed that the head-groups of PG molecules play an important role in the maintenance of the appropriate physiological activity of thylakoid membrane.

  9. Molecular analysis of a thylakoid K+ channel

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    2000-05-01

    The work undertaken during the prior granting period sought to use a novel probe to identify and clone plant ion (K) channels. It was also proposed that in vitro biochemical studies of cation transport across purified preparations of thylakoid membrane be employed to characterize a putative K channel in this membrane system. Over the last several years (including those of the previous grant period), an enormous data base of partially-sequenced mRNAs and numerous genomes (including those of plants) has evolved and provides a powerful alternative to this brute-force approach to identify and clone cDNAs ending physiologically important membrane proteins such as channels. The utility of searching genetic databases for relevant sequences, in addition to the difficulty of working with membrane proteins, led to changes in research focus during the prior granting period, and has resulted in the identification of a new class of plant ion channels, which will be the focus of research during the proposed new granting period.

  10. Nitrogen control of chloroplast differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Schmidt, G.W.

    1992-07-01

    This project is directed toward understanding how the availability of nitrogen affects the accumulation of chloroplast pigments and proteins functioning in energy transduction and carbon metabolism. Molecular analyses performed with Chlamydomonas reinhardtii grown in a continuous culture system such that ammonium concentration is maintained at a low steady-state concentration so as to limit cell division. As compared to chloroplasts from cells of non-limiting nitrogen provisions, chloroplasts of N-limited cells are profoundly chlorophyll-deficient but still assimilate carbon for deposition of as starch and as storage lipids. Chlorophyll deficiency arises by limiting accumulation of appropriate nuclear-encoded mRNAs of and by depressed rates of translation of chloroplast mRNAs for apoproteins of reaction centers. Chloroplast translational effects can be partially ascribed to diminished rates of chlorophyll biosynthesis in N-limited cells, but pigment levels are not determinants for expression of the nuclear light-harvesting protein genes. Consequently, other signals that are responsive to nitrogen availability mediate transcriptional or post-transcriptional processes for accumulation of the mRNAs for LHC apoproteins and other mRNAs whose abundance is dependent upon high nitrogen levels. Conversely, limited nitrogen availability promotes accumulation of other proteins involved in carbon metabolism and oxidative electron transport in chloroplasts. Hence, thylakoids of N-limited cells exhibit enhanced chlororespiratory activities wherein oxygen serves as the electron acceptor in a pathway that involves plastoquinone and other electron carrier proteins that remain to be thoroughly characterized. Ongoing and future studies are also outlined.

  11. Two roles of thylakoid lipids in modifying the activity of herbicides which inhibit photosystem II

    Energy Technology Data Exchange (ETDEWEB)

    Kupatt, C.C. Jr.

    1985-01-01

    Thylakoid lipids may modify the activity of herbicides which inhibit electron transport at the Q/sub B/ protein of photosystem II in two ways: (1) lipids can act as a hydrophobic barrier to a binding site localized close to the loculus of the membrane, and (2) changes in lipid composition can reduce the ability of inhibitors to block electron transport, possibly due to a change in the conformation of the Q/sub B/ protein. The herbicide binding site was localized close to the locular side of the thylakoid membrane by determining the activity of a number of substituted phenylurea and s-triazine herbicides in inverted and non-inverted thylakoids. Quantitative structure-activity relationship analysis showed that inversion of thylakoids reduced the requirement of molecular lipophilicity deemed necessary for phenylurea activity in non-inverted membranes, whereas s-triazines exhibited no differences in the lipophilicity requirement in thylakoid membranes of either orientation. The binding affinity of /sup 14/C-diuron was reduced in bicarbonate-depleted thylakoids relative to reconstituted or control membranes, as is the case with atrazine binding. These observations support a model of the herbicide binding site containing both common and herbicide family specific binding domains. Thylakoids isolated either from detached lambs quarters (Chenopodium album L.) leaves, treated with SAN 6706, or from soybean (Glycine max L.), with norflurazon or pyrazon applied preemergence, exhibited decreased susceptibility to atrazine. The ability of lipid-modifying treatments to decrease the atrazine susceptibility of field-grown soybeans was also investigated.

  12. Blue-light mediated accumulation of nuclear-encoded transcripts coding for proteins of the thylakoid membrane is absent in the phytochrome-deficient aurea mutant of tomato.

    Science.gov (United States)

    Oelmüller, R; Kendrick, R E; Briggs, W R

    1989-08-01

    Polyclonal antibodies against pea phytochrome detect 2 protein bands (about 116 and 120 kDa) on blots of crude protein extracts and protein of microsomal preparations of dark-grown tomato seedlings. Both protein bands are undetectable in Western blots of the aurea mutant extracts. Neither protein band is detectable after isogenic wild-type seedlings are illuminated with 3 h of red light, either in the crude extract or in the membrane fraction of the irradiated seedlings; this result is consistent with the hypothesis that both bands are phytochrome. When dark-grown wild-type seedlings are illuminated with 3 h of red light or blue light against a red light background, the transcript levels for chlorophyll a/b-binding proteins of photosystem I and II, plastocyanin, and the subunit II of photosystem I increase. In all cases, the same fluence rate of blue light is much more effective than red light alone, a result that indicates the involvement of a blue/UV-A light photoreceptor in addition to the involvement of the far-red-absorbing form of phytochrome, Pfr. The aurea mutant responds neither to red light nor to blue light. Thus, no Pfr-independent induction of the four transcripts by a blue/UV-A light photoreceptor can be measured in the aurea mutant.

  13. Identification and Expression Analysis of Chloroplast p-psbB Gene Differentially Expressed in Wild Ginseng

    Directory of Open Access Journals (Sweden)

    Doo-Young Kim

    2012-03-01

    Full Text Available Panax ginseng is a well-known herbal medicine in traditional Asian medicine. Although wild ginseng is widely accepted to be more active than cultivated ginseng in chemoprevention,little has actually been reported on the difference between wild ginseng and cultivated ginseng. Using suppressive subtraction hybridization, we cloned the p-psbB gene as a candidate target gene for a wild ginseng-specific gene. Here, we report that one of the clones isolated in this screen was the chloroplast p-psbB gene, a chlorophyll a-binding inner antenna protein in the photosystem II complex, located in the lipid matrix of the thylakoid membrane. Real-time results showed that the expression of the p-psbB gene was significantly up-regulated in wild ginseng as compared to cultivated ginseng. Thus, the p-psbB gene may be one of the important markers of wild ginseng.

  14. The rapid yellowing of spruce at a mountain site in the Central Black Forest (Germany). Combined effects of Mg deficiency and ozone on biochemical, physiological and structural properties of the chloroplasts.

    Science.gov (United States)

    Siefermann-Harms, Dorothea; Boxler-Baldoma, Carmen; von Wilpert, Klaus; Heumann, Hans-Günther

    2004-04-01

    Biochemical, physiological and ultrastructural changes of the chloroplasts were examined in the course of the rapid yellowing process of spruce (Picea abies (L.) Karst.) at a Mg-deficient and ozone polluted mountain site (Schöllkopf mountain, Central Black Forest, Germany, 840 m a.s.l.). While at an early stage of yellowing the chlorophyll (Chl) content of the needles decreased slowly, significant changes occurred in the chloroplasts: The lability of the light-harvesting Chl a/b protein complex LHC II increased; the thylakoid cross-sectional area of chloroplasts in the outer mesophyll of the needles decreased, and their Chl fluorescence showed typical changes like the decrease of Fv/Fm and the increase of the photoinhibitory Fv quenching. Later on, the Chl content decreased rapidly, the changes in the chloroplasts continued and the needles turned yellow. Lutein and the pigments of the xanthophyll cycle were enhanced in relation to Chl a. Light and dark reactions of the xanthophyll cycle were highly active indicating efficient proton pumping and NADPH formation. The ratio of nonappressed to appressed thylakoid membranes increased with decreasing Fv/Fm suggesting that structural and fluorescence properties of the chloroplasts were related. The response of the needles to defined shading and improved Mg supply was also examined. The combined effects of strong sun light, low levels of non-Chl-bound Mg (Mg(free)) and ozone concentrations exceeding 80 microg m(-3) are shown to be necessary to induce the rapid yellowing process. For needles with Mg(free) < 0.12 mg g(-1) needle dry matter, the lability of the LHC II was correlated with the ozone concentration suggesting that the destabilization of the LHC II plays a central role in the rapid yellowing process.

  15. Chloroplast actin filaments organize meshwork on the photorelocated chloroplasts in the moss Physcomitrella patens.

    Science.gov (United States)

    Yamashita, Hiroko; Sato, Yoshikatsu; Kanegae, Takeshi; Kagawa, Takatoshi; Wada, Masamitsu; Kadota, Akeo

    2011-02-01

    Cytoskeleton dynamics during phototropin-dependent chloroplast photorelocation movement was analyzed in protonemal cells of actin- and microtubule-visualized lines of Physcomitrella patens expressing GFP- or tdTomato-talin and GFP-tubulin. Using newly developed epi- and trans-microbeam irradiation systems that permit fluorescence observation of the cell under blue microbeam irradiation inducing chloroplast relocation, it was revealed that meshwork of actin filaments formed at the chloroplast-accumulating area both in the avoidance and accumulation movements. The structure disappeared soon when blue microbeam was turned off, and it was not induced under red microbeam irradiation that did not evoke chloroplast relocation movement. In contrast, no apparent change in microtubule organization was detected during the movements. The actin meshwork was composed of short actin filaments distinct from the cytoplasmic long actin cables and was present between the chloroplasts and plasma membrane. The short actin filaments emerged from around the chloroplast periphery towards the center of chloroplast. Showing highly dynamic behavior, the chloroplast actin filaments (cp-actin filaments) were rapidly organized into meshwork on the chloroplast surface facing plasma membrane. The actin filament configuration on a chloroplast led to the formation of actin meshwork area in the cell as the chloroplasts arrived at and occupied the area. After establishment of the meshwork, cp-actin filaments were still highly dynamic, showing appearance, disappearance, severing and bundling of filaments. These results indicate that the cp-actin filaments have significant roles in the chloroplast movement and positioning in the cell.

  16. Influence of lanthanum on chloroplast ultrastructure of soybean leaves under ultraviolet-B stress

    Institute of Scientific and Technical Information of China (English)

    PENG Qian; ZHOU Qing

    2009-01-01

    In order to investigate the effects of lanthanum(Ⅲ) on cell ultrastructure of soybean leaves under elevated ultraviolet-B irradiation (UV-B, 280-320 rim), the chloroplast ultrastructure of soybean seedlings was studied by hydroponics under laboratory conditions. The re-sults showed that the thylakoid in chloroplast was orderly and clearly as soybean leaves were pretreated by La(Ⅲ). The thylakoid was indis-tinctly disordered, expanded and even indiscoverable in the chloroplast under UV-B stress. The impact on the thylakoid by the high in-tensity UV-B irradiation (T2) was bigger than that by the low intensity UV-B irradiation (T1). However, the destruction of the chloroplast structure caused by UV-B stress was alleviated by La(Ⅲ), and the arrangement of the thylakoid in the chloroplast became orderly and clearly. The effect of the alleviation by La(Ⅲ) under the low intensity UV-B irradiation (T1) was better than that under the high intensity UV-B irradiation (T2).

  17. Consumption of thylakoid-rich spinach extract reduces hunger, increases satiety and reduces cravings for palatable food in overweight women.

    Science.gov (United States)

    Stenblom, Eva-Lena; Egecioglu, Emil; Landin-Olsson, Mona; Erlanson-Albertsson, Charlotte

    2015-08-01

    Green-plant membranes, thylakoids, have previously been found to increase postprandial release of the satiety hormone GLP-1, implicated in reward signaling. The purpose of this study was to investigate how treatment with a single dose of thylakoids before breakfast affects homeostatic as well as hedonic hunger, measured as wanting and liking for palatable food (VAS). We also examined whether treatment effects were correlated to scores for eating behavior. Compared to placebo, intake of thylakoids significantly reduced hunger (21% reduction, p satiety (14% increase, p hunger, associated with overeating and obesity. Individuals scoring higher for emotional eating behavior may have enhanced treatment effect on cravings for palatable food.

  18. PSB27: A thylakoid protein enabling Arabidopsis to adapt to changing light intensity

    Energy Technology Data Exchange (ETDEWEB)

    Hou, Xin [Univ. of California, Berkeley, CA (United States). Dept of Plant and Microbial Biology; Garcia, Veder J. [Univ. of California, Berkeley, CA (United States). Dept of Plant and Microbial Biology; Buchanan, Bob B. [Univ. of California, Berkeley, CA (United States). Dept of Plant and Microbial Biology; Luan, Sheng [Univ. of California, Berkeley, CA (United States). Dept of Plant and Microbial Biology

    2016-08-22

    Project Title: Immunophilins in the assembly and maintenance of photosynthetic electron transport chain in Arabidopsis Applicant: The Regents of the University of California PI: Sheng Luan, University of California at Berkeley Photosynthetic light energy conversion entails coordinated function of complex molecular machines that capture and convert light energy into chemical forms through photosynthetic electron transport chain. Each molecular machine, such as photosystem II (PSII), may consist of dozens of protein subunits and small molecule cofactors. Despite advanced understanding of the structure and function of these complexes, little is known about “How individual proteins and cofactors assemble into a functional machine and how do these molecular machines maintain their structure and function under a highly hazardous lumenal environment.” Our studies on immunophilins have unexpectedly contributed to the understanding of this question. Originally defined as cellular receptors for immunosuppressants, immunophilins have been discovered in a wide range of organisms from bacteria, fungi, plants, to animals. Immunophilins function in protein folding processes as chaperones and foldases. Arabidopsis genome encodes ca. 50 immunophilins. The most striking finding is that 16 immunophilin members are targeted to chloroplast thylakoid lumen, by far the largest group in the lumenal proteome. What is the function of immunophilins in the thylakoid lumen? Our studies have demonstrated critical roles for several immunophilins in the biogenesis and maintenance of photosynthetic complexes such as PSII. These studies have made a critical link between immunophilins and the assembly of photosynthetic machines and thus opened up a new area of research in photosynthesis. Our goal is to dissect the roles of immunophilins and their partners in the assembly and maintenance of the photosynthetic electron transport chain. The specific objectives for this funding period will be: 1. To

  19. Calcineurin B-like Protein CBL10 Directly Interacts with TOC34 (Translocon of the Outer Membrane of the Chloroplasts) and Decreases Its GTPase Activity in Arabidopsis

    Science.gov (United States)

    Cho, Joo Hyuk; Lee, Jeong Hwan; Park, Yoon Kook; Choi, Mi Na; Kim, Kyung-Nam

    2016-01-01

    As calcium sensor relays in plants, calcineurin B-like (CBL) proteins provide an important contribution to decoding Ca2+ signatures elicited by a variety of abiotic stresses. Currently, it is well known that CBLs perceive and transmit the Ca2+ signals mainly to a group of serine/threonine protein kinases called CBL-interacting protein kinases (CIPKs). In this study, we report that the CBL10 member of this family has a novel interaction partner besides the CIPK proteins. Yeast two-hybrid screening with CBL10 as bait identified an Arabidopsis cDNA clone encoding a TOC34 protein, which is a member of the TOC (Translocon of the Outer membrane of the Chloroplasts) complex and possesses the GTPase activity. Further analyses showed that in addition to CBL10, CBL7 also interacts with TOC34 at much lower strength in the yeast two-hybrid system. However, the rest of the CBL family members failed to interact with TOC34. Bimolecular fluorescence complementation (BiFC) analysis verified that the CBL10-TOC34 interaction occurs at the outer membrane of chloroplasts in vivo. In addition, we also demonstrated that CBL10 physically associates with TOC34 in vitro, resulting in a significant decrease in the GTPase activity of the TOC34 protein. Taken together, our findings clearly indicate that a member of the CBL family, CBL10, can modulate not only the CIPK members but also TOC34, allowing the CBL family to relay the Ca2+ signals in more diverse ways than currently known. PMID:28018422

  20. The Primitive Thylakoid-Less Cyanobacterium Gloeobacter Is a Common Rock-Dwelling Organism.

    Directory of Open Access Journals (Sweden)

    Jan Mareš

    Full Text Available Cyanobacteria are an ancient group of photosynthetic prokaryotes, which are significant in biogeochemical cycles. The most primitive among living cyanobacteria, Gloeobacter violaceus, shows a unique ancestral cell organization with a complete absence of inner membranes (thylakoids and an uncommon structure of the photosynthetic apparatus. Numerous phylogenetic papers proved its basal position among all of the organisms and organelles capable of plant-like photosynthesis (i.e., cyanobacteria, chloroplasts of algae and plants. Hence, G. violaceus has become one of the key species in evolutionary study of photosynthetic life. It also numbers among the most widely used organisms in experimental photosynthesis research. Except for a few related culture isolates, there has been little data on the actual biology of Gloeobacter, being relegated to an "evolutionary curiosity" with an enigmatic identity. Here we show that members of the genus Gloeobacter probably are common rock-dwelling cyanobacteria. On the basis of morphological, ultrastructural, pigment, and phylogenetic comparisons of available Gloeobacter strains, as well as on the basis of three new independent isolates and historical type specimen, we have produced strong evidence as to the close relationship of Gloeobacter to a long known rock-dwelling cyanobacterial morphospecies Aphanothece caldariorum. Our results bring new clues to solving the 40 year old puzzle of the true biological identity of Gloeobacter violaceus, a model organism with a high value in several biological disciplines. A probable broader distribution of Gloeobacter in common wet-rock habitats worldwide is suggested by our data, and its ecological meaning is discussed taking into consideration the background of cyanobacterial evolution. We provide observations of previously unknown genetic variability and phenotypic plasticity, which we expect to be utilized by experimental and evolutionary researchers worldwide.

  1. The Primitive Thylakoid-Less Cyanobacterium Gloeobacter Is a Common Rock-Dwelling Organism

    Science.gov (United States)

    Mareš, Jan; Hrouzek, Pavel; Kaňa, Radek; Ventura, Stefano; Strunecký, Otakar; Komárek, Jiří

    2013-01-01

    Cyanobacteria are an ancient group of photosynthetic prokaryotes, which are significant in biogeochemical cycles. The most primitive among living cyanobacteria, Gloeobacter violaceus, shows a unique ancestral cell organization with a complete absence of inner membranes (thylakoids) and an uncommon structure of the photosynthetic apparatus. Numerous phylogenetic papers proved its basal position among all of the organisms and organelles capable of plant-like photosynthesis (i.e., cyanobacteria, chloroplasts of algae and plants). Hence, G. violaceus has become one of the key species in evolutionary study of photosynthetic life. It also numbers among the most widely used organisms in experimental photosynthesis research. Except for a few related culture isolates, there has been little data on the actual biology of Gloeobacter, being relegated to an “evolutionary curiosity” with an enigmatic identity. Here we show that members of the genus Gloeobacter probably are common rock-dwelling cyanobacteria. On the basis of morphological, ultrastructural, pigment, and phylogenetic comparisons of available Gloeobacter strains, as well as on the basis of three new independent isolates and historical type specimen, we have produced strong evidence as to the close relationship of Gloeobacter to a long known rock-dwelling cyanobacterial morphospecies Aphanothece caldariorum. Our results bring new clues to solving the 40 year old puzzle of the true biological identity of Gloeobacter violaceus, a model organism with a high value in several biological disciplines. A probable broader distribution of Gloeobacter in common wet-rock habitats worldwide is suggested by our data, and its ecological meaning is discussed taking into consideration the background of cyanobacterial evolution. We provide observations of previously unknown genetic variability and phenotypic plasticity, which we expect to be utilized by experimental and evolutionary researchers worldwide. PMID:23823729

  2. Unassisted membrane insertion as the initial step in DeltapH/Tat-dependent protein transport.

    Science.gov (United States)

    Hou, Bo; Frielingsdorf, Stefan; Klösgen, Ralf Bernd

    2006-02-01

    In the thylakoid membrane of chloroplasts as well as in the cytoplasmic membrane of bacteria, the DeltapH/Tat-dependent protein transport pathway is responsible for the translocation of folded proteins. Using the chimeric 16/23 protein as model substrate in thylakoid transport experiments, we dissected the transport process into several distinct steps that are characterized by specific integral translocation intermediates. Formation of the early translocation intermediate Ti-1, which still exposes the N and the C terminus to the stroma, is observed with thylakoids pretreated with (i) solutions of chaotropic salts or alkaline pH, (ii) protease, or (iii) antibodies raised against TatA, TatB, or TatC. Membrane insertion takes place even into liposomes, demonstrating that proteinaceous components are not required. This suggests that Tat-dependent transport may be initiated by the unassisted insertion of the substrate into the lipid bilayer, and that interaction with the Tat translocase takes place only in later stages of the process.

  3. Nucleus-encoded light-harvesting chlorophyll a/b proteins are imported normally into chlorophyll b-free chloroplasts of Arabidopsis.

    Science.gov (United States)

    Nick, Sabine; Meurer, Jörg; Soll, Jürgen; Ankele, Elisabeth

    2013-05-01

    Chloroplast-located proteins which are encoded by the nuclear genome have to be imported from the cytosol into the organelle in a posttranslational manner. Among these nuclear-encoded chloroplast proteins are the light-harvesting chlorophyll a/b-binding proteins (LHCPs). After translation in the cytosol, precursor proteins of LHCPs are imported via the TOC/TIC translocase, processed to their mature size to insert into thylakoid membranes where they recruit chlorophylls a and b to form pigment-protein complexes. The translocation of proteins is a highly regulated process which employs several regulators. To analyze whether CAO (chlorophyll a oxigenase) which converts chlorophyll a to chlorophyll b at the inner chloroplast membrane, is one of these regulators, we performed import reactions utilizing a homozygous loss-of-function mutant (cao-1). We imported in vitro translated and (35)S-labeled precursor proteins of light-harvesting proteins of photosystem II LHCB1, LHCB4, and LHCB5 into chloroplasts isolated from cao-1 and show that import of precursor proteins and their processing to mature forms are not impaired in the mutant. Therefore, regulation of the import machinery cannot be responsible for the decreased steady-state levels of light-harvesting complex (LHC) proteins. Regulation does not take place at the transcriptional level either, because Lhcb mRNAs are not down-regulated. Additionally, reduced steady-state levels of LHCPs also do not occur due to posttranslational turnover of non-functional LHCPs in chloroplasts. Taken together, our data show that plants in the absence of CAO and therefore devoid of chlorophyll b are not influenced in their import behavior of LHC proteins.

  4. Salt shock-inducible Photosystem I cyclic electron transfer in Synechocystis PCC6803 relies on binding of ferredoxin : NADP(+) reductase to the thylakoid membranes via its CpcD phycobilisome-linker homologous N-terminal domain

    NARCIS (Netherlands)

    van Thor, JJ; Jeanjean, R; Havaux, M; Sjollema, KA; Joset, F; Hellingwerf, KJ; Matthijs, HCP

    2000-01-01

    Relative to ferredoxin:NADP(+) reductase (FNR) from chloroplasts, the comparable enzyme in cyanobacteria contains an additional 9 kDa domain at its amino-terninus, The domain is homologous to the phycocyanin associated linker polypeptide CpcD of the light harvesting phycobilisome antennae. The pheno

  5. Evolutionarily evolved discriminators in the 3-TPR domain of the Toc64 family involved in protein translocation at the outer membrane of chloroplasts and mitochondria.

    Science.gov (United States)

    Mirus, Oliver; Bionda, Tihana; von Haeseler, Arndt; Schleiff, Enrico

    2009-08-01

    Transport of polypeptides across membranes is a general and essential cellular process utilised by molecular machines. At least one component of these complexes contains a domain composed of three tetratricopeptide repeat (3-TPR) motifs. We have focussed on the receptor Toc64 to elucidate the evolved functional specifications of its 3-TPR domain. Toc64 is a component of the Toc core complex and functionally replaces Tom70 at the outer membrane of mitochondria in plants. Its 3-TPR domain recognises the conserved C-terminus of precursor-bound chaperones. We built homology models of the 3-TPR domain of chloroplastic Toc64 from different species and of the mitochondrial isoform from Arabidopsis. Guided by modelling, we identified residues essential for functional discrimination of the differently located isoforms to be located almost exclusively on the convex surface of the 3-TPR domain. The only exception is at568Ser/ps557Met, which is positioned in the ligand-binding groove. The functional implications of the homology models are discussed.

  6. Microoxic Niches within the Thylakoid Stroma of Air-Grown Chlamydomonas reinhardtii Protect [FeFe]-Hydrogenase and Support Hydrogen Production under Fully Aerobic Environment1[OPEN

    Science.gov (United States)

    Liran, Oded; Milrad, Yuval; Eilenberg, Haviva; Weiner, Iddo

    2016-01-01

    Photosynthetic hydrogen production in the microalga Chlamydomonas reinhardtii is catalyzed by two [FeFe]-hydrogenase isoforms, HydA1 and HydA2, both irreversibly inactivated upon a few seconds exposure to atmospheric oxygen. Until recently, it was thought that hydrogenase is not active in air-grown microalgal cells. In contrast, we show that the entire pool of cellular [FeFe]-hydrogenase remains active in air-grown cells due to efficient scavenging of oxygen. Using membrane inlet mass spectrometry, 18O2 isotope, and various inhibitors, we were able to dissect the various oxygen uptake mechanisms. We found that both chlororespiration, catalyzed by plastid terminal oxidase, and Mehler reactions, catalyzed by photosystem I and Flavodiiron proteins, significantly contribute to oxygen uptake rate. This rate is considerably enhanced with increasing light, thus forming local anaerobic niches at the proximity of the stromal face of the thylakoid membrane. Furthermore, we found that in transition to high light, the hydrogen production rate is significantly enhanced for a short duration (100 s), thus indicating that [FeFe]-hydrogenase functions as an immediate sink for surplus electrons in aerobic as well as in anaerobic environments. In summary, we show that an anaerobic locality in the chloroplast preserves [FeFe]-hydrogenase activity and supports continuous hydrogen production in air-grown microalgal cells. PMID:27443604

  7. The chloroplast signal recognition particle (CpSRP) pathway as a tool to minimize chlorophyll antenna size and maximize photosynthetic productivity.

    Science.gov (United States)

    Kirst, Henning; Melis, Anastasios

    2014-01-01

    The concept of the Truncated Light-harvesting chlorophyll Antenna (TLA) size, as a tool by which to maximize sunlight utilization and photosynthetic productivity in microalgal mass cultures or high-density plant canopies, is discussed. TLA technology is known to improve sunlight-to-product energy conversion efficiencies and is hereby exemplified by photosynthetic productivity estimates of wild type and a TLA strain under simulated mass culture conditions. Recent advances in the generation of TLA-type mutants by targeting genes of the chloroplast signal-recognition particle (CpSRP) pathway, affecting the thylakoid membrane assembly of light-harvesting proteins, are also summarized. Two distinct CpSRP assembly pathways are recognized, one entailing post-translational, the other a co-translational mechanism. Differences between the post-translational and co-translational integration mechanisms are outlined, as these pertain to the CpSRP-mediated assembly of thylakoid membrane protein complexes in higher plants and green microalgae. The applicability of the CpSRP pathway genes in efforts to generate TLA-type strains with enhanced solar energy conversion efficiency in photosynthesis is evaluated.

  8. Functional Implications of Photosystem II Crystal Formation in Photosynthetic Membranes*

    Science.gov (United States)

    Tietz, Stefanie; Puthiyaveetil, Sujith; Enlow, Heather M.; Yarbrough, Robert; Wood, Magnus; Semchonok, Dmitry A.; Lowry, Troy; Li, Zhirong; Jahns, Peter; Boekema, Egbert J.; Lenhert, Steven; Niyogi, Krishna K.; Kirchhoff, Helmut

    2015-01-01

    The structural organization of proteins in biological membranes can affect their function. Photosynthetic thylakoid membranes in chloroplasts have the remarkable ability to change their supramolecular organization between disordered and semicrystalline states. Although the change to the semicrystalline state is known to be triggered by abiotic factors, the functional significance of this protein organization has not yet been understood. Taking advantage of an Arabidopsis thaliana fatty acid desaturase mutant (fad5) that constitutively forms semicrystalline arrays, we systematically test the functional implications of protein crystals in photosynthetic membranes. Here, we show that the change into an ordered state facilitates molecular diffusion of photosynthetic components in crowded thylakoid membranes. The increased mobility of small lipophilic molecules like plastoquinone and xanthophylls has implications for diffusion-dependent electron transport and photoprotective energy-dependent quenching. The mobility of the large photosystem II supercomplexes, however, is impaired, leading to retarded repair of damaged proteins. Our results demonstrate that supramolecular changes into more ordered states have differing impacts on photosynthesis that favor either diffusion-dependent electron transport and photoprotection or protein repair processes, thus fine-tuning the photosynthetic energy conversion. PMID:25897076

  9. Functional Implications of Photosystem II Crystal Formation in Photosynthetic Membranes.

    Science.gov (United States)

    Tietz, Stefanie; Puthiyaveetil, Sujith; Enlow, Heather M; Yarbrough, Robert; Wood, Magnus; Semchonok, Dmitry A; Lowry, Troy; Li, Zhirong; Jahns, Peter; Boekema, Egbert J; Lenhert, Steven; Niyogi, Krishna K; Kirchhoff, Helmut

    2015-05-29

    The structural organization of proteins in biological membranes can affect their function. Photosynthetic thylakoid membranes in chloroplasts have the remarkable ability to change their supramolecular organization between disordered and semicrystalline states. Although the change to the semicrystalline state is known to be triggered by abiotic factors, the functional significance of this protein organization has not yet been understood. Taking advantage of an Arabidopsis thaliana fatty acid desaturase mutant (fad5) that constitutively forms semicrystalline arrays, we systematically test the functional implications of protein crystals in photosynthetic membranes. Here, we show that the change into an ordered state facilitates molecular diffusion of photosynthetic components in crowded thylakoid membranes. The increased mobility of small lipophilic molecules like plastoquinone and xanthophylls has implications for diffusion-dependent electron transport and photoprotective energy-dependent quenching. The mobility of the large photosystem II supercomplexes, however, is impaired, leading to retarded repair of damaged proteins. Our results demonstrate that supramolecular changes into more ordered states have differing impacts on photosynthesis that favor either diffusion-dependent electron transport and photoprotection or protein repair processes, thus fine-tuning the photosynthetic energy conversion.

  10. The regulation of TiO2 nanoparticles on the expression of light-harvesting complex II and photosynthesis of chloroplasts of Arabidopsis thaliana.

    Science.gov (United States)

    Ze, Yuguan; Liu, Chao; Wang, Ling; Hong, Mengmeng; Hong, Fashui

    2011-11-01

    Recent studies demonstrated that titanium dioxide nanoparticles (TiO2 NPs) could significantly promote photosynthesis and plant growth, but its mechanism is still unclear. In this article, we studied the mechanism of light absorption and transfer of chloroplasts of Arabidopsis thaliana caused by TiO2 NPs treated. The results showed that TiO2 NPs could induce significant increases of light-harvesting complex II (LHCII) b gene expression and LHCII II content on the thylakoid membrane in A. thaliana, and the increases in LHCII were higher than the non-nano TiO2 (bulk-TiO2) treatment. Meanwhile, spectroscopy assays indicated that TiO2 NPs obviously increased the absorption peak intensity of the chloroplast in red and blue region, the fluorescence quantum yield near 680 nm, the excitation peak intensity near 440 and 480 nm and/or near 650 and 680 nm of the chloroplast. TiO2 NPs treatment could reduce F480/F440 ratio and increase F650/F680 ratio and accelerate the rate of whole chain electron transport and oxygen evolution of the chloroplast. However, the photosynthesis improvement of the non-nanoTiO2 treatment was far less effective than TiO2 NPs treatment. Taken together, TiO2 NPs could promote the light absorption of chloroplast, regulate the distribution of light energy from PS I to PS II by increasing LHCII and accelerate the transformation from light energy to electronic energy, water photolysis, and oxygen evolution.

  11. The chlorophyll a fluorescence induction pattern in chloroplasts upon repetitive single turnover excitations: accumulation and function of QB-nonreducing centers.

    Science.gov (United States)

    Vredenberg, Wim; Kasalicky, Vojtech; Durchan, Milan; Prasil, Ondrej

    2006-03-01

    The increase of chlorophyll fluorescence yield in chloroplasts in a 12.5 Hz train of saturating single turnover flashes and the kinetics of fluorescence yield decay after the last flash have been analyzed. The approximate twofold increase in Fm relative to Fo, reached after 30-40 flashes, is associated with a proportional change in the slow (1-20 s) component of the multiphasic decay. This component reflects the accumulation of a sizeable fraction of QB-nonreducing centers. It is hypothesized that the generation of these centers occurs in association with proton transport across the thylakoid membrane. The data are quantitatively consistent with a model in which the fluorescence quenching of QB-nonreducing centers is reversibly released after second excitation and electron trapping on the acceptor side of Photosystem II.

  12. Biogenesis of photosynthetic complexes in the chloroplast of Chlamydomonas reinhardtii requires ARSA1, a homolog of prokaryotic arsenite transporter and eukaryotic TRC40 for guided entry of tail-anchored proteins.

    Science.gov (United States)

    Formighieri, Cinzia; Cazzaniga, Stefano; Kuras, Richard; Bassi, Roberto

    2013-03-01

    as1, for antenna size mutant 1, was obtained by insertion mutagenesis of the unicellular green alga Chlamydomonas reinhardtii. This strain has a low chlorophyll content, 8% with respect to the wild type, and displays a general reduction in thylakoid polypeptides. The mutant was found to carry an insertion into a homologous gene, prokaryotic arsenite transporter (ARSA), whose yeast and mammal counterparts were found to be involved in the targeting of tail-anchored (TA) proteins to cytosol-exposed membranes, essential for several cellular functions. Here we present the characterization in a photosynthetic organism of an insertion mutant in an ARSA-homolog gene. The ARSA1 protein was found to be localized in the cytosol, and yet its absence in as1 leads to a small chloroplast and a strongly decreased chlorophyll content per cell. ARSA1 appears to be required for optimal biogenesis of photosynthetic complexes because of its involvement in the accumulation of TOC34, an essential component of the outer chloroplast membrane translocon (TOC) complex, which, in turn, catalyzes the import of nucleus-encoded precursor polypeptides into the chloroplast. Remarkably, the effect of the mutation appears to be restricted to biogenesis of chlorophyll-binding polypeptides and is not compensated by the other ARSA homolog encoded by the C. reinhardtii genome, implying a non-redundant function.

  13. Thylakoid redox signals are integrated into organellar-gene-expression-dependent retrograde signalling in the prors1-1 mutant

    Directory of Open Access Journals (Sweden)

    Luca eTadini

    2012-12-01

    Full Text Available Perturbations in organellar gene expression (OGE and the thylakoid redox state (TRS activate retrograde signalling pathways that adaptively modify nuclear gene expression (NGE, according to developmental and metabolic needs. The prors1-1 mutation in Arabidopsis down-regulates the expression of the nuclear gene Prolyl-tRNA Synthetase1 (PRORS1 which acts in both plastids and mitochondria, thereby impairing protein synthesis in both organelles and triggering OGE-dependent retrograde signalling. Because the mutation also affects thylakoid electron transport, TRS-dependent signals may likewise have an impact on the changes in NGE observed in this genotype. In this study, we have investigated whether signals related to TRS are actually integrated into the OGE-dependent retrograde signalling pathway. To this end, the chaos mutation (for chlorophyll a/b binding protein harvesting-organelle specific, which shows a partial loss of PSII antennae proteins and thus a reduction in PSII light absorption capability, was introduced into the prors1-1 mutant background. The resulting double mutant displayed a prors1-1-like reduction in plastid translation rate and a chaos-like decrease in PSII antenna size, whereas the hyper-reduction of the thylakoid electron transport chain, caused by the prors1-1 mutation, was alleviated, as determined by monitoring chlorophyll (Chl fluorescence and thylakoid phosphorylation. Interestingly, a substantial fraction of the nucleus-encoded photosynthesis genes down-regulated in the prors1-1 mutant are expressed at nearly wild-type rates in prors1-1 chaos leaves, and this recovery is reflected in the steady-state levels of their protein products in the chloroplast. We therefore conclude that signals related to photosynthetic electron transport and TRS, and indirectly to carbohydrate metabolism and energy balance, are indeed fed into the OGE-dependent retrograde pathway to modulate NGE and adjust the abundance of chloroplast proteins.

  14. Immunogold localization of acyl carrier protein in plants and Escherichia coli: Evidence for membrane association in plants.

    Science.gov (United States)

    Slabas, A R; Smith, C G

    1988-08-01

    Immunogold labelling was used to study the distribution of acyl carrier protein (ACP) in Escherichia coli and a variety of plant tissues. In E. coli, ACP is distributed throughout the cytoplasm, confirming the observation of S. Jackowski et al. (1985, J. Bacteriol., 162, 5-8_. In the mesocarp of Avocado (Persea americana) and maturing seeds of oil-seed rape (Brassica napus cv. Jet Neuf), over 95% of the ACP is localised to plastids. The protein is almost exclusively located in the chloroplasts of leaf material from oil-seed rape. Approximately 80% of the gold particles associated with the ACP were further localized to the thylakoid membrane of the chloroplast. Since acetyl-CoA carboxylase has been reported to be localized to the thylakoid membrane (C.G. Kannangara and C.J. Jensen, 1975, Eur. J. Biochem., 54, 25-30), these results are consistent with the view that the two sequential enzymes in fatty-acid synthesis are in close spacial proximity.

  15. Effect of dimethyl phthalate (DMP) on germination, antioxidant system, and chloroplast ultrastructure in Cucumis sativus L.

    Science.gov (United States)

    Zhang, Ying; Zhang, Hui; Sun, Xin; Wang, Lei; Du, Na; Tao, Yue; Sun, Guoqiang; Erinle, Kehinde O; Wang, Pengjie; Zhou, Changjian; Duan, Shuwei

    2016-01-01

    Pollution of agricultural soils caused by widely employed plastic products, such as phthalic acid esters (PAEs), are becoming widespread in China, and they have become a threat to human health and the environment. However, little information is available on the influence of PAEs on vegetable crops. In this study, effects of different dimethyl phthalate (DMP) treatments (0, 30, 50, 100, and 200 mg L(-1)) on seed germination and growth of cucumber seedlings were investigated. Although germination rate showed no significant difference compared to control, seed germination time was significantly delayed at DMP greater than 50 mg L(-1). Concentrations of DMP greater than 30 mg L(-1) reduced cucumber lateral root length and number. The measurement of five physiological indexes in cucumber leaves with increasing DMP concentration revealed a decrease in leaf chlorophyll content, while proline and H2O2 contents increased. Peroxidase (POD) and catalase (CAT) activities increased in cucumber plants under 30 and 50 mg L(-1) DMP treatments compared to control; while after a 7-day treatment, these activities were seriously reduced under 100 and 200 mg L(-1) DMP treatments. According to transmission electron microscopy (TEM) micrographic images, the control and 30 mg L(-1) DMP treatments caused no change to leaf chloroplast shape with well-structured thylakoid membrane and parallel pattern of lamellae. At concentrations higher than 30 mg L(-1), DMP altered the ultrastructure of chloroplast, damaged membrane structure, disordered the lamellae, and increased the number and volume of starch grains. Moreover, the envelope of starch grains began to degrade under 200 mg L(-1) DMP treatment.

  16. Effects of 24-epibrassinolide on the photosynthetic characteristics, antioxidant system, and chloroplast ultrastructure in Cucumis sativus L. under Ca(NO(3))(2) stress.

    Science.gov (United States)

    Yuan, Lingyun; Shu, Sheng; Sun, Jin; Guo, Shirong; Tezuka, Takafumi

    2012-09-01

    The effects of 0.1 μM 24-epibrassinolide (EBL) on plant growth (plant height, leaf area, fresh weight, and dry weight), chlorophyll content, photosynthetic characteristics, antioxidant enzymes, and chloroplast ultrastructure were investigated using cucumber seedlings (Cucumis sativus L. cv. Jinyou No. 4) with 80 mM Ca(NO(3))(2) to induce stress. The presence of Ca(NO(3))(2) caused significant reductions in net photosynthetic rate (P(N)), stomatal conductance (Gs), intercellular CO(2) concentration (Ci), and transpiration rate (Tr) of leaves. In addition, Ca(NO(3))(2) markedly reduced the chlorophyll content and inhibited photochemical activity, including the actual photochemical efficiency (ΦPSII). In contrast, EBL increased the chlorophyll content, especially chlorophyll b, and minimized the harmful effects on photosynthesis caused by the Ca(NO(3))(2). The application of EBL to the plants subjected to Ca(NO(3))(2)-enhanced photochemical activity. EBL protected the photosynthetic membrane system from oxidative damage due to up-regulating the capacity of the antioxidant systems. Microscopic analyses revealed that Ca(NO(3))(2) affected the structure of the photosynthetic apparatus and membrane system and induced damage of granal thylakoid layers, while EBL recovered the typical shape of chloroplasts and promoted the formation of grana. Taken together, EBL compensated for damage/losses by Ca(NO(3))(2) due to the regulation of photosynthetic characteristics and the antioxidant system.

  17. Trichloroacetate affects the EPR SignalⅡslow and SignalⅠin the thylakoid of Chlamydomonas reinhardtii

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    One electron paramagnetic resonance (EPR) signal, named SignalⅡslow, originates from the oxidized Tyrosine 160 (YDo) of D2 polypeptide of photosystemⅡ reaction center. After adding high concentration trichloroacetate (TCA) to the Chlamydomonas reinhardtii thylakoid suspension, this signal was abolished in a minute. Treatment of TCA also removes a few of polypeptides, including three extrinsic polypeptides of oxygen-evolving complex, from the thylakoid membrane. Based upon the analysis of the microenvironment around YD with a three-dimensional model, it is indicated that relatively high hydrophobicity of this microenvironment may be the essential prerequisite for TCA to affect YD. It has been observed that TCA treatment also retards the decay of the SignalⅠ, produced by the oxidized reaction center chlorophyll dimer (P700+) of photosys- temⅠ.

  18. Long-day photoperiod induced unhealthy development of chloroplasts in the photoperiod-sensitive genie male-sterile rice

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    By measurement of photochemical activities of chloroplasts and observation on supramolecular archi tecture of thylakoids in chloroplasts, it was found that compared with the effects of short-day photoperiod, long-day pho toperiod could induce normal development of chloroplasts in seedlings of NK58S (photoperiod-sensitive genie male-sterile rice) and NK58 (original line) which do not enter the photoperiod sensitive phase and in seedlings of NK58 just enter the photoperiod-sensitive phase. However, it could induce unhealthy development of chloroplasts in seedlings of NK58S which also just enter the photoperiod sensitive phase. This special effect of long-day photoperiod on the development of chloroplasts in NK58S is probably one of main reasons why long-day photoperiod induces rale-sterility in NK58S and normal fertility in NK58.

  19. Analysis of Protein Import into Chloroplasts Isolated from Stressed Plants.

    Science.gov (United States)

    Ling, Qihua; Jarvis, Paul

    2016-11-01

    Chloroplasts are organelles with many vital roles in plants, which include not only photosynthesis but numerous other metabolic and signaling functions. Furthermore, chloroplasts are critical for plant responses to various abiotic stresses, such as salinity and osmotic stresses. A chloroplast may contain up to ~3,000 different proteins, some of which are encoded by its own genome. However, the majority of chloroplast proteins are encoded in the nucleus and synthesized in the cytosol, and these proteins need to be imported into the chloroplast through translocons at the chloroplast envelope membranes. Recent studies have shown that the chloroplast protein import can be actively regulated by stress. To biochemically investigate such regulation of protein import under stress conditions, we developed the method described here as a quick and straightforward procedure that can easily be achieved in any laboratory. In this method, plants are grown under normal conditions and then exposed to stress conditions in liquid culture. Plant material is collected, and chloroplasts are then released by homogenization. The crude homogenate is separated by density gradient centrifugation, enabling isolation of the intact chloroplasts. Chloroplast yield is assessed by counting, and chloroplast intactness is checked under a microscope. For the protein import assays, purified chloroplasts are incubated with (35)S radiolabeled in vitro translated precursor proteins, and time-course experiments are conducted to enable comparisons of import rates between genotypes under stress conditions. We present data generated using this method which show that the rate of protein import into chloroplasts from a regulatory mutant is specifically altered under osmotic stress conditions.

  20. Nitrogen control of chloroplast differentiation. Annual progress report

    Energy Technology Data Exchange (ETDEWEB)

    Schmidt, G.W.

    1992-07-01

    This project is directed toward understanding how the availability of nitrogen affects the accumulation of chloroplast pigments and proteins functioning in energy transduction and carbon metabolism. Molecular analyses performed with Chlamydomonas reinhardtii grown in a continuous culture system such that ammonium concentration is maintained at a low steady-state concentration so as to limit cell division. As compared to chloroplasts from cells of non-limiting nitrogen provisions, chloroplasts of N-limited cells are profoundly chlorophyll-deficient but still assimilate carbon for deposition of as starch and as storage lipids. Chlorophyll deficiency arises by limiting accumulation of appropriate nuclear-encoded mRNAs of and by depressed rates of translation of chloroplast mRNAs for apoproteins of reaction centers. Chloroplast translational effects can be partially ascribed to diminished rates of chlorophyll biosynthesis in N-limited cells, but pigment levels are not determinants for expression of the nuclear light-harvesting protein genes. Consequently, other signals that are responsive to nitrogen availability mediate transcriptional or post-transcriptional processes for accumulation of the mRNAs for LHC apoproteins and other mRNAs whose abundance is dependent upon high nitrogen levels. Conversely, limited nitrogen availability promotes accumulation of other proteins involved in carbon metabolism and oxidative electron transport in chloroplasts. Hence, thylakoids of N-limited cells exhibit enhanced chlororespiratory activities wherein oxygen serves as the electron acceptor in a pathway that involves plastoquinone and other electron carrier proteins that remain to be thoroughly characterized. Ongoing and future studies are also outlined.

  1. Fe deficiency induced changes in rice (Oryza sativa L.) thylakoids.

    Science.gov (United States)

    Wang, Yuwen; Xu, Chao; Li, Kang; Cai, Xiaojie; Wu, Min; Chen, Guoxiang

    2017-01-01

    Iron deficiency is an important abiotic stress that limits productivity of crops all over the world. We selected a hybrid rice (Oryza sativa L.), LYPJ, which is super high-yield and widely cultured in China, to investigate changes in the components and structure of thylakoid membranes and photosynthetic performance in response to iron deficiency. Our results demonstrated that photosystem I (PSI) is the primary target for iron deficiency, while the changes in photosystem II (PSII) are important for rebuilding a balance in disrupted energy utilization and dissipation caused by differential degradation of photosynthetic components. The result of immunoblot analysis suggested that the core subunit PsaA declined drastically, while PsbA remained relatively stable. Furthermore, several organizational changes of the photosynthetic apparatus were found by BN-PAGE, including a marked decrease in the PSI core complexes, the Cytb 6 /f complex, and the trimeric form of the LHCII antenna, consistent with the observed unstacking grana. The fluorescence induction analysis indicated a descending PSII activity with energy dissipation enhanced markedly. In addition, we proposed that the crippled CO2 assimilation could be compensated by the enhanced of phosphoenolpyruvate carboxylase (PEPC), which is suggested by the decreased ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO) and photosynthetic efficiency.

  2. A Zea mays 39-kDa thylakoid transglutaminase catalyses the modification by polyamines of light-harvesting complex II in a light-dependent way.

    Science.gov (United States)

    Della Mea, M; Di Sandro, A; Dondini, L; Del Duca, S; Vantini, F; Bergamini, C; Bassi, R; Serafini-Fracassini, D

    2004-09-01

    A transglutaminase (TGase; EC 2.3.2.13) activity, which shared many properties with the TGase activity of the Helianthus tuberosus chloroplast, was observed in the Zea mays L. chloroplast and in its fractions. This activity was found to be prevalent in thylakoids; bis-(glutamyl) spermidine and bis-(glutamyl) putrescine were the main polyamine conjugates formed. Light stimulated the endogenous thylakoid activity. Putrescine, spermidine and spermine were conjugated to the isolated light-harvesting complex of photosystem II (LHCII) with different degrees of efficiency, spermine being the polyamine most efficiently conjugated. A TGase with a light-sensitive activity was identified in the photosystem II-enriched fraction. Its partial purification on a sucrose gradient allowed the separation of a 39-kDa band, which was immunorecognised by two anti-TGase antibodies (Ab-3 and rat prostatic gland-TGase). Both a colorimetric and a radiometric assay for TGase activity, the former carried out in the presence of biotinylated cadaverine and the latter in the presence of polyamines labelled with radioactive isotopes and resulting in the isolation of glutamyl-polyamines, further confirmed that the thylakoid enzyme is indeed a calcium-dependent transglutaminase (Thyl-TGase). At variance with guinea pig liver and erythrocyte TGases, which are insensitive to light, the activity of the thylakoid transglutaminase is affected by light. Moreover, this enzyme, when tested with purified LHCII as substrate, catalysed the production of mono- and bis-glutamyl-polyamines in equal amounts, whereas the 'animal' enzymes produced mainly mono-derivatives. Herein, it is discussed whether this light sensitivity is due to the enzyme or the substrate.

  3. Short actin-based mechanism for light-directed chloroplast movement in Arabidopsis

    OpenAIRE

    Kadota, Akeo; Yamada, Noboru; Suetsugu, Noriyuki; Hirose, Mana; Saito, Chieko; Shoda, Keiko; Ichikawa, Satoshi; Kagawa, Takatoshi; Nakano, Akihiko; Wada, Masamitsu

    2009-01-01

    Organelle movement is essential for proper function of living cells. In plants, these movements generally depend on actin filaments, but the underlying mechanism is unknown. Here, in Arabidopsis, we identify associations of short actin filaments along the chloroplast periphery on the plasma membrane side associated with chloroplast photorelocation and anchoring to the plasma membrane. We have termed these chloroplast-actin filaments (cp-actin filaments). Cp-actin filaments emerge from the chl...

  4. Small chloroplast-targeted DnaJ proteins are involved in optimization of photosynthetic reactions in Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Piippo Mirva

    2010-03-01

    Full Text Available Abstract Background DnaJ proteins participate in many metabolic pathways through dynamic interactions with various components of these processes. The role of three small chloroplast-targeted DnaJ proteins, AtJ8 (At1 g80920, AtJ11 (At4 g36040 and AtJ20 (At4 g13830, was investigated here using knock-out mutants of Arabidopsis thaliana. Photochemical efficiency, capacity of CO2 assimilation, stabilization of Photosystem (PS II dimers and supercomplexes under high light illumination, energy distribution between PSI and PSII and phosphorylation of PSII-LHCII proteins, global gene expression profiles and oxidative stress responses of these DnaJ mutants were analyzed. Results Knockout of one of these proteins caused a series of events including a decrease in photosynthetic efficiency, destabilization of PSII complexes and loss of control for balancing the redox reactions in chloroplasts. Data obtained with DNA microarray analysis demonstrated that the lack of one of these DnaJ proteins triggers a global stress response and therefore confers the plants greater tolerance to oxidative stress induced by high light or methyl viologen treatments. Expression of a set of genes encoding enzymes that detoxify reactive oxygen species (ROS as well as a number of stress-related transcription factors behaved in the mutants at growth light similarly to that when wild-type (WT plants were transferred to high light. Also a set of genes related to redox regulation were upregulated in the mutants. On the other hand, although the three DnaJ proteins reside in chloroplasts, the expression of most genes encoding thylakoid membrane proteins was not changed in the mutants. Conclusion It is proposed that the tolerance of the DnaJ protein knockout plants to oxidative stress occurs at the expense of the flexibility of photosynthetic reactions. Despite the fact that the effects of the individual protein knockout on the response of plants to high light treatment are quite similar

  5. Auxin and chloroplast movements.

    Science.gov (United States)

    Eckstein, Aleksandra; Krzeszowiec, Weronika; Waligórski, Piotr; Gabryś, Halina

    2016-03-01

    Auxin is involved in a wide spectrum of physiological processes in plants, including responses controlled by the blue light photoreceptors phototropins: phototropic bending and stomatal movement. However, the role of auxin in phototropin-mediated chloroplast movements has never been studied. To address this question we searched for potential interactions between auxin and the chloroplast movement signaling pathway using different experimental approaches and two model plants, Arabidopsis thaliana and Nicotiana tabacum. We observed that the disturbance of auxin homeostasis by shoot decapitation caused a decrease in chloroplast movement parameters, which could be rescued by exogenous auxin application. In several cases, the impairment of polar auxin transport, by chemical inhibitors or in auxin carrier mutants, had a similar negative effect on chloroplast movements. This inhibition was not correlated with changes in auxin levels. Chloroplast relocations were also affected by the antiauxin p-chlorophenoxyisobutyric acid and mutations in genes encoding some of the elements of the SCF(TIR1)-Aux/IAA auxin receptor complex. The observed changes in chloroplast movement parameters are not prominent, which points to a modulatory role of auxin in this process. Taken together, the obtained results suggest that auxin acts indirectly to regulate chloroplast movements, presumably by regulating gene expression via the SCF(TIR1)-Aux/IAA-ARF pathway. Auxin does not seem to be involved in controlling the expression of phototropins.

  6. Age-dependent variation in membrane lipid synthesis in leaves of garden pea (Pisum sativum L.)

    DEFF Research Database (Denmark)

    Hellgren, Lars; Sandelius, A.S.

    2001-01-01

    leaf of older plants, acetate was predominantly allocated into phosphatidylglycerol (PG), which remained the major radiolabelled lipid during the 3 d studied. The proportion of radioactivity recovered in MGDG decreased with increasing plant age up to 20 d, suggesting that, in expanded leaves, MGDG...... is more stable and requires renewal to a lower extent than PG. When the second oldest leaf approached senescence, labelling of MGDG again increased, indicating an increased need for thylakoid repair. The proportion of acetate allocated into phosphatidylethanolamine and free sterols was largest in leaves...... of 18-26-d-old plants and in the youngest leaves, respectively. Thus, these results demonstrate that the distribution of newly synthesized fatty acids between acyl lipid synthesis in the chloroplast and extraplastidial membranes strongly varies with leaf age, as do the proportion utilized for sterol...

  7. Artificially acquired chlorophyll b is highly acceptable to the thylakoid-lacking cyanobacterium, Gloeobacter violaceus PCC 7421.

    Science.gov (United States)

    Araki, Mie; Akimoto, Seiji; Mimuro, Mamoru; Tsuchiya, Tohru

    2014-08-01

    Unicellular cyanobacterium Gloeobacter violaceus is an only known oxygenic photosynthetic organism that lacks thylakoid membrane. Molecular phylogenetic analyses indicate that G. violaceus is an early-branching cyanobacterium within cyanobacterial clade. Therefore, the photosynthetic system of G. violaceus is considered to be partly similar to that of the ancestral cyanobacteria that would lack thylakoid membrane. G. violaceus possesses chlorophyll (Chl) a as the only chlorophyll species like most cyanobacteria. It was proposed that the ancestral oxygenic photosynthetic organism had not only Chl a and phycobilins but also Chl b. However, no organism which contains both Chl a and Chl b and lacks thylakoid membrane has been found in nature. Therefore, we introduced the chlorophyllide a oxygenase gene responsible for Chl b biosynthesis into G. violaceus. In the resultant transformant, Chl b accumulated at approximately 11% of total Chl independent of growth phase. Photosystem I complexes isolated from the transformant contained Chl b at 9.9% of total Chl. The presence of Chl b in the photosystem I complexes did not inhibit trimer formation. Furthermore, time-resolved fluorescence spectrum demonstrated that Chl b transferred energy to Chl a in the photosystem I complexes and did not disturb the energy transfer among the Chl a molecules. These results show that G. violaceus is tolerant to artificially produced Chl b and suggest the flexibility of photosystem for Chl composition in the ancestral oxygenic photosynthetic organism.

  8. Phaseolin expression in tobacco chloroplast reveals an autoregulatory mechanism in heterologous protein translation.

    Science.gov (United States)

    De Marchis, Francesca; Bellucci, Michele; Pompa, Andrea

    2016-02-01

    Plastid DNA engineering is a well-established research area of plant biotechnology, and plastid transgenes often give high expression levels. However, it is still almost impossible to predict the accumulation rate of heterologous protein in transplastomic plants, and there are many cases of unsuccessful transgene expression. Chloroplasts regulate their proteome at the post-transcriptional level, mainly through translation control. One of the mechanisms to modulate the translation has been described in plant chloroplasts for the chloroplast-encoded subunits of multiprotein complexes, and the autoregulation of the translation initiation of these subunits depends on the availability of their assembly partners [control by epistasy of synthesis (CES)]. In Chlamydomonas reinhardtii, autoregulation of endogenous proteins recruited in the assembly of functional complexes has also been reported. In this study, we revealed a self-regulation mechanism triggered by the accumulation of a soluble recombinant protein, phaseolin, in the stroma of chloroplast-transformed tobacco plants. Immunoblotting experiments showed that phaseolin could avoid this self-regulation mechanism when targeted to the thylakoids in transplastomic plants. To inhibit the thylakoid-targeted phaseolin translation as well, this protein was expressed in the presence of a nuclear version of the phaseolin gene with a transit peptide. Pulse-chase and polysome analysis revealed that phaseolin mRNA translation on plastid ribosomes was repressed due to the accumulation in the stroma of the same soluble polypeptide imported from the cytosol. We suggest that translation autoregulation in chloroplast is not limited to heteromeric protein subunits but also involves at least some of the foreign soluble recombinant proteins, leading to the inhibition of plastome-encoded transgene expression in chloroplast.

  9. Overexpression of thylakoidal ascorbate peroxidase shows enhanced resistance to chilling stress in tomato.

    Science.gov (United States)

    Duan, Ming; Feng, Hai-Long; Wang, Li-Yan; Li, Dong; Meng, Qing-Wei

    2012-06-15

    Photosynthesis provides a strong reducing power and a high risk for generation of reactive oxygen species (ROS) particularly under chilling stress. Ascorbate peroxidases (APXs) reduce H(2)O(2) to water and play an important role in the antioxidant system of plants. Though thylakoid ascorbate peroxidase (tAPX) has been thought to be key regulator of intracellular levels of H(2)O(2), its physiological significance in the response to chilling stress is still under discussion. To study the contribution of tAPX to the ROS scavenging, a tomato thylakoidal ascorbate peroxidase gene (LetAPX) was isolated and transgenic tomatoes were obtained. The LetAPX-GFP fusion protein was targeted to chloroplast in Arabidopsis mesophyll protoplast. RNA blotting analysis revealed that the LetAPX transcript expression was up-regulated by chilling, high light, exogenous salicylic acid (SA) and methyl viologen (MV). Over expression of LetAPX in tomatoes conferred tolerance to chilling stress by maintaining higher reduced glutathione (GSH) content, chlorophyll and APX activities compared with wild type (WT) plants. Furthermore, transgenic plants showed lower levels of hydrogen peroxide (H(2)O(2)) and ion leakage, lower malendialdehyde (MDA) content, higher net photosynthetic rate (Pn) and higher maximal photochemical efficiency of PSII (Fv/Fm). The oxidizable P700 decreased more obviously in WT than that in transgenic plants under chilling stress in low irradiance. The results suggested that over expression of tAPX played a key role both in alleviating photo inhibition of PSI and PSII and enhancing their tolerance to chilling stress.

  10. Why have chloroplasts developed a unique motility system?

    Science.gov (United States)

    Suetsugu, Noriyuki; Dolja, Valerian V; Wada, Masamitsu

    2010-10-01

    Organelle movement in plants is dependent on actin filaments with most of the organelles being transported along the actin cables by class XI myosins. Although chloroplast movement is also actin filament-dependent, a potential role of myosin motors in this process is poorly understood. Interestingly, chloroplasts can move in any direction, and change the direction within short time periods, suggesting that chloroplasts use the newly formed actin filaments rather than preexisting actin cables. Furthermore, the data on myosin gene knockouts and knockdowns in Arabidopsis and tobacco do not support myosins' XI role in chloroplast movement. Our recent studies revealed that chloroplast movement and positioning are mediated by the short actin filaments localized at chloroplast periphery (cp-actin filaments) rather than cytoplasmic actin cables. The accumulation of cp-actin filaments depends on kinesin-like proteins, KAC1 and KAC2, as well as on a chloroplast outer membrane protein CHUP1. We propose that plants evolved a myosin XI-independent mechanism of the actin-based chloroplast movement that is distinct from the mechanism used by other organelles.

  11. Investigating cytoskeletal function in chloroplast protrusion formation in the arctic-alpine plant Oxyria digyna.

    Science.gov (United States)

    Holzinger, A; Wasteneys, G O; Lütz, C

    2007-05-01

    Arctic and alpine plants like Oxyria digyna have to face enhanced environmental stress. This study compared leaves from Oxyria digyna collected in the Arctic at Svalbard (78 degrees N) and in the Austrian Alps (47 degrees N) at cellular, subcellular, and ultrastructural levels. Oxyria digyna plants collected in Svalbard had significantly thicker leaves than the samples collected in the Austrian Alps. This difference was generated by increased thickness of the palisade and spongy mesophyll layers in the arctic plants, while epidermal cells had no significant size differences between the two habitats. A characteristic feature of arctic, alpine, and cultivated samples was the occurrence of broad stroma-filled chloroplast protrusions, 2 - 5 microm broad and up to 5 microm long. Chloroplast protrusions were in close spatial contact with other organelles including mitochondria and microbodies. Mitochondria were also present in invaginations of the chloroplasts. A dense network of cortical microtubules found in the mesophyll cells suggested a potential role for microtubules in the formation and function of chloroplast protrusions. No direct interactions between microtubules and chloroplasts, however, were observed and disruption of the microtubule arrays with the anti-microtubule agent oryzalin at 5 - 10 microM did not alter the appearance or dynamics of chloroplast protrusions. These observations suggest that, in contrast to studies on stromule formation in Nicotiana, microtubules are not involved in the formation and morphology of chloroplast protrusions in Oxyria digyna. The actin microfilament-disrupting drug latrunculin B (5 - 10 microM for 2 h) arrested cytoplasmic streaming and altered the cytoplasmic integrity of mesophyll cells. However, at the ultrastructural level, stroma-containing, thylakoid-free areas were still visible, mostly at the concave sides of the chloroplasts. As chloroplast protrusions were frequently found to be mitochondria-associated in Oxyria

  12. Two kinesin-like proteins mediate actin-based chloroplast movement in Arabidopsis thaliana.

    Science.gov (United States)

    Suetsugu, Noriyuki; Yamada, Noboru; Kagawa, Takatoshi; Yonekura, Hisashi; Uyeda, Taro Q P; Kadota, Akeo; Wada, Masamitsu

    2010-05-11

    Organelle movement is essential for efficient cellular function in eukaryotes. Chloroplast photorelocation movement is important for plant survival as well as for efficient photosynthesis. Chloroplast movement generally is actin dependent and mediated by blue light receptor phototropins. In Arabidopsis thaliana, phototropins mediate chloroplast movement by regulating short actin filaments on chloroplasts (cp-actin filaments), and the chloroplast outer envelope protein CHUP1 is necessary for cp-actin filament accumulation. However, other factors involved in cp-actin filament regulation during chloroplast movement remain to be determined. Here, we report that two kinesin-like proteins, KAC1 and KAC2, are essential for chloroplasts to move and anchor to the plasma membrane. A kac1 mutant showed severely impaired chloroplast accumulation and slow avoidance movement. A kac1kac2 double mutant completely lacked chloroplast photorelocation movement and showed detachment of chloroplasts from the plasma membrane. KAC motor domains are similar to those of the kinesin-14 subfamily (such as Ncd and Kar3) but do not have detectable microtubule-binding activity. The C-terminal domain of KAC1 could interact with F-actin in vitro. Instead of regulating microtubules, KAC proteins mediate chloroplast movement via cp-actin filaments. We conclude that plants have evolved a unique mechanism to regulate actin-based organelle movement using kinesin-like proteins.

  13. Iron stabilizes thylakoid protein-pigment complexes in Indian mustard during Cd-phytoremediation as revealed by BN-SDS-PAGE and ESI-MS/MS.

    Science.gov (United States)

    Qureshi, M Irfan; D'Amici, Gian Maria; Fagioni, Marco; Rinalducci, Sara; Zolla, Lello

    2010-07-01

    Two-dimensional BN-SDS-PAGE, ESI-MS/MS and electron microscopy (EM) were used to study the role of iron (Fe) under cadmium (Cd) stress in retention of thylakoidal multiprotein complexes (MPCs) and chloroplast ultrastructure of Indian mustard, a moderate hyperaccumulator plant. Mustard was grown hydroponically with or without iron for 17 days and then exposed to CdCl2 for 3 days. Fe deficiency led to an increase in oxidative stress and damage to chloroplast/thylakoids accompanied by a decrease in chlorophyll content; exposure of plants to Cd further enhanced the oxidative stress and Cd accumulation (more in -Fe plants). However, the presence of iron aided plants in the suppression of oxidative stress and retention of chloroplasts and chlorophylls under Cd stress. Proteomic analyses by 2D BN-SDS-PAGE and mass spectrometry showed that Fe deficiency considerably decreased the amount of LHCII trimer, ATPase-F1 portion, cyt b6/f and RuBisCO. No or less reduction, was observed for PSI(RCI+LHCI), the PSII-core monomer, and the PSII subcomplex, while an increase in the LHCII monomer was noted. Under iron deficiency, Cd proved to be very deleterious to MPCs, except for the PSII subcomplex, the LHCII monomer and free proteins which were increased. Iron proved to be very protective in retaining almost all the complexes. MPCs showed greater susceptibility to Cd than Fe deficiency, mainly at the level of RuBisCO and cyt b6/f; an increase in the amount of the PSII subcomplex, LHCII monomer and free proteins indicates differences in the mechanisms affected by Fe deficiency and Cd stress when compared to Fe-fed plants. This study furthers our understanding of the sites actually damaged in MPCs under Fe deficiency and Cd stress. A role emerges for iron in the protection of MPCs and, hence, of the chloroplast. The present study also indicates the importance of iron for efficient phytoextraction/phytoremediation.

  14. Interaction of actin and the chloroplast protein import apparatus.

    Science.gov (United States)

    Jouhet, Juliette; Gray, John C

    2009-07-10

    Actin filaments are major components of the cytoskeleton and play numerous essential roles, including chloroplast positioning and plastid stromule movement, in plant cells. Actin is present in pea chloroplast envelope membrane preparations and is localized at the surface of the chloroplasts, as shown by agglutination of intact isolated chloroplasts by antibodies to actin. To identify chloroplast envelope proteins involved in actin binding, we have carried out actin co-immunoprecipitation and co-sedimentation experiments on detergent-solubilized pea chloroplast envelope membranes. Proteins co-immunoprecipitated with actin were identified by mass spectrometry and by Western blotting and included the Toc159, Toc75, Toc34, and Tic110 components of the TOC-TIC protein import apparatus. A direct interaction of actin with Escherichia coli-expressed Toc159, but not Toc33, was shown by co-sedimentation experiments, suggesting that Toc159 is the component of the TOC complex that interacts with actin on the cytosolic side of the outer envelope membrane. The physiological significance of this interaction is unknown, but it may play a role in the import of nuclear-encoded photosynthesis proteins.

  15. Carotenoid-chlorophyll coupling and fluorescence quenching correlate with protein packing density in grana-thylakoids.

    Science.gov (United States)

    Holleboom, Christoph-Peter; Yoo, Sunny; Liao, Pen-Nan; Compton, Ian; Haase, Winfried; Kirchhoff, Helmut; Walla, Peter Jomo

    2013-09-26

    The regulation of light-harvesting in photosynthesis under conditions of varying solar light irradiation is essential for the survival and fitness of plants and algae. It has been proposed that rearrangements of protein distribution in the stacked grana region of thylakoid membranes connected to changes in the electronic pigment-interaction play a key role for this regulation. In particular, carotenoid-chlorophyll interactions seem to be crucial for the down-regulation of photosynthetic light-harvesting. So far, it has been difficult to determine the influence of the dense protein packing found in native photosynthetic membrane on these interactions. We investigated the changes of the electronic couplings between carotenoids and chlorophylls and the quenching in grana thylakoids of varying protein packing density by two-photon spectroscopy, conventional chlorophyll fluorometry, low-temperature fluorescence spectroscopy, and electron micrographs of freeze-fracture membranes. We observed an increasing carotenoid-chlorophyll coupling and fluorescence quenching with increasing packing density. Simultaneously, the antennas size and excitonic connectivity of Photosystem II increased with increasing quenching and carotenoid-chlorophyll coupling whereas isolated, decoupled LHCII trimers decreased. Two distinct quenching data regimes could be identified that show up at different protein packing densities. In the regime corresponding to higher protein packing densities, quenching is strongly correlated to carotenoid-chlorophyll interactions whereas in the second regime, a weak correlation is apparent with low protein packing densities. Native membranes are in the strong-coupling data regime. Consequently, PSII and LHCII in grana membranes of plants are already quenched by protein crowding. We concluded that this ensures efficient electronic connection of all pigment-protein complexes for intermolecular energy transfer to the reaction centers and allows simultaneously

  16. Reference: 757 [Arabidopsis Phenome Database[Archive

    Lifescience Database Archive (English)

    Full Text Available of prothylakoids and transition into mature thylakoid stacks. The AtTerC gene encodes an integral membrane ...nt chloroplasts lacked thylakoids, did not have grana stacks, and showed numerous globular structures of var

  17. Effects of Glycerol on the Fluorescence Spectra and Chloroplast Ultrastructure of Phaeodactylum tricornutum (Bacillariophyta)

    Institute of Scientific and Technical Information of China (English)

    Xiao-Juan Liu; Shun-Shan Duan; Ai-Fen Li; Kai-Feng Sun

    2009-01-01

    Responses of the photosynthetic activity of Phaeodactylum tricornutum (Bacillariophyta) to organic carbon glycerol were investigated. The growth rate, photosynthetic pigments, 77 K fluorescence spectra, and chloroplast ultrastructure of P. tricornutum were examined under photoautotrophic, mixotrophic, and photoheterotrophic conditions. The results showed that the specific growth rate was the fastest under mixotrophic conditions. The cell photosynthetic pigment content and values of Chl a/Chl c were reduced under mixotrophic and photoheterotrophic conditions. The value of carotenoid/Chl a was enhanced under mixotrophic conditions, but was decreased under photoheterotrophic conditions. In comparison with photoautotrophic conditions, the fluorescence emission peaks and fluorescence excitation peaks were not shifted. The relative fluorescence of photosystem (PS) Ⅰ and PS Ⅱ and the values of F6851F710 and F685/F738 were decreased. Chloroplast thylakoid pairs were less packed under mixotrophic and photoheterotrophic conditions. There was a strong correlation between degree of chloroplast thylakoid packing and the excitation energy kept in PS Ⅱ. These results suggested that the PS Ⅱ activity was reduced by glycerol under mixotrophic conditions, thereby leading to repression of the photosynthetic activity.

  18. Nitrogen control of chloroplast differentiation. Final report

    Energy Technology Data Exchange (ETDEWEB)

    Schmidt, G.W.

    1998-05-01

    This project was directed toward understanding at the physiological, biochemical and molecular levels of how photosynthetic organisms adapt to long-term nitrogen-deficiency conditions is quite incomplete even though limitation of this nutrient is the most commonly restricts plant growth and development. For our work on this problem, the unicellular green alga, Chlamydomonas reinhardtii, was grown in continuous cultures in which steady-state levels of nitrogen can be precisely controlled. N-limited cells exhibit the classical symptoms of deficiency of this nutrient, chlorosis and slow growth rates, and respond to nitrogen provision by rapid greening and chloroplast differentiation. We have addressed three aspects of this problem: (1) the regulation of pigment synthesis; (2) control of expression of nuclear genes encoding photosynthetic proteins; (3) changes in metabolic and electron transport pathways that enable sustained CO{sub 2} fixation even though they cannot be readily converted into amino and nucleic acids. For the last, principle components are: (a) enhanced mitochondrial respiratory activity intimately associated with photosynthates, and (b) the occurrence in thylakoids of a supplemental electron transport pathway that facilitates reduction of the plastoquinone pool. Together, these distinguishing features of N-limited cells are likely to enable cell survival, especially under conditions of high irradiance stress.

  19. Chloroplast Redox Poise

    DEFF Research Database (Denmark)

    Steccanella, Verdiana

    the redox status of the plastoquinone pool and chlorophyll biosynthesis. Furthermore, in the plant cell, the equilibrium between redox reactions and ROS signals is also maintained by various balancing mechanisms among which the thioredoxin reductase-thioredoxin system (TR-Trx) stands out as a mediator......The redox state of the chloroplast is maintained by a delicate balance between energy production and consumption and is affected by the need to avoid increased production of reactive oxygen species (ROS). Redox power and ROS generated in the chloroplast are essential for maintaining physiological...... metabolic pathways and for optimizing chloroplast functions. The redox poise of photosynthetic electron transport components like plastoquinone is crucial to initiate signaling cascades and might also be involved in key biosynthetic pathways such as chlorophyll biosynthesis. We, therefore, explored...

  20. Automatic Chloroplast Movement Analysis.

    Science.gov (United States)

    Johansson, Henrik; Zeidler, Mathias

    2016-01-01

    In response to low or high intensities of light, the chloroplasts in the mesophyll cells of the leaf are able to increase or decrease their exposure to light by accumulating at the upper and lower sides or along the side walls of the cell respectively. This movement, regulated by the phototropin blue light photoreceptors phot1 and phot2, results in a decreased or increased transmission of light through the leaf. This way the plant is able to optimize harvesting of the incoming light or avoid damage caused by excess light. Here we describe a method that indirectly measures the movement of chloroplasts by taking advantage of the resulting change in leaf transmittance. By using a microplate reader, quantitative measurements of chloroplast accumulation or avoidance can be monitored over time, for multiple samples with relatively little hands-on time.

  1. Functional analysis of chloroplast early light inducible proteins (ELIPs)

    Energy Technology Data Exchange (ETDEWEB)

    Wetzel, Carolyn M

    2005-02-22

    The objectives of this project were to characterize gene expression patterns of early light inducible protein (ELIP) genes in Arabidopsis thaliana and in Lycopersicon esculentum, to identify knock mutants of the 2 ELIP genes in Arabidopsis, and to characterize the effects of the knockouts. Expression in Arabidopsis was studied in response to thylakoid electron transport chain (PETC) capacity, where it was found that there is a signal for expression associated with reduction of the PETC. Expression in response to salt was also studied, with different responses of the two gene copies. Knockout lines for ELIP1 and ELIP2 have been identified and are being characterized. In tomato, it was found that the single-copy ELIP gene is highly expressed in ripening fruit during the chloroplast-to-chromoplast transition. Studies of expression in tomato ripening mutants are ongoing.

  2. Moderate heat stress of Arabidopsis thaliana leaves causes chloroplast swelling and plastoglobule formation.

    Science.gov (United States)

    Zhang, Ru; Wise, Robert R; Struck, Kimberly R; Sharkey, Thomas D

    2010-08-01

    Photosynthesis is inhibited by heat stress. This inhibition is rapidly reversible when heat stress is moderate but irreversible at higher temperature. Absorbance changes can be used to detect a variety of biophysical parameters in intact leaves. We found that moderate heat stress caused a large reduction of the apparent absorbance of green light in light-adapted, intact Arabidopsis thaliana leaves. Three mechanisms that can affect green light absorbance of leaves, namely, zeaxanthin accumulation (absorbance peak at 505 nm), the electrochromic shift (ECS) of carotenoid absorption spectra (peak at 518 nm), and light scattering (peak at 535 nm) were investigated. The change of green light absorbance caused by heat treatment was not caused by changes of zeaxanthin content nor by the ECS. The formation of non-photochemical quenching (NPQ), chloroplast movements, and chloroplast swelling and shrinkage can all affect light scattering inside leaves. The formation of NPQ under high temperature was not well correlated with the heat-induced absorbance change, and light microscopy revealed no appreciable changes of chloroplast location because of heat treatment. Transmission electron microscopy results showed swollen chloroplasts and increased number of plastoglobules in heat-treated leaves, indicating that the structural changes of chloroplasts and thylakoids are significant results of moderate heat stress and may explain the reduced apparent absorbance of green light under moderately high temperature.

  3. Regulation of chloroplast biogenesis: the immutans mutant of Arabidopsis

    Energy Technology Data Exchange (ETDEWEB)

    Rodermel, Steven

    2015-11-16

    The immutans (im) variegation mutant of Arabidopsis is an ideal model to gain insight into factors that control chloroplast biogenesis. im defines the gene for PTOX, a plastoquinol terminal oxidase that participates in control of thylakoid redox. Here, we report that the im defect can be suppressed during the late stages of plant development by gigantea (gi2), which defines the gene for GIGANTEA (GI), a central component of the circadian clock that plays a poorly-understood role in diverse plant developmental processes. imgi2 mutants are late-flowering and display other well-known phenotypes associated with gi2, such as starch accumulation and resistance to oxidative stress. We show that the restoration of chloroplast biogenesis in imgi2 is caused by a developmental-specific de-repression of cytokinin signaling that involves crosstalk with signaling pathways mediated by gibberellin (GA) and SPINDLY (SPY), a GA response inhibitor. Suppression of the plastid defect in imgi2 is likely caused by a relaxation of excitation pressures in developing plastids by factors contributed by gi2, including enhanced rates of photosynthesis and increased resistance to oxidative stress. Interestingly, the suppression phenotype of imgi can be mimicked by crossing im with the starch accumulation mutant, sex1, perhaps because sex1 utilizes pathways similar to gi. We conclude that our studies provide a direct genetic linkage between GIGANTEA and chloroplast biogenesis, and we construct a model of interactions between signaling pathways mediated by gi, GA, SPY, cytokinins, and sex1 that are required for chloroplast biogenesis.

  4. Phylogenetic analysis of the thylakoid ATP/ADP carrier reveals new insights into its function restricted to green plants

    Directory of Open Access Journals (Sweden)

    Cornelia eSpetea

    2012-01-01

    Full Text Available ATP is the common energy currency of cellular metabolism in all living organisms. Most of them synthesize ATP in the cytosol or on the mitochondrial inner membrane, whereas land plants, algae and cyanobacteria also produce it on the thylakoid membrane during the light-dependent reactions of photosynthesis. From the site of synthesis, ATP is transported to the site of utilization via intracellular membranes transporters. One major type of ATP transporter is represented by the mitochondrial ADP/ATP carrier family. Here we review a recently characterized member, namely the thylakoid ATP/ADP carrier from Arabidopsis thaliana (AtTAAC. Thus far, no orthologues of this carrier have been characterized in other organisms, although similar sequences can be recognized in many sequenced genomes. Protein Sequence database searches and phylogenetic analyses indicate the absence of TAAC in cyanobacteria and its appearance early in the evolution of photosynthetic eukaryotes. The TAAC clade is composed of carriers found in land plants and some green algae, but no proteins from other photosynthetic taxa, such as red algae, brown algae and diatoms. This implies that TAAC-like sequences arose only once before the divergence of green algae and land plants. Based on these findings, it is proposed that TAAC may have evolved in response to the need of a new activity in higher photosynthetic eukaryotes. This activity may provide the energy to drive reactions during biogenesis and turnover of photosynthetic complexes, which are heterogenously distributed in a thylakoid membrane system composed of appressed and non-appressed regions.

  5. Albino Leaf1 That Encodes the Sole Octotricopeptide Repeat Protein Is Responsible for Chloroplast Development1[OPEN

    Science.gov (United States)

    Tan, Jianjie; Xing, Yi; Liu, Changhong; Chen, Qiaoling; Zhu, Haitao; Wang, Jiang; Zhang, Jingliu; Zhang, Guiquan

    2016-01-01

    Chloroplast, the photosynthetic organelle in plants, plays a crucial role in plant development and growth through manipulating the capacity of photosynthesis. However, the regulatory mechanism of chloroplast development still remains elusive. Here, we characterized a mutant with defective chloroplasts in rice (Oryza sativa), termed albino leaf1 (al1), which exhibits a distinct albino phenotype in leaves, eventually leading to al1 seedling lethality. Electronic microscopy observation demonstrated that the number of thylakoids was reduced and the structure of thylakoids was disrupted in the al1 mutant during rice development, which eventually led to the breakdown of chloroplast. Molecular cloning revealed that AL1 encodes the sole octotricopeptide repeat protein (RAP) in rice. Genetic complementation of Arabidopsis (Arabidopsis thaliana) rap mutants indicated that the AL1 protein is a functional RAP. Further analysis illustrated that three transcript variants were present in the AL1 gene, and the altered splices occurred at the 3′ untranslated region of the AL1 transcript. In addition, our results also indicate that disruption of the AL1 gene results in an altered expression of chloroplast-associated genes. Consistently, proteomic analysis demonstrated that the abundance of photosynthesis-associated proteins is altered significantly, as is that of a group of metabolism-associated proteins. More specifically, we found that the loss of AL1 resulted in altered abundances of ribosomal proteins, suggesting that RAP likely also regulates the homeostasis of ribosomal proteins in rice in addition to the ribosomal RNA. Taken together, we propose that AL1, particularly the AL1a and AL1c isoforms, plays an essential role in chloroplast development in rice. PMID:27208287

  6. Effect of Salts and Electron Transport on the Conformation of Isolated Chloroplasts. II. Electron Microscopy 1

    Science.gov (United States)

    Izawa, Seikichi; Good, Norman E.

    1966-01-01

    Spinach chloroplasts isolated in media containing salts and the rare chloroplasts which are still within their envelopes alike retain grana similar to those seen in chloroplasts in situ. Chloroplasts isolated in low-salt media lose their grana without losing any chlorophyll. These grana-free chloroplasts are considerably swollen and consist almost entirely of continuous sheets of paired-membrane structures. These double structures, the lamellae, are only loosely held together, primarily at the edges, by tenuous material which does not react with permanganate. Addition of salts (methylamine hydrochloride, NaCl, MgCl2) to the grana-free low-salt chloroplasts provide strong interlamellar attractions. These attractions result in a stacking of the lamellae which is sometimes almost random but sometimes results in regular structures indistinguishable from the original grana. The phosphorylation-uncoupler atebrin causes further swelling of the chloroplasts in the absence of electron transport by increasing the space between the paired membranes of the lamellae. The rapid electron transport (Hill reaction) made possible by atebrin-uncoupling is associated with a great decrease in chloroplast volume. This decrease results from a collapsing together of the widely separated lamellar membrane pairs. The pairs approach each other so closely that they usually appear as a single membrane when viewed with the electron microscope. The much slower electron transport which occurs in the absence of uncouplers is associated with a similar but smaller decrease in the space between the lamellar membrane pairs. Chloroplasts swell during the rapid electron transport made possible by the phosphorylation-uncoupler methylamine. This swelling is accompanied by a degree of membrane distortion which precludes an interpretation of the mechanism. As with atebrin-faciliated electron transport, obviously paired membranes disappear but it is not yet clear whether this is by association or

  7. Below-ambient levels of UV induce chloroplast structural change and alter starch metabolism.

    Science.gov (United States)

    Fagerberg, W R

    2007-01-01

    Electromagnetic radiation (EMR) in the 400-700 nm bandwidth of photosynthetically active radiation (PAR) has been established as an important source of energy for photosynthesis and environmental signals regulating many aspects of green-plant life. Above-ambient levels of UV-B radiation (290-320 nm) under high-PAR conditions have been shown to elicit responses in chloroplasts of Brassica napus similar to those of chloroplasts at low-PAR exposure (W. Fagerberg and J. Bornman, Physiol. Plant. 101: 833-844, 1997). The question arises as to whether UV at normal levels can also evoke similar responses. Here we provide evidence that even below-ambient levels of UV-B (1/28 ambient; Durham, N.H., U.S.A., 1200 hours, March) were capable of inducing an increase in thylakoid surface area relative to the chloroplast volume typical of a low-PAR response (shade response) in sunflowers. This response occurred even though leaves were concurrently exposed to PAR levels that normally induce a "sun" or high-PAR response in the absence of UV-B. Subambient levels of UV-B were also associated with a decrease in chloroplast and starch volume. Exposure to levels of UV-A 1/10 of ambient appeared to enhance the high-PAR response of the chloroplast, characterized by an increase in the amounts of stored starch, an increase in chloroplast volume density ratio values, and a decrease in thylakoid surface area density ratios relative to the high-light controls. These effects were opposite to those seen in UV-B-exposed tissue. In a general sense, subambient levels of UV-B evoked a response similar to that elicited by low-PAR irradiance, while subambient UV-A elicited responses similar to those typical of high-PAR irradiance. The fact that below-ambient levels of UV altered a normal chloroplast structural response to PAR provides evidence that UV may be an important environmental signal for plants.

  8. Silicon alleviates cadmium toxicity by enhanced photosynthetic rate and modified bundle sheath's cell chloroplasts ultrastructure in maize.

    Science.gov (United States)

    Vaculík, Marek; Pavlovič, Andrej; Lux, Alexander

    2015-10-01

    Silicon was shown to alleviate the negative effects of various biotic and abiotic stresses on plant growth. Although the positive role of Si on toxic and heavy metal Cd has been already described, the mechanisms have been explained only partially and still remain unclear. In the present study we investigated the effect of Si on photosynthetic-related processes in maize exposed to two different levels of Cd via measurements of net photosynthetic rate (AN), chlorophyll a fluorescence and pigment analysis, as well as studies of leaf tissue anatomy and cell ultrastructure using bright-field and transmission electron microscopy. We found that Si actively alleviated the toxic syndromes of Cd by increasing AN, effective photochemical quantum yield of photosystem II (ϕPSII) and content of assimilation pigments, although did not decrease the concentration of Cd in leaf tissues. Cadmium did not affect the leaf anatomy and ultrastructure of leaf mesophyll's cell chloroplasts; however, Cd negatively affected thylakoid formation in chloroplasts of bundle sheath cells, and this was alleviated by Si. Improved thylakoid formation in bundle sheath's cell chloroplasts may contribute to Si-induced enhancement of photosynthesis and related increase in biomass production in C4 plant maize.

  9. Short actin-based mechanism for light-directed chloroplast movement in Arabidopsis.

    Science.gov (United States)

    Kadota, Akeo; Yamada, Noboru; Suetsugu, Noriyuki; Hirose, Mana; Saito, Chieko; Shoda, Keiko; Ichikawa, Satoshi; Kagawa, Takatoshi; Nakano, Akihiko; Wada, Masamitsu

    2009-08-04

    Organelle movement is essential for proper function of living cells. In plants, these movements generally depend on actin filaments, but the underlying mechanism is unknown. Here, in Arabidopsis, we identify associations of short actin filaments along the chloroplast periphery on the plasma membrane side associated with chloroplast photorelocation and anchoring to the plasma membrane. We have termed these chloroplast-actin filaments (cp-actin filaments). Cp-actin filaments emerge from the chloroplast edge and exhibit rapid turnover. The presence of cp-actin filaments depends on an actin-binding protein, chloroplast unusual positioning1 (CHUP1), localized on the chloroplast envelope. chup1 mutant lacked cp-actin filaments but showed normal cytoplasmic actin filaments. When irradiated with blue light to induce chloroplast movement, cp-actin filaments relocalize to the leading edge of chloroplasts before and during photorelocation and are regulated by 2 phototropins, phot1 and phot2. Our findings suggest that plants evolved a unique actin-based mechanism for organelle movement.

  10. The hetero-hexameric nature of a chloroplast AAA+ FtsH protease contributes to its thermodynamic stability.

    Directory of Open Access Journals (Sweden)

    Ofer Moldavski

    Full Text Available FtsH is an evolutionary conserved membrane-bound metalloprotease complex. While in most prokaryotes FtsH is encoded by a single gene, multiple FtsH genes are found in eukaryotes. Genetic and biochemical data suggest that the Arabidopsis chloroplast FtsH is a hetero-hexamer. This raises the question why photosynthetic organisms require a heteromeric complex, whereas in most bacteria a homomeric one is sufficient. To gain structural information of the possible complexes, the Arabidopsis FtsH2 (type B and FtsH5 (type A were modeled. An in silico study with mixed models of FtsH2/5 suggests that heteromeric hexamer structure with ratio of 4:2 is more likely to exists. Specifically, calculation of the buried surface area at the interfaces between neighboring subunits revealed that a hetero-complex should be thermodynamically more stable than a homo-hexamer, due to the presence of additional hydrophobic and hydrophilic interactions. To biochemically assess this model, we generated Arabidopsis transgenic plants, expressing epitope-tagged FtsH2 and immuno-purified the protein. Mass-spectrometry analysis showed that FtsH2 is associated with FtsH1, FtsH5 and FtsH8. Interestingly, we found that 'type B' subunits (FtsH2 and FtsH8 were 2-3 fold more abundant than 'type A' (FtsH1 and FtsH5. The biochemical data corroborate the in silico model and suggest that the thylakoid FtsH hexamer is composed of two 'type A' and four 'type B' subunits.

  11. Purification of a novel lipoxygenase from eggplant (Solanum melongena) fruit chloroplasts.

    Science.gov (United States)

    Pérez-Gilabert, Manuela; López-Nicolás, José Manuel; García Carmona, Francisco

    2001-03-01

    A novel membrane lipoxygenase (LOX; EC 1.13.11.12) from eggplant (Solanum melongena L. cv. Belleza negra) fruit chloroplasts has been purified 20-fold to a specific activity of 207 enzymatic units per mg of protein with a yield of 72%. The purification was carried out by sonicating the chloroplastic membranes in the presence of Triton X-114 followed by phase partitioning and anion exchange chromatography. The purified membrane LOX preparation consisted of a single major band with an apparent molecular mass of 97 kDa after sodium dodecyl sulfate polyacrylamide gel electrophoresis. The results obtained using intact chloroplasts indicate that the enzyme is not localized in the stroma. When the enzyme reacts with linoleic acid, it produces a single peak, which comigrates with standard 9-hydroperoxy-octadecadienoic acid. A physiological role for this chloroplastic LOX is proposed.

  12. Genetic Analysis of Chloroplast Translation

    Energy Technology Data Exchange (ETDEWEB)

    Barkan, Alice

    2005-08-15

    The assembly of the photosynthetic apparatus requires the concerted action of hundreds of genes distributed between the two physically separate genomes in the nucleus and chloroplast. Nuclear genes coordinate this process by controlling the expression of chloroplast genes in response to developmental and environmental cues. However, few regulatory factors have been identified. We used mutant phenotypes to identify nuclear genes in maize that modulate chloroplast translation, a key control point in chloroplast gene expression. This project focused on the nuclear gene crp1, required for the translation of two chloroplast mRNAs. CRP1 is related to fungal proteins involved in the translation of mitochondrial mRNAs, and is the founding member of a large gene family in plants, with {approx}450 members. Members of the CRP1 family are defined by a repeated 35 amino acid motif called a ''PPR'' motif. The PPR motif is closely related to the TPR motif, which mediates protein-protein interactions. We and others have speculated that PPR tracts adopt a structure similar to that of TPR tracts, but with a substrate binding surface adapted to bind RNA instead of protein. To understand how CRP1 influences the translation of specific chloroplast mRNAs, we sought proteins that interact with CRP1, and identified the RNAs associated with CRP1 in vivo. We showed that CRP1 is associated in vivo with the mRNAs whose translation it activates. To explore the functions of PPR proteins more generally, we sought mutations in other PPR-encoding genes: mutations in the maize PPR2 and PPR4 were shown to disrupt chloroplast ribosome biogenesis and chloroplast trans-splicing, respectively. These and other results suggest that the nuclear-encoded PPR family plays a major role in modulating the expression of the chloroplast genome in higher plants.

  13. Dynamics of Chloroplast Translation during Chloroplast Differentiation in Maize.

    Directory of Open Access Journals (Sweden)

    Prakitchai Chotewutmontri

    2016-07-01

    Full Text Available Chloroplast genomes in land plants contain approximately 100 genes, the majority of which reside in polycistronic transcription units derived from cyanobacterial operons. The expression of chloroplast genes is integrated into developmental programs underlying the differentiation of photosynthetic cells from non-photosynthetic progenitors. In C4 plants, the partitioning of photosynthesis between two cell types, bundle sheath and mesophyll, adds an additional layer of complexity. We used ribosome profiling and RNA-seq to generate a comprehensive description of chloroplast gene expression at four stages of chloroplast differentiation, as displayed along the maize seedling leaf blade. The rate of protein output of most genes increases early in development and declines once the photosynthetic apparatus is mature. The developmental dynamics of protein output fall into several patterns. Programmed changes in mRNA abundance make a strong contribution to the developmental shifts in protein output, but output is further adjusted by changes in translational efficiency. RNAs with prioritized translation early in development are largely involved in chloroplast gene expression, whereas those with prioritized translation in photosynthetic tissues are generally involved in photosynthesis. Differential gene expression in bundle sheath and mesophyll chloroplasts results primarily from differences in mRNA abundance, but differences in translational efficiency amplify mRNA-level effects in some instances. In most cases, rates of protein output approximate steady-state protein stoichiometries, implying a limited role for proteolysis in eliminating unassembled or damaged proteins under non-stress conditions. Tuned protein output results from gene-specific trade-offs between translational efficiency and mRNA abundance, both of which span a large dynamic range. Analysis of ribosome footprints at sites of RNA editing showed that the chloroplast translation machinery

  14. Dynamics of Chloroplast Translation during Chloroplast Differentiation in Maize.

    Science.gov (United States)

    Chotewutmontri, Prakitchai; Barkan, Alice

    2016-07-01

    Chloroplast genomes in land plants contain approximately 100 genes, the majority of which reside in polycistronic transcription units derived from cyanobacterial operons. The expression of chloroplast genes is integrated into developmental programs underlying the differentiation of photosynthetic cells from non-photosynthetic progenitors. In C4 plants, the partitioning of photosynthesis between two cell types, bundle sheath and mesophyll, adds an additional layer of complexity. We used ribosome profiling and RNA-seq to generate a comprehensive description of chloroplast gene expression at four stages of chloroplast differentiation, as displayed along the maize seedling leaf blade. The rate of protein output of most genes increases early in development and declines once the photosynthetic apparatus is mature. The developmental dynamics of protein output fall into several patterns. Programmed changes in mRNA abundance make a strong contribution to the developmental shifts in protein output, but output is further adjusted by changes in translational efficiency. RNAs with prioritized translation early in development are largely involved in chloroplast gene expression, whereas those with prioritized translation in photosynthetic tissues are generally involved in photosynthesis. Differential gene expression in bundle sheath and mesophyll chloroplasts results primarily from differences in mRNA abundance, but differences in translational efficiency amplify mRNA-level effects in some instances. In most cases, rates of protein output approximate steady-state protein stoichiometries, implying a limited role for proteolysis in eliminating unassembled or damaged proteins under non-stress conditions. Tuned protein output results from gene-specific trade-offs between translational efficiency and mRNA abundance, both of which span a large dynamic range. Analysis of ribosome footprints at sites of RNA editing showed that the chloroplast translation machinery does not generally

  15. Conditional Depletion of the Chlamydomonas Chloroplast ClpP Protease Activates Nuclear Genes Involved in Autophagy and Plastid Protein Quality Control.

    Science.gov (United States)

    Ramundo, Silvia; Casero, David; Mühlhaus, Timo; Hemme, Dorothea; Sommer, Frederik; Crèvecoeur, Michèle; Rahire, Michèle; Schroda, Michael; Rusch, Jannette; Goodenough, Ursula; Pellegrini, Matteo; Perez-Perez, Maria Esther; Crespo, José Luis; Schaad, Olivier; Civic, Natacha; Rochaix, Jean David

    2014-05-01

    Plastid protein homeostasis is critical during chloroplast biogenesis and responses to changes in environmental conditions. Proteases and molecular chaperones involved in plastid protein quality control are encoded by the nucleus except for the catalytic subunit of ClpP, an evolutionarily conserved serine protease. Unlike its Escherichia coli ortholog, this chloroplast protease is essential for cell viability. To study its function, we used a recently developed system of repressible chloroplast gene expression in the alga Chlamydomonas reinhardtii. Using this repressible system, we have shown that a selective gradual depletion of ClpP leads to alteration of chloroplast morphology, causes formation of vesicles, and induces extensive cytoplasmic vacuolization that is reminiscent of autophagy. Analysis of the transcriptome and proteome during ClpP depletion revealed a set of proteins that are more abundant at the protein level, but not at the RNA level. These proteins may comprise some of the ClpP substrates. Moreover, the specific increase in accumulation, both at the RNA and protein level, of small heat shock proteins, chaperones, proteases, and proteins involved in thylakoid maintenance upon perturbation of plastid protein homeostasis suggests the existence of a chloroplast-to-nucleus signaling pathway involved in organelle quality control. We suggest that this represents a chloroplast unfolded protein response that is conceptually similar to that observed in the endoplasmic reticulum and in mitochondria.

  16. Conditional Depletion of the Chlamydomonas Chloroplast ClpP Protease Activates Nuclear Genes Involved in Autophagy and Plastid Protein Quality Control[W

    Science.gov (United States)

    Ramundo, Silvia; Casero, David; Mühlhaus, Timo; Hemme, Dorothea; Sommer, Frederik; Crèvecoeur, Michèle; Rahire, Michèle; Schroda, Michael; Rusch, Jannette; Goodenough, Ursula; Pellegrini, Matteo; Perez-Perez, Maria Esther; Crespo, José Luis; Schaad, Olivier; Civic, Natacha; Rochaix, Jean David

    2014-01-01

    Plastid protein homeostasis is critical during chloroplast biogenesis and responses to changes in environmental conditions. Proteases and molecular chaperones involved in plastid protein quality control are encoded by the nucleus except for the catalytic subunit of ClpP, an evolutionarily conserved serine protease. Unlike its Escherichia coli ortholog, this chloroplast protease is essential for cell viability. To study its function, we used a recently developed system of repressible chloroplast gene expression in the alga Chlamydomonas reinhardtii. Using this repressible system, we have shown that a selective gradual depletion of ClpP leads to alteration of chloroplast morphology, causes formation of vesicles, and induces extensive cytoplasmic vacuolization that is reminiscent of autophagy. Analysis of the transcriptome and proteome during ClpP depletion revealed a set of proteins that are more abundant at the protein level, but not at the RNA level. These proteins may comprise some of the ClpP substrates. Moreover, the specific increase in accumulation, both at the RNA and protein level, of small heat shock proteins, chaperones, proteases, and proteins involved in thylakoid maintenance upon perturbation of plastid protein homeostasis suggests the existence of a chloroplast-to-nucleus signaling pathway involved in organelle quality control. We suggest that this represents a chloroplast unfolded protein response that is conceptually similar to that observed in the endoplasmic reticulum and in mitochondria. PMID:24879428

  17. The TOC complex: preprotein gateway to the chloroplast.

    Science.gov (United States)

    Andrès, Charles; Agne, Birgit; Kessler, Felix

    2010-06-01

    Photosynthetic eukaryotes strongly depend on chloroplast metabolic pathways. Most if not all involve nuclear encoded proteins. These are synthesized as cytosolic preproteins with N-terminal, cleavable targeting sequences (transit peptide). Preproteins are imported by a major pathway composed of two proteins complexes: TOC and TIC (Translocon of the Outer and Inner membranes of the Chloroplasts, respectively). These selectively recognize the preproteins and facilitate their transport across the chloroplast envelope. The TOC core complex consists of three types of components, each belonging to a small family: Toc34, Toc75 and Toc159. Toc34 and Toc159 isoforms represent a subfamily of the GTPase superfamily. The members of the Toc34 and Toc159 subfamily act as GTP-dependent receptors at the chloroplast surface and distinct members of each occur in defined, substrate-specific TOC complexes. Toc75, a member of the Omp85 family, is conserved from prokaryotes and functions as the unique protein-conducting channel at the outer membrane. In this review we will describe the current state of knowledge regarding the composition and function of the TOC complex.

  18. Membraner

    DEFF Research Database (Denmark)

    Bach, Finn

    2009-01-01

    Notatet giver en kort introduktion til den statiske virkemåde af membraner og membrankonstruktioner......Notatet giver en kort introduktion til den statiske virkemåde af membraner og membrankonstruktioner...

  19. Chloroplast avoidance movement is not functional in plants grown under strong sunlight.

    Science.gov (United States)

    Higa, Takeshi; Wada, Masamitsu

    2016-04-01

    Chloroplast movement in nine climbing plant species was investigated. It is thought that chloroplasts generally escape from strong light to avoid photodamage but accumulate towards weak light to perform photosynthesis effectively. Unexpectedly, however, the leaves of climbing plants grown under strong sunlight showed very low or no chloroplast photorelocation responses to either weak or strong blue light when detected by red light transmittance through leaves. Direct observations of Cayratia japonica leaves, for example, revealed that the average number of chloroplasts in upper periclinal walls of palisade tissue cells was only 1.2 after weak blue-light irradiation and almost all of the chloroplasts remained at the anticlinal wall, the state of chloroplast avoidance response. The leaves grown under strong light have thin and columnar palisade tissue cells comparing with the leaves grown under low light. Depending on our analyses and our schematic model, the thinner cells in a unit leaf area have a wider total plasma membrane area, such that more chloroplasts can exist on the plasma membrane in the thinner cells than in the thicker cells in a unit leaf-area basis. The same strategy might be used in other plant leaves grown under direct sunlight.

  20. The Arabidopsis Thylakoid Protein PAM68 Is Required for Efficient D1 Biogenesis and Photosystem II Assembly[W

    Science.gov (United States)

    Armbruster, Ute; Zühlke, Jessica; Rengstl, Birgit; Kreller, Renate; Makarenko, Elina; Rühle, Thilo; Schünemann, Danja; Jahns, Peter; Weisshaar, Bernd; Nickelsen, Jörg; Leister, Dario

    2010-01-01

    Photosystem II (PSII) is a multiprotein complex that functions as a light-driven water:plastoquinone oxidoreductase in photosynthesis. Assembly of PSII proceeds through a number of distinct intermediate states and requires auxiliary proteins. The photosynthesis affected mutant 68 (pam68) of Arabidopsis thaliana displays drastically altered chlorophyll fluorescence and abnormally low levels of the PSII core subunits D1, D2, CP43, and CP47. We show that these phenotypes result from a specific decrease in the stability and maturation of D1. This is associated with a marked increase in the synthesis of RC (the PSII reaction center-like assembly complex) at the expense of PSII dimers and supercomplexes. PAM68 is a conserved integral membrane protein found in cyanobacterial and eukaryotic thylakoids and interacts in split-ubiquitin assays with several PSII core proteins and known PSII assembly factors. Biochemical analyses of thylakoids from Arabidopsis and Synechocystis sp PCC 6803 suggest that, during PSII assembly, PAM68 proteins associate with an early intermediate complex that might contain D1 and the assembly factor LPA1. Inactivation of cyanobacterial PAM68 destabilizes RC but does not affect larger PSII assembly complexes. Our data imply that PAM68 proteins promote early steps in PSII biogenesis in cyanobacteria and plants, but their inactivation is differently compensated for in the two classes of organisms. PMID:20923938

  1. The Arabidopsis thylakoid protein PAM68 is required for efficient D1 biogenesis and photosystem II assembly.

    Science.gov (United States)

    Armbruster, Ute; Zühlke, Jessica; Rengstl, Birgit; Kreller, Renate; Makarenko, Elina; Rühle, Thilo; Schünemann, Danja; Jahns, Peter; Weisshaar, Bernd; Nickelsen, Jörg; Leister, Dario

    2010-10-01

    Photosystem II (PSII) is a multiprotein complex that functions as a light-driven water:plastoquinone oxidoreductase in photosynthesis. Assembly of PSII proceeds through a number of distinct intermediate states and requires auxiliary proteins. The photosynthesis affected mutant 68 (pam68) of Arabidopsis thaliana displays drastically altered chlorophyll fluorescence and abnormally low levels of the PSII core subunits D1, D2, CP43, and CP47. We show that these phenotypes result from a specific decrease in the stability and maturation of D1. This is associated with a marked increase in the synthesis of RC (the PSII reaction center-like assembly complex) at the expense of PSII dimers and supercomplexes. PAM68 is a conserved integral membrane protein found in cyanobacterial and eukaryotic thylakoids and interacts in split-ubiquitin assays with several PSII core proteins and known PSII assembly factors. Biochemical analyses of thylakoids from Arabidopsis and Synechocystis sp PCC 6803 suggest that, during PSII assembly, PAM68 proteins associate with an early intermediate complex that might contain D1 and the assembly factor LPA1. Inactivation of cyanobacterial PAM68 destabilizes RC but does not affect larger PSII assembly complexes. Our data imply that PAM68 proteins promote early steps in PSII biogenesis in cyanobacteria and plants, but their inactivation is differently compensated for in the two classes of organisms.

  2. Diversity in biosynthetic pathways of galactolipids in the light of endosymbiotic origin of chloroplasts

    Directory of Open Access Journals (Sweden)

    Naoki eSato

    2016-02-01

    Full Text Available Cyanobacteria and chloroplasts perform oxygenic photosynthesis, and share a common origin. Galactolipids are present in the photosynthetic membranes of both cyanobacteria and chloroplasts, but the biosynthetic pathways of the galactolipids are significantly different in the two systems. In this minireview, we explain the history of the discovery of the cyanobacterial pathway, and present a probable scenario of the evolution of the two pathways.

  3. MapA, an iron-regulated, cytoplasmic membrane protein in the cyanobacterium Synechococcus sp. strain PCC7942.

    Science.gov (United States)

    Webb, R; Troyan, T; Sherman, D; Sherman, L A

    1994-08-01

    Growth of Synechococcus sp. strain PCC 7942 in iron-deficient media leads to the accumulation of an approximately 34-kDa protein. The gene encoding this protein, mapA (membrane-associated protein A), has been cloned and sequenced (GenBank accession number, L01621). The mapA transcript is not detectable in normally grown cultures but is stably accumulated by cells grown in iron-deficient media. However, the promoter sequence for this gene does not resemble other bacterial iron-regulated promoters described to date. The carboxyl-terminal region of the derived amino acid sequence of MapA resembles bacterial proteins involved in iron acquisition, whereas the amino-terminal end of MapA has a high degree of amino acid identity with the abundant, chloroplast envelope protein E37. An approach employing improved cellular fractionation techniques as well as electron microscopy and immunocytochemistry was essential in localizing MapA protein to the cytoplasmic membrane of Synechococcus sp. strain PCC 7942. When these cells were grown under iron-deficient conditions, a significant fraction of MapA could also be localized to the thylakoid membranes.

  4. Chloroplast in Plant-Virus Interaction

    Science.gov (United States)

    Zhao, Jinping; Zhang, Xian; Hong, Yiguo; Liu, Yule

    2016-01-01

    In plants, the chloroplast is the organelle that conducts photosynthesis. It has been known that chloroplast is involved in virus infection of plants for approximate 70 years. Recently, the subject of chloroplast-virus interplay is getting more and more attention. In this article we discuss the different aspects of chloroplast-virus interaction into three sections: the effect of virus infection on the structure and function of chloroplast, the role of chloroplast in virus infection cycle, and the function of chloroplast in host defense against viruses. In particular, we focus on the characterization of chloroplast protein-viral protein interactions that underlie the interplay between chloroplast and virus. It can be summarized that chloroplast is a common target of plant viruses for viral pathogenesis or propagation; and conversely, chloroplast and its components also can play active roles in plant defense against viruses. Chloroplast photosynthesis-related genes/proteins (CPRGs/CPRPs) are suggested to play a central role during the complex chloroplast-virus interaction. PMID:27757106

  5. Chloroplast evolution: secondary symbiogenesis and multiple losses.

    Science.gov (United States)

    Cavalier-Smith, T

    2002-01-22

    Chloroplasts originated from cyanobacteria only once, but have been laterally transferred to other lineages by symbiogenetic cell mergers. Such secondary symbiogenesis is rarer and chloroplast losses commoner than often assumed.

  6. Phototropin encoded by a single-copy gene mediates chloroplast photorelocation movements in the liverwort Marchantia polymorpha.

    Science.gov (United States)

    Komatsu, Aino; Terai, Mika; Ishizaki, Kimitsune; Suetsugu, Noriyuki; Tsuboi, Hidenori; Nishihama, Ryuichi; Yamato, Katsuyuki T; Wada, Masamitsu; Kohchi, Takayuki

    2014-09-01

    Blue-light-induced chloroplast photorelocation movement is observed in most land plants. Chloroplasts move toward weak-light-irradiated areas to efficiently absorb light (the accumulation response) and escape from strong-light-irradiated areas to avoid photodamage (the avoidance response). The plant-specific kinase phototropin (phot) is the blue-light receptor for chloroplast movements. Although the molecular mechanisms for chloroplast photorelocation movement have been analyzed, the overall aspects of signal transduction common to land plants are still unknown. Here, we show that the liverwort Marchantia polymorpha exhibits the accumulation and avoidance responses exclusively induced by blue light as well as specific chloroplast positioning in the dark. Moreover, in silico and Southern-blot analyses revealed that the M. polymorpha genome encodes a single PHOT gene, MpPHOT, and its knockout line displayed none of the chloroplast photorelocation movements, indicating that the sole MpPHOT gene mediates all types of movement. Mpphot was localized on the plasma membrane and exhibited blue-light-dependent autophosphorylation both in vitro and in vivo. Heterologous expression of MpPHOT rescued the defects in chloroplast movement of phot mutants in the fern Adiantum capillus-veneris and the seed plant Arabidopsis (Arabidopsis thaliana). These results indicate that Mpphot possesses evolutionarily conserved regulatory activities for chloroplast photorelocation movement. M. polymorpha offers a simple and versatile platform for analyzing the fundamental processes of phototropin-mediated chloroplast photorelocation movement common to land plants.

  7. A TIR-NBS protein encoded by Arabidopsis Chilling Sensitive 1 (CHS1) limits chloroplast damage and cell death at low temperature.

    Science.gov (United States)

    Zbierzak, Anna Maria; Porfirova, Svetlana; Griebel, Thomas; Melzer, Michael; Parker, Jane E; Dörmann, Peter

    2013-08-01

    Survival of plants at low temperature depends on mechanisms for limiting physiological damage and maintaining growth. We mapped the chs1-1 (chilling sensitive1-1) mutation in Arabidopsis accession Columbia to the TIR-NBS gene At1g17610. In chs1-1, a single amino acid exchange at the CHS1 N-terminus close to the conserved TIR domain creates a stable mutant protein that fails to protect leaves against chilling stress. The sequence of another TIR-NBS gene (At5g40090) named CHL1 (CHS1-like 1) is related to that of CHS1. Over-expression of CHS1 or CHL1 alleviates chilling damage and enhances plant growth at moderate (24°C) and chilling (13°C) temperatures, suggesting a role for both proteins in growth homeostasis. chs1-1 mutants show induced salicylic acid production and defense gene expression at 13°C, indicative of autoimmunity. Genetic analysis of chs1-1 in combination with defense pathway mutants shows that chs1-1 chilling sensitivity requires the TIR-NBS-LRR and basal resistance regulators encoded by EDS1 and PAD4 but not salicylic acid. By following the timing of metabolic, physiological and chloroplast ultrastructural changes in chs1-1 leaves during chilling, we have established that alterations in photosynthetic complexes and thylakoid membrane integrity precede leaf cell death measured by ion leakage. At 24°C, the chs1-1 mutant appears normal but produces a massive necrotic response to virulent Pseudomonas syringae pv. tomato infection, although this does not affect bacterial proliferation. Our results suggest that CHS1 acts at an intersection between temperature sensing and biotic stress pathway activation to maintain plant performance over a range of conditions.

  8. Proton equilibration in the chloroplast modulates multiphasic kinetics of nonphotochemical quenching of fluorescence in plants.

    Science.gov (United States)

    Joliot, Pierre A; Finazzi, Giovanni

    2010-07-13

    In plants, the major route for dissipating excess light is the nonphotochemical quenching of absorbed light (NPQ), which is associated with thylakoid lumen acidification. Our data offer an interpretation for the complex relationship between changes in luminal pH and the NPQ response. Upon steady-state illumination, fast NPQ relaxation in the dark reflects the equilibration between the electrochemical proton gradient established in the light and the cellular ATP/ADP+Pi ratio. This is followed by a slower phase, which reflects the decay of the proton motive force at equilibrium, due to gradual cellular ATP consumption. In transient conditions, a sustained lag appears in both quenching onset and relaxation, which is modulated by the size of the antenna complexes of photosystem II and by cyclic electron flow around photosystem I. We propose that this phenomenon reflects the signature of protonation of specific domains in the antenna and of slow H(+) diffusion in the different domains of the chloroplast.

  9. Evolution of chloroplast vesicle transport.

    Science.gov (United States)

    Westphal, Sabine; Soll, Jürgen; Vothknecht, Ute C

    2003-02-01

    Vesicle traffic plays a central role in eukaryotic transport. The presence of a vesicle transport system inside chloroplasts of spermatophytes raises the question of its phylogenetic origin. To elucidate the evolution of this transport system we analyzed organisms belonging to different lineages that arose from the first photosynthetic eukaryote, i.e. glaucocystophytes, chlorophytes, rhodophytes, and charophytes/embryophytes. Intriguingly, vesicle transport is not apparent in any group other than embryophytes. The transfer of this eukaryotic-type vesicle transport system from the cytosol into the chloroplast thus seems a late evolutionary development that was acquired by land plants in order to adapt to new environmental challenges.

  10. THRUMIN1 is a light-regulated actin-bundling protein involved in chloroplast motility.

    Science.gov (United States)

    Whippo, Craig W; Khurana, Parul; Davis, Phillip A; DeBlasio, Stacy L; DeSloover, Daniel; Staiger, Christopher J; Hangarter, Roger P

    2011-01-11

    Chloroplast movement in response to changing light conditions optimizes photosynthetic light absorption. This repositioning is stimulated by blue light perceived via the phototropin photoreceptors and is transduced to the actin cytoskeleton. Some actin-based motility systems use filament reorganizations rather than myosin-based translocations. Recent research favors the hypothesis that chloroplast movement is driven by actin reorganization at the plasma membrane, but no proteins affecting chloroplast movements have been shown to associate with both the plasma membrane and actin filaments in vivo. Here we identified THRUMIN1 as a critical link between phototropin photoreceptor activity at the plasma membrane and actin-dependent chloroplast movements. THRUMIN1 bundles filamentous actin in vitro, and it localizes to the plasma membrane and displays light- and phototropin-dependent localization to microfilaments in vivo. These results suggest that phototropin-induced actin bundling via THRUMIN1 is important for chloroplast movement. A mammalian homolog of THRUMIN1, GRXCR1, has been implicated in auditory responses and hair cell stereocilla development as a regulator of actin architecture. Studies of THRUMIN1 will help elucidate the function of this family of eukaryotic proteins.

  11. Effects of Ce3+ on Chloroplast Senescence of Spinach under Light

    Institute of Scientific and Technical Information of China (English)

    Yang Fan; Ma Zhenni; Liu Chao; Wu Cheng; Zhou Juan; Gao Fengqing; Hong Fashui

    2005-01-01

    The effects of Ce3+ on the chloroplast senescence of spinach under light were studied. The results show that when the chloroplasts are illuminated for 1, 5 and 10 min with 500 μmol·cm-2·min-1 light intensity, the oxygen evolution rate is rapidly increased. When the chloroplasts are treated for 20, 30 and 40 min with 500 μmol·cm-2·min-1 light intensity, the oxygen evolution rate is gradually decreased. While spinach is treated with 16 μmol·L-1 Ce3+, the rate of oxygen evolution of chloroplasts in different illumination time (1,5, 10, 20, 30, 40 min) is higher than that of control, and when illumination time is over 10 min, the reduction of the oxygen evolution rate is lower than that of control. It suggests that Ce3+ treatment can protect chloroplasts from aging for long time illumination. The mechanism research results indicate that Ce3+ treatment can significantly decrease accumulation of active oxygen free radicals such as O2·- and H2O2, and reduce the level of malondialdehyde (MDA), and maintain stability of membrane structure of chloroplast under light. It is shown that the redox took place between cerium and free radicals, which are eliminated in a large number, leading to protect the membrane from peroxidating.

  12. Dependence of chlorophyll P700 redox transients during the induction period on the transmembrane distribution of protons in chloroplasts of pea leaves

    NARCIS (Netherlands)

    Bulychev, A. A.; Cherkashin, A. A.; Rubin, A. B.

    2010-01-01

    Differential absorbance measurements and fluorometry were applied to examine the impact of dicyclohexylcarbodiimide (DCCD, an inhibitor of H(+) conductance in thylakoid membranes) and nigericin (a K(+)/H(+) antiporter) on photoinduced redox state transients of chlorophyll P700 and the induction curv

  13. Programmed chloroplast destruction during leaf senescence involves 13-lipoxygenase (13-LOX).

    Science.gov (United States)

    Springer, Armin; Kang, ChulHee; Rustgi, Sachin; von Wettstein, Diter; Reinbothe, Christiane; Pollmann, Stephan; Reinbothe, Steffen

    2016-03-22

    Leaf senescence is the terminal stage in the development of perennial plants. Massive physiological changes occur that lead to the shut down of photosynthesis and a cessation of growth. Leaf senescence involves the selective destruction of the chloroplast as the site of photosynthesis. Here, we show that 13-lipoxygenase (13-LOX) accomplishes a key role in the destruction of chloroplasts in senescing plants and propose a critical role of its NH2-terminal chloroplast transit peptide. The 13-LOX enzyme identified here accumulated in the plastid envelope and catalyzed the dioxygenation of unsaturated membrane fatty acids, leading to a selective destruction of the chloroplast and the release of stromal constituents. Because 13-LOX pathway products comprise compounds involved in insect deterrence and pathogen defense (volatile aldehydes and oxylipins), a mechanism of unmolested nitrogen and carbon relocation is suggested that occurs from leaves to seeds and roots during fall.

  14. Localization of Membrane Proteins in the Cyanobacterium Synechococcus sp. PCC7942 (Radial Asymmetry in the Photosynthetic Complexes).

    Science.gov (United States)

    Sherman, D. M.; Troyan, T. A.; Sherman, L. A.

    1994-09-01

    Localization of membrane proteins in the cyanobacterium Synechococcus sp. PCC7942 was determined by transmission electron microscopy utilizing immunocytochemistry with cells prepared by freeze-substitution. This preparation procedure maintained cellular morphology and permitted detection of cellular antigens with high sensitivity and low background. Synechococcus sp. PCC7942 is a unicellular cyanobacterium with thylakoids organized in concentric layers toward the periphery of the cell. Cytochrome oxidase was localized almost entirely in the cytoplasmic membrane, whereas a carotenoprotein (P35) was shown to be a cell wall component. The major photosystem II (PSII) proteins (D1, D2 CP43, and CP47) were localized throughout the thylakoids. Proteins of the Cyt b6/f complex were found to have a similar distribution. Thylakoid luminal proteins, such as the Mn-stabilizing protein, were located primarily in the thylakoid, but a small, reproducible fraction was found in the outer compartment. The photosystem I (PSI) reaction center proteins and the ATP synthase proteins were found associated mostly with the outermost thylakoid and with the cytoplasmic membrane. These results indicated that the photosynthetic apparatus is not evenly distributed throughout the thylakoids. Rather, there is a radial asymmetry such that much of the PSI and the ATPase synthase is located in the outermost thylakoid. The relationship of this structure to the photosynthetic mechanism is discussed. It is suggested that the photosystems are separated because of kinetic differences between PSII and PSI, as hypothesized by H.-W. Trissl and C. Wilhelm (Trends Biochem Sci [1993] 18:415-419).

  15. Chloroplast proteins without cleavable transit peptides: rare exceptions or a major constituent of the chloroplast proteome?

    Science.gov (United States)

    Armbruster, Ute; Hertle, Alexander; Makarenko, Elina; Zühlke, Jessica; Pribil, Mathias; Dietzmann, Angela; Schliebner, Ivo; Aseeva, Elena; Fenino, Elena; Scharfenberg, Michael; Voigt, Christian; Leister, Dario

    2009-11-01

    Most chloroplast proteins (cp proteins) are nucleus-encoded, synthesized on cytosolic ribosomes as precursor proteins containing a presequence (cTP), and post-translationally imported via the Tic/Toc complex into the organelle, where the cTP is removed. Only a few unambiguous instances of cp proteins that do not require cTPs (non-canonical cp proteins) have been reported so far. However, the survey of data from large-scale proteomic studies presented here suggests that the fraction of such proteins in the total cp proteome might be as large as approximately 30%. To explore this discrepancy, we chose a representative set of 28 putative non-canonical cp proteins, and used in vitro import and Red Fluorescent Protein (RFP)-fusion assays to determine their sub-cellular destinations. Four proteins, including embryo defective 1211, glycolate oxidase 2, protein disulfide isomerase-like protein (PDII), and a putative glutathione S-transferase, could be unambiguously assigned to the chloroplast. Several others ('potential cp proteins') were found to be imported into chloroplasts in vitro, but failed to localize to the organelle when RFP was fused to their C-terminal ends. Extrapolations suggest that the fraction of cp proteins that enter the inner compartments of the organelle, although they lack a cTP, might be as large as 11.4% of the total cp proteome. Our data also support the idea that cytosolic proteins that associate with the cp outer membrane might account for false positive cp proteins obtained in earlier studies.

  16. The 1.7 Å resolution structure of At2g44920, a pentapeptide-repeat protein in the thylakoid lumen of Arabidopsis thaliana

    Energy Technology Data Exchange (ETDEWEB)

    Ni, Shuisong; McGookey, Michael E.; Tinch, Stuart L.; Jones, Alisha N.; Jayaraman, Seetharaman; Tong, Liang; Kennedy, Michael A. (Miami U); (Columbia)

    2012-01-09

    At2g44920 belongs to a diverse family (Pfam PF00805) of pentapeptide-repeat proteins (PRPs) that are present in all known organisms except yeast. PRPs contain at least eight tandem-repeating sequences of five amino acids with an approximate consensus sequence (STAV)(D/N)(L/F)(S/T/R)(X). Recent crystal structures show that PRPs adopt a highly regular four-sided right-handed {beta}-helical structure consisting mainly of type II and type IV {beta}-turns, sometimes referred to as a repeated five-residue (or Rfr) fold. Among sequenced genomes, PRP genes are most abundant in cyanobacteria, leading to speculation that PRPs play an important role in the unique lifestyle of photosynthetic cyanobacteria. Despite the recent structural characterization of several cyanobacterial PRPs, most of their functions remain unknown. Plants, whose chloroplasts are of cyanobacterial origin, have only four PRP genes in their genomes. At2g44920 is one of three PRPs located in the thylakoid lumen. Here, the crystal structure of a double methionine mutant of residues 81-224 of At2g44920, the naturally processed fragment of one of its full-length isoforms, is reported at 1.7 {angstrom} resolution. The structure of At2g44920 consists of the characteristic Rfr fold with five uninterrupted coils made up of 25 pentapeptide repeats and {alpha}-helical elements capping both termini. A disulfide bridge links the two {alpha}-helices with a conserved loop between the helical elements at its C-terminus. This structure represents the first structure of a PRP protein whose subcellular location has been experimentally confirmed to be the thylakoid lumen in a plant species.

  17. External Ca(2+) is essential for chloroplast movement induced by mechanical stimulation but not by light stimulation.

    Science.gov (United States)

    Sato, Y; Wada, M; Kadota, A

    2001-10-01

    In the fern Adiantum capillus-veneris, chloroplast movement is induced by mechanical stimulation as well as by light stimulation. Directional movement of both types depends on an actin-based motile system. To investigate the physiological relationship between mechanical and light signaling in the regulation of chloroplast movement, we examined the mechano-response of chloroplasts whose motility had been already restricted after photo-relocation. Chloroplast mechano-avoidance movement was induced under all of the photo-relocation conditions tested, indicating that mechano-specific signals generated by mechanical stimulation dominate over the light signals and reactivate the motility of chloroplasts. When the effects of external Ca(2+) on the induction of mechano- and light responses were examined, strikingly different requirements of external Ca(2+) were found for each. In medium without Ca(2+), the mechano-response was suppressed but no effects were observed on photo-response. Mechano-relocation movement of chloroplasts was inhibited by 100 microM lanthanum (La(3+)), a plasma membrane calcium channel blocker, and by 10 microM gadolinium (Gd(3+)), a stretch-activated channel blocker. However, the same concentrations of these drugs did not affect the photo-relocation movement at all. These results suggest that the influx of external Ca(2+) is crucial for the early signaling step of chloroplast mechano-relocation but not for that of photo-relocation. This is the first report showing the separation of signaling pathways in mechano- and photo-relocation of chloroplasts.

  18. Light-Induced Movements of Chloroplasts and Nuclei Are Regulated in Both Cp-Actin-Filament-Dependent and -Independent Manners in Arabidopsis thaliana.

    Directory of Open Access Journals (Sweden)

    Noriyuki Suetsugu

    Full Text Available Light-induced chloroplast movement and attachment to the plasma membrane are dependent on actin filaments. In Arabidopsis thaliana, the short actin filaments on the chloroplast envelope, cp-actin filaments, are essential for chloroplast movement and positioning. Furthermore, cp-actin-filament-mediated chloroplast movement is necessary for the strong-light-induced nuclear avoidance response. The proteins CHLOROPLAST UNUSUAL POSITIONING 1 (CHUP1, KINESIN-LIKE PROTEIN FOR ACTIN-BASED CHLOROPLAST MOVEMENT 1 (KAC1 and KAC2 are required for the generation and/or maintenance of cp-actin filaments in Arabidopsis. In land plants, CHUP1 and KAC family proteins play pivotal roles in the proper movement of chloroplasts and their attachment to the plasma membrane. Here, we report similar but distinct phenotypes in chloroplast and nuclear photorelocation movements between chup1 and kac1kac2 mutants. Measurement of chloroplast photorelocation movement indicated that kac1kac2, but not chup1, exhibited a clear strong-light-induced increase in leaf transmittance changes. The chloroplast movement in kac1kac2 depended on phototropin 2, CHUP1 and two other regulators for cp-actin filaments, PLASTID MOVEMENT IMPAIRED 1 and THRUMIN 1. Furthermore, kac1kac2 retained a weak but significant nuclear avoidance response although chup1 displayed a severe defect in the nuclear avoidance response. The kac1kac2chup1 triple mutant was completely defective in both chloroplast and nuclear avoidance responses. These results indicate that CHUP1 and the KACs function somewhat independently, but interdependently mediate both chloroplast and nuclear photorelocation movements.

  19. Light-Induced Movements of Chloroplasts and Nuclei Are Regulated in Both Cp-Actin-Filament-Dependent and -Independent Manners in Arabidopsis thaliana.

    Science.gov (United States)

    Suetsugu, Noriyuki; Higa, Takeshi; Gotoh, Eiji; Wada, Masamitsu

    2016-01-01

    Light-induced chloroplast movement and attachment to the plasma membrane are dependent on actin filaments. In Arabidopsis thaliana, the short actin filaments on the chloroplast envelope, cp-actin filaments, are essential for chloroplast movement and positioning. Furthermore, cp-actin-filament-mediated chloroplast movement is necessary for the strong-light-induced nuclear avoidance response. The proteins CHLOROPLAST UNUSUAL POSITIONING 1 (CHUP1), KINESIN-LIKE PROTEIN FOR ACTIN-BASED CHLOROPLAST MOVEMENT 1 (KAC1) and KAC2 are required for the generation and/or maintenance of cp-actin filaments in Arabidopsis. In land plants, CHUP1 and KAC family proteins play pivotal roles in the proper movement of chloroplasts and their attachment to the plasma membrane. Here, we report similar but distinct phenotypes in chloroplast and nuclear photorelocation movements between chup1 and kac1kac2 mutants. Measurement of chloroplast photorelocation movement indicated that kac1kac2, but not chup1, exhibited a clear strong-light-induced increase in leaf transmittance changes. The chloroplast movement in kac1kac2 depended on phototropin 2, CHUP1 and two other regulators for cp-actin filaments, PLASTID MOVEMENT IMPAIRED 1 and THRUMIN 1. Furthermore, kac1kac2 retained a weak but significant nuclear avoidance response although chup1 displayed a severe defect in the nuclear avoidance response. The kac1kac2chup1 triple mutant was completely defective in both chloroplast and nuclear avoidance responses. These results indicate that CHUP1 and the KACs function somewhat independently, but interdependently mediate both chloroplast and nuclear photorelocation movements.

  20. Chloroplast avoidance movement reduces photodamage in plants.

    Science.gov (United States)

    Kasahara, Masahiro; Kagawa, Takatoshi; Oikawa, Kazusato; Suetsugu, Noriyuki; Miyao, Mitsue; Wada, Masamitsu

    When plants are exposed to light levels higher than those required for photosynthesis, reactive oxygen species are generated in the chloroplasts and cause photodamage. This can occur even under natural growth conditions. To mitigate photodamage, plants have developed several protective mechanisms. One is chloroplast avoidance movement, in which chloroplasts move from the cell surface to the side walls of cells under high light conditions, although experimental support is still awaited. Here, using different classes of mutant defective in chloroplast avoidance movement, we show that these mutants are more susceptible to damage in high light than wild-type plants. Damage of the photosynthetic apparatus and subsequent bleaching of leaf colour and necrosis occur faster under high light conditions in the mutants than in wild-type plants. We conclude that chloroplast avoidance movement actually decreases the amount of light absorption by chloroplasts, and might therefore be important to the survival of plants under natural growth conditions.

  1. Blue-light-induced rapid chloroplast de-anchoring in Vallisneria epidermal cells

    Institute of Scientific and Technical Information of China (English)

    Yuuki Sakai; Shin-Ichiro Inoue; Akiko Harada; Ken-Ichiro Shimazaki; Shingo Takagi

    2015-01-01

    In the outer periclinal cytoplasm of leaf epidermal cells of an aquatic angiosperm Vallisneria, blue light induces “chloroplast de‐anchoring”, a rapid decline in the resistance of chloroplasts against centrifugal force. Chloroplast deanchoring is known induced within 1 min of irradiation with high‐fluence‐rate blue light specifically, preceding the commencement of chloroplasts migration toward the anticlinal cytoplasm. However, its regulatory mechanism has remained elusive, although pharmacological analysis suggested that a calcium release from intracellular calcium stores is necessary for the response. In search of the responsible photoreceptors, immunoblotting analysis using antibodies against phototropins demonstrated that cross‐reactive polypeptides of 120‐kDa exist in the plasma‐membrane fraction prepared from the leaves. In vitro phosphorylation analysis revealed that 120‐kDa polypeptides were phosphorylated by exposure to blue light in a fluence‐dependent manner. The blue‐light‐induced phosphorylation activity was sensitive to a Ser/Thr kinase inhibitor, staurosporine, and unusually was retained at a high level for a long time in darkness. Furthermore, phototropin gene homologs (Vallisneria PHOTOTROPIN1 and PHOTOTROPIN2) expressed in leaves were isolated. We propose that calciumregulated chloroplast de‐anchoring, possibly mediated by phototropins, is an initial process of the blue‐light‐induced avoidance response of chloroplasts in Vallisneria.

  2. Loss‐of‐function mutation of rice SLAC7 decreases chloroplast stability and induces a photoprotection mechanism in rice

    OpenAIRE

    Fan, Xiaolei; Wu, Jiemin; Chen, Taiyu; Tie, Weiwei; Chen, Hao; ZHOU, Fei; Lin, Yongjun

    2015-01-01

    Abstract Plants absorb sunlight to power the photochemical reactions of photosynthesis, which can potentially damage the photosynthetic machinery. However, the mechanism that protects chloroplasts from the damage remains unclear. In this work, we demonstrated that rice (Oryza sativa L.) SLAC7 is a generally expressed membrane protein. Loss‐of‐function of SLAC7 caused continuous damage to the chloroplasts of mutant leaves under normal light conditions. Ion leakage indicators related to leaf da...

  3. Mechanisms of Protein Synthesis in Chloroplasts: How to Design Translatable mRNAs in Chloroplasts

    Institute of Scientific and Technical Information of China (English)

    M. Sugiura

    2007-01-01

    @@ Chloroplast transformation provides a powerful tool to produce useful proteins in plants. After completion of the chloroplast genome sequencing from tobacco plants (Shinozaki et al., 1986, Yukawa et al., 2005), Pal Maliga group developed the high-frequency chloroplast transformation system in tobacco (Svab and Maliga, 1993).

  4. Isolation of Chloroplasts from Plant Protoplasts.

    Science.gov (United States)

    Lung, Shiu-Cheung; Smith, Matthew D; Chuong, Simon D X

    2015-10-01

    Chloroplasts can be isolated from higher plants directly following homogenization; however, the resulting yield, purity, and intactness are often low, necessitating a large amount of starting material. This protocol is optimized to produce a high yield of pure chloroplasts from isolated Arabidopsis protoplasts. The two-part method is a simple, scaled-down, and low-cost procedure that readily provides healthy mesophyll protoplasts, which are then ruptured to release intact chloroplasts. Chloroplasts isolated using this method are competent for use in biochemical, cellular, and molecular analyses.

  5. Chloroplast Dynamics and Photosynthetic Efficiency: Final Technical Report

    Energy Technology Data Exchange (ETDEWEB)

    Hanson, Maureen [Cornell Univ., Ithaca, NY (United States)

    2016-11-03

    This project investigated the mechanism by which chloroplasts position themselves to maximize solar energy utilization, to enhance gas exchange, to minimize environmental stress, and to promote efficient exchange of metabolites with other compartments within the plant cell. Chloroplasts move within leaf cells to optimize light levels, moving toward levels of light useful for photosynthesis while moving away from excess light. Plastids sometimes extend their reach by sending out projections (stromules) that can connect anchor chloroplasts in position within the cell or provide close contacts with plasma membrane, mitochondria, peroxisomes, endoplasmic reticulum, and the nucleus. The intracellular location of chloroplasts in relation to other organelles with which they share biosynthetic pathways, such as peroxisomes and mitochondria in photorespiration, affects metabolite flow. This work contributed to the knowledge of the mechanisms of organelle movement and anchoring in specific locations in plant cells and how proteins traffic within the cell. We identified two domains on 12 of the 13 Arabidopsis myosins that were similar to the vacuole-binding (V) domain characterized in yeast and to the DIL domain characterized in yeast and mouse as required for secretory vesicle or melanosome movement, respectively. Because all of the Arabidopsis regions with homology to the V domain contain the amino acid sequence PAL, we refer to this region as the Arabidopsis PAL domain. We have used the yeast Myo2p tail structural information to model the 12 myosin XI tail domains containing the homologous PAL and DIL domains. Eight YFP::DIL domain fusions labeled peroxisomes; none labeled mitochondria or chloroplasts. Six myosin XI Vacuole domains labeled mitochondria and seven labeled Golgi bodies. The Arabidopsis myosin XI-F PAL domain and the homologous myosin XI-F PAL domain from N. benthamiana labels chloroplasts and stromules in N. benthamiana leaves. Using an Arabidopsis line

  6. Recherche de nouveaux systèmes de transport à travers l'enveloppe du chloroplaste. Caractérisation de nouvelles protéines hydrophobes.

    OpenAIRE

    Seigneurin-Berny, Daphné

    2000-01-01

    The chloroplast is an organelle totally integrated in the metabolism of the plant cell. It contains its own metabolic pathways like photosynthesis, aminoacid synthesis. The chloroplast is limited by the envelope composed of two membranes and an intermembrane space. Envelope membranes are the site of transport of metabolites, ions, proteins and information between the plastid and the cytosol. Then, they contain many transport systems, but only some of them have been characterised. Hydrophobici...

  7. In vitro comparative kinetic analysis of the chloroplast Toc GTPases.

    Science.gov (United States)

    Reddick, L Evan; Vaughn, Michael D; Wright, Sarah J; Campbell, Ian M; Bruce, Barry D

    2007-04-13

    A unique aspect of protein transport into plastids is the coordinate involvement of two GTPases in the translocon of the outer chloroplast membrane (Toc). There are two subfamilies in Arabidopsis, the small GTPases (Toc33 and Toc34) and the large acidic GTPases (Toc90, Toc120, Toc132, and Toc159). In chloroplasts, Toc34 and Toc159 are implicated in precursor binding, yet mechanistic details are poorly understood. How the GTPase cycle is modulated by precursor binding is complex and in need of careful dissection. To this end, we have developed novel in vitro assays to quantitate nucleotide binding and hydrolysis of the Toc GTPases. Here we present the first systematic kinetic characterization of four Toc GTPases (cytosolic domains of atToc33, atToc34, psToc34, and the GTPase domain of atToc159) to permit their direct comparison. We report the KM, Vmax, and Ea values for GTP hydrolysis and the Kd value for nucleotide binding for each protein. We demonstrate that GTP hydrolysis by psToc34 is stimulated by chloroplast transit peptides; however, this activity is not stimulated by homodimerization and is abolished by the R133A mutation. Furthermore, we show peptide stimulation of hydrolytic rates are not because of accelerated nucleotide exchange, indicating that transit peptides function as GTPase-activating proteins and not guanine nucleotide exchange factors in modulating the activity of psToc34. Finally, by using the psToc34 structure, we have developed molecular models for atToc33, atToc34, and atToc159G. By combining these models with the measured enzymatic properties of the Toc GTPases, we provide new insights of how the chloroplast protein import cycle may be regulated.

  8. A large population of small chloroplasts in tobacco leaf cells allows more effective chloroplast movement than a few enlarged chloroplasts.

    Science.gov (United States)

    Jeong, Won Joong; Park, Youn-Il; Suh, KyeHong; Raven, John A; Yoo, Ook Joon; Liu, Jang Ryol

    2002-05-01

    We generated transgenic tobacco (Nicotiana tabacum cv Xanthi) plants that contained only one to three enlarged chloroplasts per leaf mesophyll cell by introducing NtFtsZ1-2, a cDNA for plastid division. These plants were used to investigate the advantages of having a large population of small chloroplasts rather than a few enlarged chloroplasts in a leaf mesophyll cell. Despite the similarities in photosynthetic components and ultrastructure of photosynthetic machinery between wild-type and transgenic plants, the overall growth of transgenic plants under low- and high-light conditions was retarded. In wild-type plants, the chloroplasts moved toward the face position under low light and toward the profile position under high-light conditions. However, chloroplast rearrangement in transgenic plants in response to light conditions was not evident. In addition, transgenic plant leaves showed greatly diminished changes in leaf transmittance values under both light conditions, indicating that chloroplast rearrangement was severely retarded. Therefore, under low-light conditions the incomplete face position of the enlarged chloroplasts results in decreased absorbance of light energy. This, in turn, reduces plant growth. Under high-light conditions, the amount of absorbed light exceeds the photosynthetic utilization capacity due to the incomplete profile position of the enlarged chloroplasts, resulting in photodamage to the photosynthetic machinery, and decreased growth. The presence of a large number of small and/or rapidly moving chloroplasts in the cells of higher land plants permits more effective chloroplast phototaxis and, hence, allows more efficient utilization of low-incident photon flux densities. The photosynthetic apparatus is, consequently, protected from damage under high-incident photon flux densities.

  9. GENOTYPIC VARIATION IN CHLOROPHYLL FLUORESCENCE PARAMETERS, PHOTOSYNTHESIS AND GROWTH OF TOMATO GROWN AT LOW-TEMPERATURE AND LOW IRRADIANCE

    NARCIS (Netherlands)

    JANSSEN, LHJ; VANOEVEREN, JC; VANHASSELT, PR; KUIPER, PJC

    1995-01-01

    The genetic variation in low temperature sensitivity of eight tomato genotypes grown at suboptimal temperature (19 degrees C) and at low irradiance (140 mu mol m(-2) s(-1)) was assessed at the plant, chloroplast and thylakoid membrane levels. Temperature effects on the thylakoid membrane were determ

  10. Bioelectrochemistry II: Membrane Phenomena,

    Science.gov (United States)

    1984-12-11

    techniques for studying protein-lipid interactions and molecular movements in membranes. He discussed spin labels, fluorescent probes, NMR studies and recent...transduction in chloroplasts . Re reviewed the components and reactions at the two reaction centers In photosynthesis, and carefully correlated the structure...particularly useful for considering biological problems involving charge movement (e.g., ion transport, energy transduction, and electrical excitation

  11. Defects in the Expression of Chloroplast Proteins Leads to H2O2 Accumulation and Activation of Cyclic Electron Flow around Photosystem I

    Science.gov (United States)

    Strand, Deserah D.; Livingston, Aaron K.; Satoh-Cruz, Mio; Koepke, Tyson; Enlow, Heather M.; Fisher, Nicholas; Froehlich, John E.; Cruz, Jeffrey A.; Minhas, Deepika; Hixson, Kim K.; Kohzuma, Kaori; Lipton, Mary; Dhingra, Amit; Kramer, David M.

    2017-01-01

    We describe a new member of the class of mutants in Arabidopsis exhibiting high rates of cyclic electron flow around photosystem I (CEF), a light-driven process that produces ATP but not NADPH. High cyclic electron flow 2 (hcef2) shows strongly increased CEF activity through the NADPH dehydrogenase complex (NDH), accompanied by increases in thylakoid proton motive force (pmf), activation of the photoprotective qE response, and the accumulation of H2O2. Surprisingly, hcef2 was mapped to a non-sense mutation in the TADA1 (tRNA adenosine deaminase arginine) locus, coding for a plastid targeted tRNA editing enzyme required for efficient codon recognition. Comparison of protein content from representative thylakoid complexes, the cytochrome bf complex, and the ATP synthase, suggests that inefficient translation of hcef2 leads to compromised complex assembly or stability leading to alterations in stoichiometries of major thylakoid complexes as well as their constituent subunits. Altered subunit stoichiometries for photosystem I, ratios and properties of cytochrome bf hemes, and the decay kinetics of the flash-induced thylakoid electric field suggest that these defect lead to accumulation of H2O2 in hcef2, which we have previously shown leads to activation of NDH-related CEF. We observed similar increases in CEF, as well as increases in H2O2 accumulation, in other translation defective mutants. This suggests that loss of coordination in plastid protein levels lead to imbalances in photosynthetic energy balance that leads to an increase in CEF. These results taken together with a large body of previous observations, support a general model in which processes that lead to imbalances in chloroplast energetics result in the production of H2O2, which in turn activates CEF. This activation could be from either H2O2 acting as a redox signal, or by a secondary effect from H2O2 inducing a deficit in ATP. PMID:28133462

  12. Defects in the Expression of Chloroplast Proteins Leads to H2O2 Accumulation and Activation of Cyclic Electron Flow around Photosystem I

    Energy Technology Data Exchange (ETDEWEB)

    Strand, Deserah D.; Livingston, Aaron K.; Satoh-Cruz, Mio; Koepke, Tyson; Enlow, Heather M.; Fisher, Nicholas; Froehlich, John E.; Cruz, Jeffrey A.; Minhas, Deepika; Hixson, Kim K.; Kohzuma, Kaori; Lipton, Mary; Dhingra, Amit; Kramer, David M.

    2017-01-13

    We describe a new member of the class of mutants in Arabidopsis exhibiting high rates of cyclic electron flow around photosystem I (CEF), a light-driven process that produces ATP but not NADPH. High cyclic electron flow 2 (hcef2) shows strongly increased CEF activity through the NADPH dehydrogenase complex (NDH), accompanied by increases in thylakoid proton motive force (pmf), activation of the photoprotective qE response, and the accumulation of H2O2 . Surprisingly, hcef2 was mapped to a nonsense mutation in the TADA1 (tRNA adenosine deaminase arginine) locus, coding for a plastid targeted tRNA editing enzyme required for efficient codon recognition. Comparison of protein content from representative thylakoid complexes, the cytochrome bf complex and the ATP synthase, suggests that inefficient translation of hcef2 leads to compromised complex assembly or stability leading to alterations in stoichiometries of major thylakoid complexes as well as their constituent subunits. Altered subunit stoichiometries for photosystem I, ratios and properties of cytochrome bf hemes, and the decay kinetics of the flash induced thylakoid electric field suggest that these defect lead to accumulation of H2O2 in hcef2, which we have previously shown leads to activation of NDHrelated CEF. We observed similar increases in CEF and H2O2 accumulation in other translation defective mutants, suggesting that loss of coordination in plastid protein levels lead to imbalances in the photosynthetic energy balance that leads to increased CEF. These results, together with a large body of previous observations, support a general model in which processes that imbalances in chloroplast energetics result in the production of H2O2 , which activates CEF, either as a redox signal or by inducing deficits in ATP levels.

  13. Molecular basis of chloroplast photorelocation movement.

    Science.gov (United States)

    Kong, Sam-Geun; Wada, Masamitsu

    2016-03-01

    Chloroplast photorelocation movement is an essential physiological response for sessile plant survival and the optimization of photosynthetic ability. Simple but effective experiments on the physiological, cell biological and molecular genetic aspects have been widely used to investigate the signaling components of chloroplast photorelocation movement in Arabidopsis for the past few decades. Although recent knowledge on chloroplast photorelocation movement has led us to a deeper understanding of its physiological and molecular basis, the biochemical roles of the downstream factors remain largely unknown. In this review, we briefly summarize recent advances regarding chloroplast photorelocation movement and propose that a new high-resolution approach is necessary to investigate the molecular mechanism underlying actin-based chloroplast photorelocation movement.

  14. Effects of truncated mutants of the ε subunit of chloroplast ATP synthase on the fast phase of millisecond delayed light emission of chloroplast and its ATP synthesis ability

    Institute of Scientific and Technical Information of China (English)

    ZENG Xiaomei; SHI Xiaobing; SHEN Yungang

    2004-01-01

    The ε subunit of the chloroplast ATP synthase and the truncated ε mutants which lack some amino acid residues from the N-terminus or C-terminus were overexpressed in E. coli. When the ε subunit or the truncated ε proteins was added to the spinach chloroplast suspension, both the intensity of the fast phase of millisecond delayed light emission (ms-DLE) and the cyclic and noncyclic photophosphorylation activity of chloroplast were enhanced. With an increase in the number of residues deleted from the N-terminus, the enhancement effect of the N-terminal truncated proteins decreased gradually. For the C-terminal truncated proteins, the enhancement effect increased gradually with an increase in the number of residues deleted from the C-terminus. Besides, the ATP synthesis activity of ε-deficient membrane reconstituted with the ε subunit or the truncated ε proteins was compared. The ATP synthesis activity of reconstituted membrane with the N-terminal truncated proteins decreased gradually as the number of residues deleted from the N-terminus increased. For the C-terminal truncated proteins, the ATP synthesis activity of reconstituted membrane increased gradually with an increase in the number of residues deleted from the C-terminus, but was still lower than that of the wild type ε protein. These results suggested that: (a) the N-terminal domain of the ε subunit of the chloroplast ATP synthase could affect the ATP synthesis activity of ATP synthase by regulating the efficiency of blocking proton leakage of ε subunit; and (b) the C-terminal domain of the ε subunit of the chloroplast ATP synthase had a subtle function in modulating the ATP synthesis ability of ATP synthase.

  15. Role of chloroplasts and other plastids in ageing and death of plants and animals: a tale of Vishnu and Shiva.

    Science.gov (United States)

    van Doorn, Wouter G; Yoshimoto, Kohki

    2010-04-01

    Chloroplasts (chlorophyll-containing plastids) and other plastids are found in all plants and many animals. They are crucial to the survival of plants and most of the animals that harbour them. An example of a non-photosynthesizing plastid in animals is the apicoplast in the malaria-causing Plasmodium species, which is required for survival of the parasite. Many animals (such as sea slugs, sponges, reef corals, and clams) consume prey containing chloroplasts, or feed on algae. Some of these incorporate the chloroplasts from their food, or whole algal cells, into their own cells. Other species from these groups place algal cells between their own cells. Reef-building corals often lose their intracellular algae as a result of environmental changes, resulting in coral bleaching and death. The sensitivity of the chloroplast internal membranes to temperature stress is one of the reasons for coral death. Chloroplasts can also be a causal factor in the processes leading to whole-plant death, as the knockout of a gene encoding a chloroplast protein delayed the yellowing that proceeds death in tobacco plants. It is concluded that chloroplasts and other plastids are essential to individual survival in many species, including animals, and that they also play a role in triggering death in some plant and animal species.

  16. Copper and cadmium tolerance, uptake and effect on chloroplast ultrastructure. Studies on Salix purpurea and Phragmites australis

    Energy Technology Data Exchange (ETDEWEB)

    Hakmaoui, A.; Ater, A. [Abdelmalek Essaadi Univ., Tetouan (Morocco). Dept. of Biology; Boka, K. [Eoetvoes Lorand Univ., Budapest (Hungary). Dept. of Plant Anatomy; Baron, M. [Granada Univ. (Spain). Dept. of Biochemistry and Cell and Molecular Biology of Plants

    2007-05-15

    We have compared the effect of toxic Cu and Cd concentrations on growth, metal accumulation, and chloroplast ultrastructure of willow (Salix purpurea L.) and reed [Phragmites australis (Cav.) Trin. ex Steud.]. After a 10-day treatment, both species have tolerated to some extent the lowest concentration of both metals; however, plant growth was strongly reduced at the highest Cu and Cd concentrations. These plants could be described as Cu-tolerant at the lowest concentration tested, showing a higher tolerance index in reed than in willow; in contrast, willow exhibited higher tolerance against Cd. Both plants appeared to be moderate root accumulators of Cu and Cd. Ultrastructural studies revealed special features that can provide some protection against heavy metals stress, such as ferritin aggregates in the stroma. In addition, Cu and Cd induced distortion of thylakoids, reduction of grana stacks, as well as an increased number and size of plastoglobuli and peripheral vesicles. (orig.)

  17. The Chloroplast Import Receptor Toc90 Partially Restores the Accumulation of Toc159 Client Proteins in the Arabidopsis thaliana ppi2 Mutant

    Institute of Scientific and Technical Information of China (English)

    Sibylle Infanger; Sylvain Bischof; Andreas Hiltbrunner; Birgit Agne; Sacha Baginsky; Felix Kessler

    2011-01-01

    Successful import of hundreds of nucleus-encoded proteins is essential for chloroplast biogenesis. The import of cytosolic precursor proteins relies on the Toc- (translocon at the outer chloroplast membrane) and Tic- (translocon at the inner chloroplast membrane) complexes. In Arabidopsis thaliana,precursor recognition is mainly mediated by outer membrane receptors belonging to two gene families: Toc34/33 and Toc159/132/120/90. The role in import and precursor selectivity of these receptors has been intensively studied,but the function of Toc90 still remains unclear. Here,we report the ability of Toc90 to support the import of Toc159 client proteins. We show that the overexpression of Toc90 partially complements the albino knockout of Toc159 and restores photoautotrophic growth. Several lines of evidence including proteome profiling demonstrate the import and accumulation of proteins essential for chloroplast biogenesis and functionality.

  18. A twin arginine signal peptide and the pH gradient trigger reversible assembly of the thylakoid [Delta]pH/Tat translocase.

    Science.gov (United States)

    Mori, Hiroki; Cline, Kenneth

    2002-04-15

    The thylakoid DeltapH-dependent/Tat pathway is a novel system with the remarkable ability to transport tightly folded precursor proteins using a transmembrane DeltapH as the sole energy source. Three known components of the transport machinery exist in two distinct subcomplexes. A cpTatC-Hcf106 complex serves as precursor receptor and a Tha4 complex is required after precursor recognition. Here we report that Tha4 assembles with cpTatC-Hcf106 during the translocation step. Interactions among components were examined by chemical cross-linking of intact thylakoids followed by immunoprecipitation and immunoblotting. cpTatC and Hcf106 were consistently associated under all conditions tested. In contrast, Tha4 was only associated with cpTatC and Hcf106 in the presence of a functional precursor and the DeltapH. Interestingly, a synthetic signal peptide could replace intact precursor in triggering assembly. The association of all three components was transient and dissipated upon the completion of protein translocation. Such an assembly-disassembly cycle could explain how the DeltapH/Tat system can assemble translocases to accommodate folded proteins of varied size. It also explains in part how the system can exist in the membrane without compromising its ion and proton permeability barrier.

  19. AtPHT4;4 is a chloroplast-localized ascorbate transporter in Arabidopsis.

    Science.gov (United States)

    Miyaji, Takaaki; Kuromori, Takashi; Takeuchi, Yu; Yamaji, Naoki; Yokosho, Kengo; Shimazawa, Atsushi; Sugimoto, Eriko; Omote, Hiroshi; Ma, Jian Feng; Shinozaki, Kazuo; Moriyama, Yoshinori

    2015-01-05

    Ascorbate is an antioxidant and coenzyme for various metabolic reactions in vivo. In plant chloroplasts, high ascorbate levels are required to overcome photoinhibition caused by strong light. However, ascorbate is synthesized in the mitochondria and the molecular mechanisms underlying ascorbate transport into chloroplasts are unknown. Here we show that AtPHT4;4, a member of the phosphate transporter 4 family of Arabidopsis thaliana, functions as an ascorbate transporter. In vitro analysis shows that proteoliposomes containing the purified AtPHT4;4 protein exhibit membrane potential- and Cl(-)-dependent ascorbate uptake. The AtPHT4;4 protein is abundantly expressed in the chloroplast envelope membrane. Knockout of AtPHT4;4 results in decreased levels of the reduced form of ascorbate in the leaves and the heat dissipation process of excessive energy during photosynthesis is compromised. Taken together, these observations indicate that the AtPHT4;4 protein is an ascorbate transporter at the chloroplast envelope membrane, which may be required for tolerance to strong light stress.

  20. Rapid Inactivation of Chloroplastic Ascorbate Peroxidase is Responsible for Oxidative Modification to Rubisco in Tomato (Lycopersicon esculentum) under Cadmium Stress

    Institute of Scientific and Technical Information of China (English)

    Kai-Lang Liu; Lin Shen; Jia-Qi Wang; Ji-Ping Sheng

    2008-01-01

    To investigate the sensitive site of antioxidant systems in chloroplast under cadmium stress and its consequence on reactive oxygen species production and action, the sub-organellar localization of chloroplast superoxide dismutases (SOD,EC 1.15.1.1) and ascorbic peroxidase (APX, EC 1.11.1.11) isoenzymes and changes of enzymes activities under cadmium stress were investigated in tomato seedlings. Two APX isoforms, one thylakoid-bound and one stromal, were detected. Cd at 50 μM induced a moderate increase of SOD activities but a rapid inactivation of both APX isoenzymes. APX inactivation was mainly related to the decrease of ascorbate concentration, as supported by in vitro treatment of exogenous ascorbate and APX kinetic properties under Cd stress. H2O2 accumulation in chloroplast, as a consequence of APX inactivation,was associated with a 60% loss of Rubisco (EC 4.1.1.39) activity, which could be partially accounted for by a 10% loss of Rubisco content. Protein oxidation assay found that the Rubisco large subunit was the most prominent carbonylated protein; the level of carbonylated Rubisco large subunit increased fivefold after Cd exposure. Thiol groups in the Rubisco large subunit were oxidized, as indicated by non-reducing electrophoresis. Treating crude extract with H2O2 resulted in a similar pattern of protein oxidation and thiols oxidation with that observed in Cd-treated plants. Our study indicates that APXs in the chloroplast is a highly sensitive site of antioxidant systems under Cd stress, and the inactivation of APX could be mainly responsible for oxidative modification to Rubisco and subsequent decrease in its activity.

  1. Thylakoids promote release of the satiety hormone cholecystokinin while reducing insulin in healthy humans

    DEFF Research Database (Denmark)

    Köhnke, Rickard; Lindbo, Agnes; Larsson, Therese

    2009-01-01

    (CCK, leptin and ghrelin), insulin and blood metabolites (glucose and free fatty acids). RESULTS: The CCK level increased, in particular between the 120 min time-point and onwards, the ghrelin level was reduced at 120 min and leptin level increased at 360 min after intake of the thylakoid-enriched meal...

  2. An ATP synthase harboring an atypical γ-subunit is involved in ATP synthesis in tomato fruit chromoplasts

    DEFF Research Database (Denmark)

    Pateraki, Irini; Renato, Marta; Azcõn-Bieto, Joaquín;

    2013-01-01

    Chromoplasts are non-photosynthetic plastids specialized in the synthesis and accumulation of carotenoids. During fruit ripening, chloroplasts differentiate into photosynthetically inactive chromoplasts in a process characterized by the degradation of the thylakoid membranes, and by the active sy...

  3. Analysis of chloroplast movement and relocation in Arabidopsis.

    Science.gov (United States)

    Wada, Masamitsu; Kong, Sam-Geun

    2011-01-01

    Chloroplast photorelocation movement is essential for the sessile plant survival and plays a role for efficient photosynthesis and avoiding photodamage of chloroplasts. There are several ways to observe or detect chloroplast movement directly or indirectly. Here, techniques for the induction of chloroplast movement and how to detect the responses, as well as various points of attention and advice for the experiments, are described.

  4. Recent advances in understanding the molecular mechanism of chloroplast photorelocation movement.

    Science.gov (United States)

    Kong, Sam-Geun; Wada, Masamitsu

    2014-04-01

    Plants are photosynthetic organisms that have evolved unique systems to adapt fluctuating environmental light conditions. In addition to well-known movement responses such as phototropism, stomatal opening, and nastic leaf movements, chloroplast photorelocation movement is one of the essential cellular responses to optimize photosynthetic ability and avoid photodamage. For these adaptations, chloroplasts accumulate at the areas of cells illuminated with low light (called accumulation response), while they scatter from the area illuminated with strong light (called avoidance response). Plant-specific photoreceptors (phototropin, phytochrome, and/or neochrome) mediate these dynamic directional movements in response to incident light position and intensity. Several factors involved in the mechanisms underlying the processes from light perception to actin-based movements have also been identified through molecular genetic approach. This review aims to discuss recent findings in the field relating to how chloroplasts move at molecular levels. This article is part of a Special Issue entitled: Dynamic and ultrastructure of bioenergetic membranes and their components.

  5. Phycobiliprotein diffusion in chloroplasts of cryptophyte Rhodomonas CS24.

    Science.gov (United States)

    Mirkovic, Tihana; Wilk, Krystyna E; Curmi, Paul M G; Scholes, Gregory D

    2009-04-01

    Unicellular cryptophyte algae employ antenna proteins with phycobilin chromophores in their photosynthetic machinery. The mechanism of light harvesting in these organisms is significantly different than the energy funneling processes in phycobilisomes utilized by cyanobacteria and red algae. One of the most striking features of cryptophytes is the location of the water-soluble phycobiliproteins, which are contained within the intrathylakoid spaces and are not on the stromal side of the lamellae as in the red algae and cyanobacteria. Studies of mobility of phycobiliproteins at the lumenal side of the thylakoid membranes and how their diffusional behavior may influence the energy funneling steps in light harvesting are reported. Confocal microscopy and fluorescence recovery after photobleaching (FRAP) are used to measure the diffusion coefficient of phycoerythrin 545 (PE545), the primary light harvesting protein of Rhodomonas CS24, in vivo. It is concluded that the diffusion of PE545 in the lumen is inhibited, suggesting possible membrane association or aggregation as a potential source of mobility hindrance.

  6. Chloroplast structure of the Cryptophyceae. Evidence for phycobiliproteins within intrathylakoidal spaces.

    Science.gov (United States)

    Gantt, E; Edwards, M R; Provasoli, L

    1971-02-01

    Selective extraction and morphological evidence indicate that the phycobiliproteins in three Cryptophyceaen algae (Chroomonas, Rhodomonas, and Cryptomonas) are contained within intrathylakoidal spaces and are not on the stromal side of the lamellae as in the red and blue-green algae. Furthermore, no discrete phycobilisome-type aggregates have thus far been observed in the Cryptophyceae. Structurally, although not necessarily functionally, this is a radical difference. The width of the intrathylakoidal spaces can vary but is generally about 200-300 A. While the thylakoid membranes are usually closely aligned, grana-type fusions do not occur. In Chroomonas these membranes evidence an extensive periodic display with a spacing on the order of 140-160 A. This periodicity is restricted to the membranes and has not been observed in the electron-opaque intrathylakoidal matrix.

  7. Distinct responses of chloroplasts to blue and green laser microbeam irradiations in the centric diatom Pleurosira laevis.

    Science.gov (United States)

    Shihira-Ishikawa, Ikuko; Nakamura, Takanori; Higashi, Sho-ichi; Watanabe, Masakatsu

    2007-01-01

    The centric diatom Pleurosira laevis is a large unicellular alga, in which ca 200 chloroplasts migrate toward the nuclear cytoplasm through the transvacuolar cytoplasmic strands in response to blue-light irradiation and, on the contrary, toward the cortical cytoplasm in response to green-light irradiation. We analyzed these light-induced chloroplast migrations using a scanning laser microbeam provided by a confocal microscope for intracellular irradiation. Spot irradiation of a blue laser microbeam induced rapid assemblage of chroloplasts into the nuclear cytoplasm regardless of the spot position and spot number. On the other hand, one or two spots of green laser microbeam induced chloroplast accumulation at the spots, although increasing spot numbers suppressed chloroplast accumulation at each spot. In our experimental condition, ca 1 min of blue-light irradiation was sufficient to stimulate movement, whereas green-light irradiation required uninterrupted and longer irradiation time (ca 15 min). Chloroplast assemblage induced by blue-light required extracellular Ca2+, and was inhibited by Ca2+ channel antagonists. Furthermore, higher efficiencies of chloroplast migration were obtained when a single beam spot was fragmented and scattered over wider area of plasma membrane. These observations suggested that blue-light induced a response at the plasma membrane, which subsequently activated Ca2+ permeable channels. This sequence of physiological events is identical to what was previously observed with chloroplast movement in response to mechanical stimulation. Furthermore, experiments with the cytoskeleton-disrupting agents, colchicine and cytochalasin D, indicated that blue-light-induced chloroplast movement required microtubules whereas the green-light-induced response to beam spot required actin filaments.

  8. Kinetics of structural reorganizations in multilamellarphotosynthetic membranes monitored by small-angle neutronscattering

    DEFF Research Database (Denmark)

    Nagy, Gergely; Kovacs, Laszlo; Unnep, Renata;

    2013-01-01

    We demonstrate the power of time-resolved small-angle neutron scattering experiments for the investigation of the structure and structural reorganizations of multilamellar photosynthetic membranes. In addition to briefly summarizing our results on thylakoid membranes isolated from higher plants a...

  9. Reversible membrane reorganizations during photosynthesis in vivo: revealed by small-angle neutron scattering

    DEFF Research Database (Denmark)

    Nagy, Gergely; Posselt, Dorthe; Kovacs, Laszlo

    2011-01-01

    In the present study, we determined characteristic repeat distances of the photosynthetic membranes in living cyanobacterial and eukaryotic algal cells, and in intact thylakoid membranes isolated from higher plants with time-resolved small-angle neutron scattering. This non-invasive technique rev...

  10. [Effect of metalloproteins on the photochemical activity of chloroplasts treated with polyene antibiotics].

    Science.gov (United States)

    Mutuskin, A A; Makovkina, L E; Pshenova, K V; Vostroknutova, G N

    1977-04-01

    The effects of various metall-containing proteins (plastocyanin, plantacyanin, azurine and cytochromes of the f type) on the activity of photosystem I of chloroplasts, treated with polyene antibiotics, were studied. The inhibiting effect of the polyenes, surgumycin and philipin, was completely removed by an addition of copper-containing protein plastocyanin. No similar effect was exerted by other Cu-containing proteins--azurine and plantacyanin. The cytochromes of the f type isolated from the green algae chlorella, blue-green algae spiruline and aphanezomenone, having different electrophoretic properties, restored the activity of photosystem I of chloroplasts incubated with antibiotics in a different degree. Acid cytochrome f of chlorella restored the activity by 80--100%; less acid cytochrome f from spiruline-only by 50%. The least restoring effect was exerted by aphanezomenone cytochrome, which possesses some basic properties. The chloroplasts treatment with surgumycin did not affect the isolation of the terminal enzyme of the chloroplast electron-transporting chain of ferredoxin--NADP--reductase. Possible environment of plastocyanin in the chloroplast membrane and the mechanism of photosystem I restoration are discussed.

  11. Transfer of the cytochrome P450-dependent dhurrin pathway from Sorghum bicolor into Nicotiana tabacum chloroplasts for light-driven synthesis

    DEFF Research Database (Denmark)

    Gnanasekaran, Thiyagarajan; Karcher, Daniel; Nielsen, Agnieszka Janina Zygadlo;

    2016-01-01

    Plant chloroplasts are light-driven cell factories that have great potential to act as a chassis for metabolic engineering applications. Using plant chloroplasts, we demonstrate how photosynthetic reducing power can drive a metabolic pathway to synthesise a bio-active natural product....... For this purpose, we stably engineered the dhurrin pathway from Sorghum bicolor into the chloroplasts of Nicotiana tabacum (tobacco). Dhurrin is a cyanogenic glucoside and its synthesis from the amino acid tyrosine is catalysed by two membrane-bound cytochrome P450 enzymes (CYP79A1 and CYP71E1) and a soluble...... glucosyltransferase (UGT85B1), and is dependent on electron transfer from a P450 oxidoreductase. The entire pathway was introduced into the chloroplast by integrating CYP79A1, CYP71E1, and UGT85B1 into a neutral site of the N. tabacum chloroplast genome. The two P450s and the UGT85B1 were functional when expressed...

  12. Arabidopsis thaliana leaves with altered chloroplast numbers and chloroplast movement exhibit impaired adjustments to both low and high light

    OpenAIRE

    Königer, Martina; Delamaide, Joy A.; Marlow, Elizabeth D.; Harris, Gary C.

    2008-01-01

    The effects of chloroplast number and size on the capacity for blue light-dependent chloroplast movement, the ability to increase light absorption under low light, and the susceptibility to photoinhibition were investigated in Arabidopsis thaliana. Leaves of wild-type and chloroplast number mutants with mean chloroplast numbers ranging from 120 to two per mesophyll cell were analysed. Chloroplast movement was monitored as changes in light transmission through the leaves. Light transmission wa...

  13. A comparison of rice chloroplast genomes

    DEFF Research Database (Denmark)

    Tang, Jiabin; Xia, Hong'ai; Cao, Mengliang

    2004-01-01

    ), which are both parental varieties of the super-hybrid rice, LYP9. Based on the patterns of high sequence coverage, we partitioned chloroplast sequence variations into two classes, intravarietal and intersubspecific polymorphisms. Intravarietal polymorphisms refer to variations within 93-11 or PA64S...... to intersubspecific polymorphisms. In our study, we found that the intersubspecific variations of 93-11 (indica) and PA64S (japonica) chloroplast genomes consisted of 72 single nucleotide polymorphisms and 27 insertions or deletions. The intersubspecific polymorphism rates between 93-11 and PA64S were 0.......05% for single nucleotide polymorphisms and 0.02% for insertions or deletions, nearly 8 and 10 times lower than their respective nuclear genomes. Based on the total number of nucleotide substitutions between the two chloroplast genomes, we dated the divergence of indica and japonica chloroplast genomes...

  14. Nano-scale characterization of the dynamics of the chloroplast Toc translocon.

    Science.gov (United States)

    Reddick, L Evan; Chotewutmontri, Prakitchai; Crenshaw, Will; Dave, Ashita; Vaughn, Michael; Bruce, Barry D

    2008-01-01

    Translocons are macromolecular nano-scale machines that facilitate the selective translocation of proteins across membranes. Although common in function, different translocons have evolved diverse molecular mechanisms for protein translocation. Subcellular organelles of endosymbiotic origin such as the chloroplast and mitochondria had to evolve/acquire translocons capable of importing proteins whose genes were transferred to the host genome. These gene products are expressed on cytosolic ribosomes as precursor proteins and targeted back to the organelle by an N-terminal extension called the transit peptide or presequence. In chloroplasts the transit peptide is specifically recognized by the Translocon of the Outer Chloroplast membrane (Toc) which is composed of receptor GTPases that potentially function as gate-like switches, where GTP binding and hydrolysis somehow facilitate preprotein binding and translocation. Compared to other translocons, the dynamics of the Toc translocon are probably more complex and certainly less understood. We have developed biochemical/biophysical, imaging, and computational techniques to probe the dynamics of the Toc translocon at the nanoscale. In this chapter we provide detailed protocols for kinetic and binding analysis of precursor interactions in organeller, measurement of the activity and nucleotide binding of the Toc GTPases, native electrophoretic analysis of the assembly/organization of the Toc complex, visualization of the distribution and mobility of Toc apparatus on the surface of chloroplasts, and conclude with the identification and molecular modeling Toc75 POTRA domains. With these new methodologies we discuss future directions of the field.

  15. Evolution of the chloroplast division machinery

    Institute of Scientific and Technical Information of China (English)

    Hongbo GAO; Fuli GAO

    2011-01-01

    Chloroplasts are photosynthetic organelles derived from endosymbiotic cyanobacteria during evolution.Dramatic changes occurred during the process of the formation and evolution of chloroplasts,including the large-scale gene transfer from chloroplast to nucleus.However,there are still many essential characters remaining.For the chloroplast division machinery,FtsZ proteins,Ftn2,SulA and part of the division site positioning system- MinD and MinE are still conserved.New or at least partially new proteins,such as FtsZ family proteins FtsZl and ARC3,ARC6H,ARC5,PDV1,PDV2 and MCD1,were introduced for the division of chloroplasts during evolution.Some bacterial cell division proteins,such as FtsA,MreB,Ftn6,FtsW and Ftsl,probably lost their function or were gradually lost.Thus,the chloroplast division machinery is a dynamically evolving structure with both conservation and innovation.

  16. Dating the cyanobacterial ancestor of the chloroplast.

    Science.gov (United States)

    Falcón, Luisa I; Magallón, Susana; Castillo, Amanda

    2010-06-01

    Cyanobacteria have had a pivotal role in the history of life on Earth being the first organisms to perform oxygenic photosynthesis, which changed the atmospheric chemistry and allowed the evolution of aerobic Eukarya. Chloroplasts are the cellular organelles of photoautotrophic eukaryotes in which most portions of photosynthesis occur. Although the initial suggestion that cyanobacteria are the ancestors of chloroplasts was greeted with skepticism, the idea is now widely accepted. Here we attempt to resolve and date the cyanobacterial ancestry of the chloroplast using phylogenetic analysis and molecular clocks. We found that chloroplasts form a monophyletic lineage, are most closely related to subsection-I, N(2)-fixing unicellular cyanobacteria (Order Chroococcales), and heterocyst-forming Order Nostocales cyanobacteria are their sister group. Nostocales and Chroococcales appeared during the Paleoproterozoic and chloroplasts appeared in the mid-Proterozoic. The capability of N(2) fixation in cyanobacteria may have appeared only once during the late Archaean and early Proterozoic eons. Furthermore, we found that oxygen-evolving cyanobacteria could have appeared in the Archaean. Our results suggest that a free-living cyanobacterium with the capacity to store starch through oxygenic CO(2) fixation, and to fix atmospheric N(2), would be a very important intracellular acquisition, which, as can be recounted today from several lines of evidence, would have become the chloroplast by endosymbiosis.

  17. Comparison of the chloroplast peroxidase system in the chlorophyte Chlamydomonas reinhardtii, the bryophyte Physcomitrella patens, the lycophyte Selaginella moellendorffii and the seed plant Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Baier Margarete

    2010-06-01

    Full Text Available Abstract Background Oxygenic photosynthesis is accompanied by the formation of reactive oxygen species (ROS, which damage proteins, lipids, DNA and finally limit plant yield. The enzymes of the chloroplast antioxidant system are exclusively nuclear encoded. During evolution, plastid and mitochondrial genes were post-endosymbiotically transferred to the nucleus, adapted for eukaryotic gene expression and post-translational protein targeting and supplemented with genes of eukaryotic origin. Results Here, the genomes of the green alga Chlamydomonas reinhardtii, the moss Physcomitrella patens, the lycophyte Selaginella moellendorffii and the seed plant Arabidopsis thaliana were screened for ORFs encoding chloroplast peroxidases. The identified genes were compared for their amino acid sequence similarities and gene structures. Stromal and thylakoid-bound ascorbate peroxidases (APx share common splice sites demonstrating that they evolved from a common ancestral gene. In contrast to most cormophytes, our results predict that chloroplast APx activity is restricted to the stroma in Chlamydomonas and to thylakoids in Physcomitrella. The moss gene is of retrotransposonal origin. The exon-intron-structures of 2CP genes differ between chlorophytes and streptophytes indicating an independent evolution. According to amino acid sequence characteristics only the A-isoform of Chlamydomonas 2CP may be functionally equivalent to streptophyte 2CP, while the weakly expressed B- and C-isoforms show chlorophyte specific surfaces and amino acid sequence characteristics. The amino acid sequences of chloroplast PrxII are widely conserved between the investigated species. In the analyzed streptophytes, the genes are unspliced, but accumulated four introns in Chlamydomonas. A conserved splice site indicates also a common origin of chlorobiont PrxQ. The similarity of splice sites also demonstrates that streptophyte glutathione peroxidases (GPx are of common origin. Besides

  18. Analysis of gene sequences indicates that quantity not quality of chloroplast small HSPs improves thermotolerance in C4 and CAM plants.

    Science.gov (United States)

    Shakeel, Samina N; Ul Haq, Noor; Heckathorn, Scott; Luthe, D S

    2012-10-01

    Chloroplast-localized small heat-shock proteins (Cp-sHSP) protect Photosystem II and thylakoid membranes during heat and other stresses, and Cp-sHSP production levels are related to plant thermotolerance. However, to date, a paucity of Cp-sHSP sequences from C4 or CAM species, or from other extremely heat-tolerant species, has precluded an examination to determine if Cp-sHSP genes or proteins might differ among plants with photosynthetic pathways or between heat-sensitive and heat-tolerant species. To investigate this, we isolated and characterized novel Cp-sHSP genes in four plant species: two moderately heat-tolerant C4 species, Spartina alterniflora (monocot) and Amaranthus retroflexus (eudicot), and two very heat-tolerant CAM species, Agave americana (monocot) and Ferocactus wislizenii (eudicot) (respective genes: SasHSP27.12, ArsHSP26.43, AasHSP26.85 and FwsHSP27.52) by PCR-based genome walking and cDNA RACE. Analysis of these Cp-sHSPs has confirmed the presence of conserved domains common to previously examined species. As expected, the transit peptide was found to be the most variable part of these proteins. Promoter analysis of these genes revealed differences in CAM versus C3 and C4 species that were independent of a general difference between monocots and eudicots observed for the entire protein. Heat-induced gene and protein expression indicated that Cp-sHSP protein levels were correlated with thermotolerance of photosynthetic electron transport, and that in most cases protein and transcript levels were correlated. Thus, available evidence indicates little variation in the amino acid sequence of Cp-sHSP mature proteins between heat-sensitive and -tolerant species, but that variation in Cp-sHSP protein production is related to heat tolerance or photosynthetic pathway (CAM vs. C3 and C4) and is driven by promoter differences. Key message We isolated and characterized four novel Cp-sHSP genes with promoters from wild plants, analysis has shown qualitative

  19. The effect of metal chelators on the production of hydroxyl radicals in thylakoids.

    Science.gov (United States)

    Snyrychová, Iva; Pospísil, Pavel; Naus, Jan

    2006-06-01

    The effect of metal chelators (EDTA, DTPA and Desferal) on the metal-catalyzed decomposition of hydrogen peroxide was studied using EPR spin-trapping spectroscopy. The formation of hydroxyl radicals (OH*) in both chemical (Fenton reaction) and biological (thylakoids) systems was stimulated by EDTA. DTPA promoted the generation of OH* in the presence of strong reducing agents, whereas in their absence it acted as an antioxidant. Desferal suppressed OH* production even in the presence of reductants. In our study, we have shown that metal chelators can both stimulate and suppress the formation of OH*, depending on the experimental conditions. In illuminated thylakoids we have observed prooxidant effect of EDTA and DTPA, possibly due to their reduction by some component of the electron transport chain. According to our results, metal chelators should not be used as antioxidants without prior testing of their effect in given samples.

  20. Chloroplast anchoring: its implications for the regulation of intracellular chloroplast distribution.

    Science.gov (United States)

    Takagi, Shingo; Takamatsu, Hideyasu; Sakurai-Ozato, Nami

    2009-01-01

    The intracellular distribution of organelles plays a pivotal role in the maintenance and adaptation of a wide spectrum of cellular activities in plants. Chloroplasts are a special type of organelle able to photosynthesize, capturing light energy to fix atmospheric CO2. Consequently, the intracellular positioning of chloroplasts is crucial for plant growth and development. Knowledge of the photoreceptors and cellular apparatus responsible for chloroplast movement has gradually accumulated over time, yet recent advances have allowed improved understanding. In this article, several aspects of research progress into the mechanisms for maintaining the specific intracellular distribution patterns of chloroplasts, namely, chloroplast anchoring, are summarized, together with a brief consideration of the future prospects of this subject. Our discussion covers developmental, physiological, ecophysiological, and recent cell biological research areas.

  1. Complete chloroplast genome of Sedum sarmentosum and chloroplast genome evolution in Saxifragales.

    Directory of Open Access Journals (Sweden)

    Wenpan Dong

    Full Text Available Comparative chloroplast genome analyses are mostly carried out at lower taxonomic levels, such as the family and genus levels. At higher taxonomic levels, chloroplast genomes are generally used to reconstruct phylogenies. However, little attention has been paid to chloroplast genome evolution within orders. Here, we present the chloroplast genome of Sedum sarmentosum and take advantage of several available (or elucidated chloroplast genomes to examine the evolution of chloroplast genomes in Saxifragales. The chloroplast genome of S. sarmentosum is 150,448 bp long and includes 82,212 bp of a large single-copy (LSC region, 16.670 bp of a small single-copy (SSC region, and a pair of 25,783 bp sequences of inverted repeats (IRs.The genome contains 131 unique genes, 18 of which are duplicated within the IRs. Based on a comparative analysis of chloroplast genomes from four representative Saxifragales families, we observed two gene losses and two pseudogenes in Paeonia obovata, and the loss of an intron was detected in the rps16 gene of Penthorum chinense. Comparisons among the 72 common protein-coding genes confirmed that the chloroplast genomes of S. sarmentosum and Paeonia obovata exhibit accelerated sequence evolution. Furthermore, a strong correlation was observed between the rates of genome evolution and genome size. The detected genome size variations are predominantly caused by the length of intergenic spacers, rather than losses of genes and introns, gene pseudogenization or IR expansion or contraction. The genome sizes of these species are negatively correlated with nucleotide substitution rates. Species with shorter duration of the life cycle tend to exhibit shorter chloroplast genomes than those with longer life cycles.

  2. Integrated role of ROS and Ca(+2) in blue light-induced chloroplast avoidance movement in leaves of Hydrilla verticillata (L.f.) Royle.

    Science.gov (United States)

    Majumdar, Arkajo; Kar, Rup Kumar

    2016-11-01

    Directional chloroplast photorelocation is a major physio-biochemical mechanism that allows these organelles to realign themselves intracellularly in response to the intensity of the incident light as an adaptive response. Signaling processes involved in blue light (BL)-dependent chloroplast movements were investigated in Hydrilla verticillata (L.f.) Royle leaves. Treatments with antagonists of actin filaments [2,3,5-triiodobenzoic acid (TIBA)] and microtubules (oryzalin) revealed that actin filaments, but not microtubules, play a pivotal role in chloroplast movement. Involvement of reactive oxygen species (ROS) in controlling chloroplast avoidance movement has been demonstrated, as exogenous H2O2 not only accelerated chloroplast avoidance but also could induce chloroplast avoidance even in weak blue light (WBL). Further support came from experiments with different ROS scavengers, i.e., dimethylthiourea (DMTU), KI, and CuCl2, which inhibited chloroplast avoidance, and from ROS localization using specific stains. Such avoidance was also partially inhibited by ZnCl2, an inhibitor of NADPH oxidase (NOX) as well as 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), a photosynthetic electron transport chain (ETC) inhibitor at PS II. However, methyl viologen (MV), a PS I ETC inhibitor, rather accelerated avoidance response. Exogenous calcium (Ca(+2)) induced avoidance even in WBL while inhibited chloroplast accumulation partially. On the other hand, chloroplast movements (both accumulation and avoidance) were blocked by Ca(+2) antagonists, La(3+) (inhibitor of plasma membrane Ca(+2) channel) and ethylene glycol-bis(2-aminoethylether)-N,N,N',N'-tetraacetic acid (EGTA, Ca(+2) chelator) while LiCl that affects Ca(+2) release from endosomal compartments did not show any effect. A model on integrated role of ROS and Ca(+2) (influx from apolastic space) in actin-mediated chloroplast avoidance has been proposed.

  3. Multiple complexes of nitrogen assimilatory enzymes in spinach chloroplasts: possible mechanisms for the regulation of enzyme function.

    Directory of Open Access Journals (Sweden)

    Yoko Kimata-Ariga

    Full Text Available Assimilation of nitrogen is an essential biological process for plant growth and productivity. Here we show that three chloroplast enzymes involved in nitrogen assimilation, glutamate synthase (GOGAT, nitrite reductase (NiR and glutamine synthetase (GS, separately assemble into distinct protein complexes in spinach chloroplasts, as analyzed by western blots under blue native electrophoresis (BN-PAGE. GOGAT and NiR were present not only as monomers, but also as novel complexes with a discrete size (730 kDa and multiple sizes (>120 kDa, respectively, in the stromal fraction of chloroplasts. These complexes showed the same mobility as each monomer on two-dimensional (2D SDS-PAGE after BN-PAGE. The 730 kDa complex containing GOGAT dissociated into monomers, and multiple complexes of NiR reversibly converted into monomers, in response to the changes in the pH of the stromal solvent. On the other hand, the bands detected by anti-GS antibody were present not only in stroma as a conventional decameric holoenzyme complex of 420 kDa, but also in thylakoids as a novel complex of 560 kDa. The polypeptide in the 560 kDa complex showed slower mobility than that of the 420 kDa complex on the 2D SDS-PAGE, implying the assembly of distinct GS isoforms or a post-translational modification of the same GS protein. The function of these multiple complexes was evaluated by in-gel GS activity under native conditions and by the binding ability of NiR and GOGAT with their physiological electron donor, ferredoxin. The results indicate that these multiplicities in size and localization of the three nitrogen assimilatory enzymes may be involved in the physiological regulation of their enzyme function, in a similar way as recently described cases of carbon assimilatory enzymes.

  4. PPR8522 encodes a chloroplast-targeted pentatricopeptide repeat protein necessary for maize embryogenesis and vegetative development.

    Science.gov (United States)

    Sosso, Davide; Canut, Matthieu; Gendrot, Ghislaine; Dedieu, Annick; Chambrier, Pierre; Barkan, Alice; Consonni, Gabriella; Rogowsky, Peter M

    2012-10-01

    The pentatricopeptide repeat (PPR) domain is an RNA binding domain allowing members of the PPR superfamily to participate in post-transcriptional processing of organellar RNA. Loss of PPR8522 from maize (Zea mays) confers an embryo-specific (emb) phenotype. The emb8522 mutation was isolated in an active Mutator (Mu) population and co-segregation analysis revealed that it was tightly linked to a MuDR insertion in the first exon of PPR8522. Independent evidence that disruption of PPR8522 caused the emb phenotype was provided by fine mapping to a region of 116kb containing no other gene than PPR8522 and complementation of the emb8522 mutant by a PPR8522 cDNA. The deduced PPR8522 amino acid sequence of 832 amino acids contains 10 PPR repeats and a chloroplast target peptide, the function of which was experimentally demonstrated by transient expression in Nicotiana benthamiana. Whereas mutant endosperm is apparently normal, mutant embryos deviate from normal development as early as 3 days after pollination, are reduced in size, exhibit more or less severe morphological aberrations depending on the genetic background, and generally do not germinate. The emb8522 mutation is the first to associate the loss of a PPR gene with an embryo-lethal phenotype in maize. Analyses of mutant plantlets generated by embryo-rescue experiments indicate that emb8522 also affects vegetative plant growth and chloroplast development. The loss of chloroplast transcription dependent on plastid-encoded RNA polymerase is the likely cause for the lack of an organized thylakoid network and an albino, seedling-lethal phenotype.

  5. The novel cytochrome c6 of chloroplasts: a case of evolutionary bricolage?

    Science.gov (United States)

    Howe, Christopher J; Schlarb-Ridley, Beatrix G; Wastl, Juergen; Purton, Saul; Bendall, Derek S

    2006-01-01

    Cytochrome c6 has long been known as a redox carrier of the thylakoid lumen of cyanobacteria and some eukaryotic algae that can substitute for plastocyanin in electron transfer. Until recently, it was widely accepted that land plants lack a cytochrome c6. However, a homologue of the protein has now been identified in several plant species together with an additional isoform in the green alga Chlamydomonas reinhardtii. This form of the protein, designated cytochrome c6A, differs from the 'conventional' cytochrome c6 in possessing a conserved insertion of 12 amino acids that includes two absolutely conserved cysteine residues. There are conflicting reports of whether cytochrome c6A can substitute for plastocyanin in photosynthetic electron transfer. The evidence for and against this is reviewed and the likely evolutionary history of cytochrome c6A is discussed. It is suggested that it has been converted from a primary role in electron transfer to one in regulation within the chloroplast, and is an example of evolutionary 'bricolage'.

  6. Non-invasive, whole-plant imaging of chloroplast movement and chlorophyll fluorescence reveals photosynthetic phenotypes independent of chloroplast photorelocation defects in chloroplast division mutants.

    Science.gov (United States)

    Dutta, Siddhartha; Cruz, Jeffrey A; Jiao, Yuhua; Chen, Jin; Kramer, David M; Osteryoung, Katherine W

    2015-10-01

    Leaf chloroplast movement is thought to optimize light capture and to minimize photodamage. To better understand the impact of chloroplast movement on photosynthesis, we developed a technique based on the imaging of reflectance from leaf surfaces that enables continuous, high-sensitivity, non-invasive measurements of chloroplast movement in multiple intact plants under white actinic light. We validated the method by measuring photorelocation responses in Arabidopsis chloroplast division mutants with drastically enlarged chloroplasts, and in phototropin mutants with impaired photorelocation but normal chloroplast morphology, under different light regimes. Additionally, we expanded our platform to permit simultaneous image-based measurements of chlorophyll fluorescence and chloroplast movement. We show that chloroplast division mutants with enlarged, less-mobile chloroplasts exhibit greater photosystem II photodamage than is observed in the wild type, particularly under fluctuating high levels of light. Comparison between division mutants and the severe photorelocation mutant phot1-5 phot2-1 showed that these effects are not entirely attributable to diminished photorelocation responses, as previously hypothesized, implying that altered chloroplast morphology affects other photosynthetic processes. Our dual-imaging platform also allowed us to develop a straightforward approach to correct non-photochemical quenching (NPQ) calculations for interference from chloroplast movement. This correction method should be generally useful when fluorescence and reflectance are measured in the same experiments. The corrected data indicate that the energy-dependent (qE) and photoinhibitory (qI) components of NPQ contribute differentially to the NPQ phenotypes of the chloroplast division and photorelocation mutants. This imaging technology thus provides a platform for analyzing the contributions of chloroplast movement, chloroplast morphology and other phenotypic attributes to the

  7. Chloroplasts in seeds and dark-grown seedlings of lotus.

    Science.gov (United States)

    Ushimaru, Takashi; Hasegawa, Takahiro; Amano, Toyoki; Katayama, Masao; Tanaka, Shigeyasu; Tsuji, Hideo

    2003-03-01

    In most higher plants, mature dry seeds have no chloroplasts but etioplasts. Here we show that in a hydrophyte, lotus (Nelumbo nucifera), young chloroplasts already exist in shoots of mature dry seeds and that they give rise to mature chloroplasts during germination, even in darkness. These shoots contain chlorophyll and chlorophyll-binding proteins CP1 and LHCP. The unique features of chloroplast formation in N. nucifera suggest a unique adaptive strategy for seedling development correlated with the plant's habitat.

  8. Chup1 - a chloroplast movement protein and its interactions

    OpenAIRE

    Schmidt von Braun, Serena

    2008-01-01

    The molecular mechanisms of light dependent chloroplast movement could for a long time not be unravelled. But the recent discovery of a mutant deficient in chloroplast movement sparked new impulses in the field. This study investigates the molecular mechanisms of chloroplast movement based on the protein Chup1 and the interactions of Chup1 and cytoskeletal effectors. It is demonstrated that Chup1 is exclusively and directly targeted to the chloroplast surface in an N-terminus dependent manner...

  9. TGD4 involved in endoplasmic reticulum-to-chloroplast lipid trafficking is a phosphatidic acid binding protein

    Energy Technology Data Exchange (ETDEWEB)

    Wang Z.; Xu C.; Benning, C.

    2012-05-01

    The synthesis of galactoglycerolipids, which are prevalent in photosynthetic membranes, involves enzymes at the endoplasmic reticulum (ER) and the chloroplast envelope membranes. Genetic analysis of trigalactosyldiacylglycerol (TGD) proteins in Arabidopsis has demonstrated their role in polar lipid transfer from the ER to the chloroplast. The TGD1, 2, and 3 proteins resemble components of a bacterial-type ATP-binding cassette (ABC) transporter, with TGD1 representing the permease, TGD2 the substrate binding protein, and TGD3 the ATPase. However, the function of the TGD4 protein in this process is less clear and its location in plant cells remains to be firmly determined. The predicted C-terminal {beta}-barrel structure of TGD4 is weakly similar to proteins of the outer cell membrane of Gram-negative bacteria. Here, we show that, like TGD2, the TGD4 protein when fused to DsRED specifically binds phosphatidic acid (PtdOH). As previously shown for tgd1 mutants, tgd4 mutants have elevated PtdOH content, probably in extraplastidic membranes. Using highly purified and specific antibodies to probe different cell fractions, we demonstrated that the TGD4 protein was present in the outer envelope membrane of chloroplasts, where it appeared to be deeply buried within the membrane except for the N-terminus, which was found to be exposed to the cytosol. It is proposed that TGD4 is either directly involved in the transfer of polar lipids, possibly PtdOH, from the ER to the outer chloroplast envelope membrane or in the transfer of PtdOH through the outer envelope membrane.

  10. Utilization of complete chloroplast genomes for phylogenetic studies

    NARCIS (Netherlands)

    Ramlee, Shairul Izan Binti

    2016-01-01

    Chloroplast DNA sequence polymorphisms are a primary source of data in many plant phylogenetic studies. The chloroplast genome is relatively conserved in its evolution making it an ideal molecule to retain phylogenetic signals. The chloroplast genome is also largely, but not completely, free from ot

  11. Chloroplasts can move in any direction to avoid strong light.

    Science.gov (United States)

    Tsuboi, Hidenori; Wada, Masamitsu

    2011-01-01

    Chloroplasts migrate in response to different light intensities. Under weak light, chloroplasts gather at an illuminated area to maximize light absorption and photosynthesis rates (the accumulation response). In contrast, chloroplasts escape from strong light to avoid photodamage (the avoidance response). Photoreceptors involved in these phenomena have been identified in Arabidopsis thaliana and Adiantum capillus-veneris. Chloroplast behavior has been studied in detail during the accumulation response, but not for the avoidance response. Hence, we analyzed the chloroplast avoidance response in detail using dark-adapted Adiantum capillus-veneris gametophyte cells and partial cell irradiation with a microbeam of blue light. Chloroplasts escaped from an irradiated spot. Both duration of this response and the distance of the migrated chloroplasts were proportional to the total fluence irradiated. The speed of movement during the avoidance response was dependent on the fluence rate, but the speed of the accumulation response towards the microbeam from cell periphery was constant irrespective of fluence rate. When a chloroplast was only partially irradiated with a strong microbeam, it moved away towards the non-irradiated region within a few minutes. During this avoidance response two additional microbeam irradiations were applied to different locus of the same chloroplast. Under these conditions the chloroplast changed the moving direction after a lag time of a few minutes without rolling. Taken together, these findings indicate that chloroplasts can move in any direction and never have an intrinsic polarity. Similar phenomenon was observed in chloroplasts of Arabidopsis thaliana palisade cells.

  12. Chloroplast signaling within, between and beyond cells.

    Directory of Open Access Journals (Sweden)

    Krzysztof eBobik

    2015-10-01

    Full Text Available The most conspicuous function of the plastid is oxygenic photosynthesis of chloroplasts, yet plastids are super-factories that produce a plethora of compounds that are indispensable for proper plant physiology and development. Given their origins as free-living prokaryotes, it is not surprising that the plastid possesses its own genome whose expression is essential to plastid function. This semi-autonomous character of plastids requires the existence of sophisticated regulatory mechanisms that provide reliable communication between them and other cellular compartments. Such intracellular signaling is necessary for coordinating whole-cell responses to constantly varying environmental cues and cellular metabolic needs. This is achieved by plastids acting as receivers and transmitters of specific signals that coordinate expression of the nuclear and plastid genomes according to particular needs. In this review we will consider the so-called retrograde signaling occurring between plastids and nucleus, and between plastids and other organelles. Another important role of the plastid we will discuss is the involvement of plastid signaling in biotic and abiotic stress that, in addition to influencing retrograde signaling has direct effects on several cellular compartments including the cell wall. We will also review recent evidence pointing to an intriguing function of chloroplasts in regulating intercellular symplasmic transport. Finally, we consider an intriguing yet neglected aspect of plant biology, chloroplast signaling from the perspective of the entire plant. Thus, accumulating evidence highlights that chloroplasts, with their complex signaling pathways, provide a mechanism for exquisite regulation of plant development, metabolism and responses to the environment. As chloroplast processes are targeted for engineering for improved productivity the effect of such modifications on chloroplast signaling will have to be carefully considered in order

  13. The Shibata Shift and the Transformation of Etioplasts to Chloroplasts in Wheat with Clomazone (FMC 57020) and Amiprophos-Methyl (Tokunol M) 1

    Science.gov (United States)

    Artus, Nancy N.; Ryberg, Margareta; Lindsten, Agneta; Ryberg, Hans; Sundqvist, Christer

    1992-01-01

    The Shibata shift is a change in the absorption maximum of chlorophyllide from 684 to 672 nanometers that occurs within approximately 0.5 hour of phototransformation of protochlorophyllide to chlorophyllide. Two compounds, clomazone and amiprophos-methyl, which previously have been shown to inhibit the Shibata shift in vivo, were used to look for correlations between the Shibata shift and other processes that occur during etioplast to chloroplast transformation. Leaf sections from 6-day-old etiolated wheat seedlings (Triticum aestivum L. cv Walde) were treated with 0.5 millimolar clomazone or 0.1 millimolar amiprophos-methyl in darkness. In addition to the Shibata shift, the esterification of chlorophyllide to chlorophyll and the relocation of protochlorophyllide reductase from the prolamellar bodies to the developing thylakoids were inhibited by these treatments. Prolamellar body transformation did not appear to be affected by amiprophos-methyl and was only slightly affected by clomazone. The results indicate that: (a) there is a strong correlation between the occurrence of the Shibata shift and esterification activity; (b) transformation of the prolamellar bodies does not depend on the Shibata shift; and (c) the occurrence of the Shibata shift may be a prerequisite to the relocation of protochlorophyllide reductase from prolamellar bodies to thylakoids. ImagesFigure 2Figure 3Figure 4Figure 5 PMID:16668622

  14. The Shibata Shift and the Transformation of Etioplasts to Chloroplasts in Wheat with Clomazone (FMC 57020) and Amiprophos-Methyl (Tokunol M).

    Science.gov (United States)

    Artus, N N; Ryberg, M; Lindsten, A; Ryberg, H; Sundqvist, C

    1992-01-01

    The Shibata shift is a change in the absorption maximum of chlorophyllide from 684 to 672 nanometers that occurs within approximately 0.5 hour of phototransformation of protochlorophyllide to chlorophyllide. Two compounds, clomazone and amiprophos-methyl, which previously have been shown to inhibit the Shibata shift in vivo, were used to look for correlations between the Shibata shift and other processes that occur during etioplast to chloroplast transformation. Leaf sections from 6-day-old etiolated wheat seedlings (Triticum aestivum L. cv Walde) were treated with 0.5 millimolar clomazone or 0.1 millimolar amiprophos-methyl in darkness. In addition to the Shibata shift, the esterification of chlorophyllide to chlorophyll and the relocation of protochlorophyllide reductase from the prolamellar bodies to the developing thylakoids were inhibited by these treatments. Prolamellar body transformation did not appear to be affected by amiprophos-methyl and was only slightly affected by clomazone. The results indicate that: (a) there is a strong correlation between the occurrence of the Shibata shift and esterification activity; (b) transformation of the prolamellar bodies does not depend on the Shibata shift; and (c) the occurrence of the Shibata shift may be a prerequisite to the relocation of protochlorophyllide reductase from prolamellar bodies to thylakoids.

  15. A Putative Chloroplast-Localized Ca(2+)/H(+) Antiporter CCHA1 Is Involved in Calcium and pH Homeostasis and Required for PSII Function in Arabidopsis.

    Science.gov (United States)

    Wang, Chao; Xu, Weitao; Jin, Honglei; Zhang, Taijie; Lai, Jianbin; Zhou, Xuan; Zhang, Shengchun; Liu, Shengjie; Duan, Xuewu; Wang, Hongbin; Peng, Changlian; Yang, Chengwei

    2016-08-01

    Calcium is important for chloroplast, not only in its photosynthetic but also nonphotosynthetic functions. Multiple Ca(2+)/H(+) transporters and channels have been described and studied in the plasma membrane and organelle membranes of plant cells; however, the molecular identity and physiological roles of chloroplast Ca(2+)/H(+) antiporters have remained unknown. Here we report the identification and characterization of a member of the UPF0016 family, CCHA1 (a chloroplast-localized potential Ca(2+)/H(+) antiporter), in Arabidopsis thaliana. We observed that the ccha1 mutant plants developed pale green leaves and showed severely stunted growth along with impaired photosystem II (PSII) function. CCHA1 localizes to the chloroplasts, and the levels of the PSII core subunits and the oxygen-evolving complex were significantly decreased in the ccha1 mutants compared with the wild type. In high Ca(2+) concentrations, Arabidopsis CCHA1 partially rescued the growth defect of yeast gdt1Δ null mutant, which is defective in a Ca(2+)/H(+) antiporter. The ccha1 mutant plants also showed significant sensitivity to high concentrations of CaCl2 and MnCl2, as well as variation in pH. Taken these results together, we propose that CCHA1 might encode a putative chloroplast-localized Ca(2+)/H(+) antiporter with critical functions in the regulation of PSII and in chloroplast Ca(2+) and pH homeostasis in Arabidopsis.

  16. Processing of the 5'-UTR and existence of protein factors that regulate translation of tobacco chloroplast psbN mRNA.

    Science.gov (United States)

    Kuroda, Hiroshi; Sugiura, Masahiro

    2014-12-01

    The chloroplast psbB operon includes five genes encoding photosystem II and cytochrome b 6 /f complex components. The psbN gene is located on the opposite strand. PsbN is localized in the thylakoid and is present even in the dark, although its level increases upon illumination and then decreases. However, the translation mechanism of the psbN mRNA remains unclear. Using an in vitro translation system from tobacco chloroplasts and a green fluorescent protein as a reporter protein, we show that translation occurs from a tobacco primary psbN 5'-UTR of 47 nucleotides (nt). Unlike many other chloroplast 5'-UTRs, the psbN 5'-UTR has two processing sites, at -39 and -24 upstream from the initiation site. Processing at -39 enhanced the translation rate fivefold. In contrast, processing at -24 did not affect the translation rate. These observations suggest that the two distinct processing events regulate, at least in part, the level of PsbN during development. The psbN 5'-UTR has no Shine-Dalgarno (SD)-like sequence. In vitro translation assays with excess amounts of the psbN 5'-UTR or with deleted psbN 5'-UTR sequences demonstrated that protein factors are required for translation and that their binding site is an 18 nt sequence in the 5'-UTR. Mobility shift assays using 10 other chloroplast 5'-UTRs suggested that common or similar proteins are involved in translation of a set of mRNAs lacking SD-like sequences.

  17. Velocity of chloroplast avoidance movement is fluence rate dependent.

    Science.gov (United States)

    Kagawa, Takatoshi; Wada, Masamitsu

    2004-06-01

    In Arabidopsis leaves, chloroplast movement is fluence rate dependent. At optimal, lower light fluences, chloroplasts accumulate at the cell surface to maximize photosynthetic potential. Under high fluence rates, chloroplasts avoid incident light to escape photodamage. In this paper, we examine the phenomenon of chloroplast avoidance movement in greater detail and demonstrate a proportional relationship between fluence rate and the velocity of chloroplast avoidance. In addition we show that the amount of light-activated phototropin2, the photoreceptor for the avoidance response, likely plays a role in this phenomenon, as heterozygous mutant plants show a reduced avoidance velocity compared to that of homozygous wild type plants.

  18. Chloroplast replication and growth in tobacco

    NARCIS (Netherlands)

    Verbeek-Boasson, Rosalinda

    1969-01-01

    SUMMARY AND CONCLUSIONS 1. The greening and the growth of chloroplasts as induced by light has been investigated in leaf discs from etiolated tobacco leaves in sterile culture. 2.On a medium containing salts after Murashige and Skoog plus sucrose, chlorophyll synthesis proceeds very slowly during th

  19. The potato mop-top virus TGB2 protein and viral RNA associate with chloroplasts and viral infection induces inclusions in the plastids

    Directory of Open Access Journals (Sweden)

    Graham H Cowan

    2012-12-01

    Full Text Available The potato mop-top virus (PMTV triple gene block 2 (TGB2 movement protein fused to monomeric red fluorescent protein (mRFP-TGB2 was expressed under the control of the PMTV subgenomic promoter from a PMTV vector. The subcellular localisations and interactions of mRFP-TGB2 were investigated using confocal imaging (CLSM and biochemical analysis. The results revealed associations with membranes of the endoplasmic reticulum, mobile granules, small round structures (1-2 µm in diameter and chloroplasts. Expression of mRFP-TGB2 in epidermal cells enabled cell-to-cell movement of a TGB2 defective PMTV reporter clone, indicating that the mRFP-TGB2 fusion protein was functional and required for cell-to-cell movement. Protein-lipid interaction assays revealed an association between TGB2 and lipids present in chloroplasts, consistent with microscopical observations where the plastid envelope was labelled later in infection. To further investigate the association of PMTV infection with chloroplasts, ultrastructural studies of thin sections of PMTV-infected potato and Nicotiana benthamiana leaves by electron microscopy revealed abnormal chloroplasts with cytoplasmic inclusions and terminal projections. Viral coat protein, genomic RNA and fluorescently-labelled TGB2 were detected in plastid preparations isolated from the infected leaves, and viral RNA was localised to chloroplasts in infected tissues. The results reveal a novel association of TGB2 and vRNA with chloroplasts, and suggest viral replication is associated with chloroplast membranes, and that TGB2 plays a novel role in targeting the virus to chloroplasts.

  20. Regulation of Chloroplast Protein Import by the Ubiquitin E3 Ligase SP1 Is Important for Stress Tolerance in Plants.

    Science.gov (United States)

    Ling, Qihua; Jarvis, Paul

    2015-10-05

    Chloroplasts are the organelles responsible for photosynthesis in plants [1, 2]. The chloroplast proteome comprises ∼3,000 different proteins, including components of the photosynthetic apparatus, which are highly abundant. Most chloroplast proteins are nucleus-encoded and imported following synthesis in the cytosol. Such import is mediated by multiprotein complexes in the envelope membranes that surround each organelle [3, 4]. The translocon at the outer envelope membrane of chloroplasts (TOC) mediates client protein recognition and early stages of import. The TOC apparatus is regulated by the ubiquitin-proteasome system (UPS) in a process controlled by the envelope-localized ubiquitin E3 ligase SUPPRESSOR OF PPI1 LOCUS1 (SP1) [5, 6]. Previous work showed that SP1-mediated regulation of chloroplast protein import contributes to the organellar proteome changes that occur during plant development (e.g., during de-etiolation). Here, we reveal a critical role for SP1 in plant responses to abiotic stress, which is a major and increasing cause of agricultural yield losses globally [7]. Arabidopsis plants lacking SP1 are hypersensitive to salt, osmotic, and oxidative stresses, whereas plants overexpressing SP1 are considerably more stress tolerant than wild-type. We present evidence that SP1 acts to deplete the TOC apparatus under stress conditions to limit the import of photosynthetic apparatus components, which may attenuate photosynthetic activity and reduce the potential for reactive oxygen species production and photo-oxidative damage. Our results indicate that chloroplast protein import is responsive to environmental cues, enabling dynamic regulation of the organellar proteome, and suggest new approaches for improving stress tolerance in crops.

  1. The chloroplast atpA gene cluster in Chlamydomonas reinhardtii. Functional analysis of a polycistronic transcription unit.

    Science.gov (United States)

    Drapier, D; Suzuki, H; Levy, H; Rimbault, B; Kindle, K L; Stern, D B; Wollman, F A

    1998-06-01

    Most chloroplast genes in vascular plants are organized into polycistronic transcription units, which generate a complex pattern of mono-, di-, and polycistronic transcripts. In contrast, most Chlamydomonas reinhardtii chloroplast transcripts characterized to date have been monocistronic. This paper describes the atpA gene cluster in the C. reinhardtii chloroplast genome, which includes the atpA, psbI, cemA, and atpH genes, encoding the alpha-subunit of the coupling-factor-1 (CF1) ATP synthase, a small photosystem II polypeptide, a chloroplast envelope membrane protein, and subunit III of the CF0 ATP synthase, respectively. We show that promoters precede the atpA, psbI, and atpH genes, but not the cemA gene, and that cemA mRNA is present only as part of di-, tri-, or tetracistronic transcripts. Deletions introduced into the gene cluster reveal, first, that CF1-alpha can be translated from di- or polycistronic transcripts, and, second, that substantial reductions in mRNA quantity have minimal effects on protein synthesis rates. We suggest that posttranscriptional mRNA processing is common in C. reinhardtii chloroplasts, permitting the expression of multiple genes from a single promoter.

  2. Direct measurement of calcium transport across chloroplast inner-envelope vesicles

    Energy Technology Data Exchange (ETDEWEB)

    Roh, M.H.; Shingles, R.; Cleveland, M.J.; McCarty, R.E. [Johns Hopkins Univ., Baltimore, MD (United States). Dept. of Biology

    1998-12-01

    The initial rate of Ca{sup 2+} movement across the inner-envelope membrane of pea (Pisum sativum L.) chloroplasts was directly measured by stopped-flow spectrofluorometry using membrane vesicles loaded with the Ca{sup 2+}-sensitive fluorophore fura-2. Calibration of fura-2 fluorescence was achieved by combining a ratiometric method with Ca{sup 2+}-selective minielectrodes to determine pCa values. The initial rate of Ca{sup 2+} influx in predominantly right-side-out inner-envelope membrane vesicles was greater than that in largely inside-out vesicles. Ca{sup 2+} movement was stimulated by an inwardly directed electrochemical proton gradient across the membrane vesicles, an effect that was diminished by the addition of valinomycin in the presence of K{sup +}. In addition, Ca{sup 2+} was shown to move across the membrane vesicles in the presence of K{sup +} diffusion potential gradient. The potential-stimulated rate of Ca{sup 2+} transport was slightly inhibited by diltiazem and greatly inhibited by ruthenium red. Other pharmacological agents such as LaCl{sub 3}, verapamil, and nifedipine had little or no effect. These results indicate that Ca{sup 2+} transport across the chloroplast inner envelope can occur by a potential-stimulated uniport mechanism.

  3. Dietary green-plant thylakoids decrease gastric emptying and gut transit, promote changes in the gut microbial flora, but does not cause steatorrhea

    DEFF Research Database (Denmark)

    Stenblom, Eva-Lena; Weström, Björn; Linninge, Caroline

    2016-01-01

    on GI passage, 16 rats were gavage-fed a control or thylakoid-supplemented high-fat diet (HFD) 30 min before receiving Evans blue. Another 16 rats were fed a control HFD or thylakoid HFD for two weeks prior to the intragastric challenge with Evans blue. The amount of Evans blue in the stomach...

  4. Supplementation by thylakoids to a high carbohydrate meal decreases feelings of hunger, elevates CCK levels and prevents postprandial hypoglycaemia in overweight women

    DEFF Research Database (Denmark)

    Stenblom, Eva-Lena; Montelius, Caroline; Östbring, Karolina

    2013-01-01

    over a 4-h period. Addition of thylakoids suppressed hunger motivation and increased secretion of CCK from 180 min, and prevented postprandial hypoglycaemia from 90 min following food intake. These effects indicate that thylakoids may intensify signals of satiety. This study therefore suggests...

  5. Electron spin resonance study of chloroplast photosynthetic activity in the presence of amphiphilic amines.

    Science.gov (United States)

    Sersen, F; Balgavý, P; Devínsky, F

    1990-12-01

    Electron spin resonance spectroscopy (ESR) was used to study the effects of amphiphilic amines of the carbamate, amide, and ester type and amine oxide on the photosynthetic system of spinach chloroplasts. The ESR signal II connected to the photosynthetic center PS II donor side was observed to diminish in the presence of amines, whereas that of PS I remained unchanged. The inhibition of PS II increased with the increasing of amine concentration. In the presence of amines, the light: dark chloroplast ESR signals ratio as well as the intensity of the ESR signal of unbound Mn2+ increased. It is suggested that the amphiphilic amines affect the structure of PS II and the electron transfer to PS I. The effects of the amines tested on the photosynthetic system correlate with their potency to perturb the lipid membrane structure.

  6. Dietary thylakoids reduce visceral fat mass and increase expression of genes involved in intestinal fatty acid oxidation in high-fat fed rats.

    Science.gov (United States)

    Stenblom, Eva-Lena; Egecioglu, Emil; Montelius, Caroline; Ramachandran, Deepti; Bonn, Britta; Weström, Björn; Mansouri, Abdelhak; Langhans, Wolfgang; Erlanson-Albertsson, Charlotte

    2016-09-01

    Thylakoids reduce body weight gain and body fat accumulation in rodents. This study investigated whether an enhanced oxidation of dietary fat-derived fatty acids in the intestine contributes to the thylakoid effects. Male Sprague-Dawley rats were fed a high-fat diet with (n = 8) or without thylakoids (n = 8) for 2 wk. Body weight, food intake, and body fat were measured, and intestinal mucosa was collected and analyzed. Quantitative real-time PCR was used to measure gene expression levels of key enzymes involved in fatty acid transport, fatty acid oxidation, and ketogenesis. Another set of thylakoid-treated (n = 10) and control rats (n = 10) went through indirect calorimetry. In the first experiment, thylakoid-treated rats (n = 8) accumulated 25% less visceral fat than controls. Furthermore, fatty acid translocase (Fat/Cd36), carnitine palmitoyltransferase 1a (Cpt1a), and mitochondrial 3-hydroxy-3-methylglutaryl-CoA synthase 2 (Hmgcs2) genes were upregulated in the jejunum of the thylakoid-treated group. In the second experiment, thylakoid-treated rats (n = 10) gained 17.5% less weight compared with controls and their respiratory quotient was lower, 0.86 compared with 0.91. Thylakoid-intake resulted in decreased food intake and did not cause steatorrhea. These results suggest that thylakoids stimulated intestinal fatty acid oxidation and ketogenesis, resulting in an increased ability of the intestine to handle dietary fat. The increased fatty acid oxidation and the resulting reduction in food intake may contribute to the reduced fat accumulation in thylakoid-treated animals.

  7. Direct chloroplast sequencing: comparison of sequencing platforms and analysis tools for whole chloroplast barcoding.

    Directory of Open Access Journals (Sweden)

    Marta Brozynska

    Full Text Available Direct sequencing of total plant DNA using next generation sequencing technologies generates a whole chloroplast genome sequence that has the potential to provide a barcode for use in plant and food identification. Advances in DNA sequencing platforms may make this an attractive approach for routine plant identification. The HiSeq (Illumina and Ion Torrent (Life Technology sequencing platforms were used to sequence total DNA from rice to identify polymorphisms in the whole chloroplast genome sequence of a wild rice plant relative to cultivated rice (cv. Nipponbare. Consensus chloroplast sequences were produced by mapping sequence reads to the reference rice chloroplast genome or by de novo assembly and mapping of the resulting contigs to the reference sequence. A total of 122 polymorphisms (SNPs and indels between the wild and cultivated rice chloroplasts were predicted by these different sequencing and analysis methods. Of these, a total of 102 polymorphisms including 90 SNPs were predicted by both platforms. Indels were more variable with different sequencing methods, with almost all discrepancies found in homopolymers. The Ion Torrent platform gave no apparent false SNP but was less reliable for indels. The methods should be suitable for routine barcoding using appropriate combinations of sequencing platform and data analysis.

  8. Protein trafficking to the complex chloroplasts of Euglena.

    Science.gov (United States)

    Vacula, Rostislav; Sláviková, Silvia; Schwartzbach, Steven D

    2007-01-01

    Proteins are delivered to Euglena chloroplasts using the secretory pathway. We describe analytical methods to study the intracellular trafficking of Euglena chloroplast proteins and a method to isolate preparative amounts of intact import competent chloroplasts for biochemical studies. Cells are pulse labeled with 35S-sulfate and chased with unlabeled sulfate allowing the trafficking and posttranslational processing of the labeled protein to be followed. Sucrose gradients are used to separate a 35S-labeled cell lysate into cytoplasmic, endoplasmic reticuum (ER), Golgi apparatus, chloroplast and mitochondrial fractions. Immunoprecipitation of each gradient fraction allows identification of the intracellular compartment containing a specific 35S-labeled protein at different times after synthesis delineating the trafficking pathway. Because sucrose gradients cannot be used to isolate preparative amounts of highly purified chloroplasts for biochemical characterization, a preparative high-yield procedure using Percoll gradients to isolate highly purified import competent chloroplasts is also presented.

  9. Analysis of Chloroplast Ultrastructure, Photosystem Ⅱ Light Harvesting Complexes and Chlorophyll Synthesis in a Chlorophyll-Less Rice Mutant W2555

    Institute of Scientific and Technical Information of China (English)

    XU Pei-zhou; LI Yun; YUAN Shu; ZHANG Hong-yu; WANG Xu-dong; LIN Hong-hui; WU Xian-jun

    2006-01-01

    A comparative study on chloroplast ultrastructure and light harvesting complex of photosystem Ⅱ (LHC Ⅱ) was conducted between a new rice mutant (W2555) and its wild type (WT). The chloroplasts of W2555 had less thylakoids and grana stacks compared with the wild type. There was no significant change in the composition of LHC Ⅱ polypeptide in W2555, while a decline had been noted in LHC Ⅱ content. Northern blot analysis with a specific cab gene probe showed no appreciable difference in the LHC Ⅱ mRNA level between the W2555 and its wild type. The precursors of chlorophyll synthesis, δ-aminolevulinic acid (ALA)and porphobilinogen (PBG) were over accumulated in W2555, but the other precursors were all decreased. These results indicated that the decreased level of LHC Ⅱ in the mutant W2555 was attributed to the change of cab gene transcription, but a blockage in chlorophyll biosynthesis due to the formation of uroporphyrinogen Ⅲ (Urogen Ⅲ).

  10. Phototropins and chloroplast activity in plant blue light signaling

    OpenAIRE

    Goh, Chang-Hyo

    2009-01-01

    In plants, phototropins 1 (phot1) and 2 (phot2) mediate chloroplast movement to blue light (BL). A recent report showed that phototropins (phot) are required for the expression of chloroplast genes in rice. The light-induced responses of phot1a rice mutants result in H2O2-mediated damage to chloroplast photosystems, indicating that phot-regulated responses might be associated with the other photoreceptor, such as cryptochrome (cry) BL receptor. This suggests diversification and specialization...

  11. Expressing PHB synthetic genes through chloroplast genetic engineering

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Chloroplast integration and expression vector containing expression cassettes for phbB, phbA, phbC and aadA genes was constructed and bombarded into the tobacco chloroplast genome. Transplastomic plants were analyzed with PCR and Southern blot. Their homoplastomy was also judged. Northern dot and RT-PCR analysis were employed to investigate transgene expression at transcriptional level. The results indicate that the chloroplast transformation system is compatible for poly-3-hydroxybutyrate (PHB) production.

  12. Complex chloroplast RNA metabolism: just debugging the genetic programme?

    Directory of Open Access Journals (Sweden)

    Schmitz-Linneweber Christian

    2008-08-01

    Full Text Available Abstract Background The gene expression system of chloroplasts is far more complex than that of their cyanobacterial progenitor. This gain in complexity affects in particular RNA metabolism, specifically the transcription and maturation of RNA. Mature chloroplast RNA is generated by a plethora of nuclear-encoded proteins acquired or recruited during plant evolution, comprising additional RNA polymerases and sigma factors, and sequence-specific RNA maturation factors promoting RNA splicing, editing, end formation and translatability. Despite years of intensive research, we still lack a comprehensive explanation for this complexity. Results We inspected the available literature and genome databases for information on components of RNA metabolism in land plant chloroplasts. In particular, new inventions of chloroplast-specific mechanisms and the expansion of some gene/protein families detected in land plants lead us to suggest that the primary function of the additional nuclear-encoded components found in chloroplasts is the transgenomic suppression of point mutations, fixation of which occurred due to an enhanced genetic drift exhibited by chloroplast genomes. We further speculate that a fast evolution of transgenomic suppressors occurred after the water-to-land transition of plants. Conclusion Our inspections indicate that several chloroplast-specific mechanisms evolved in land plants to remedy point mutations that occurred after the water-to-land transition. Thus, the complexity of chloroplast gene expression evolved to guarantee the functionality of chloroplast genetic information and may not, with some exceptions, be involved in regulatory functions.

  13. Nanophotonics of Chloroplasts for Bio-Inspired Solar Energy Materials

    Science.gov (United States)

    Gourley, Paul L.; Gourley, Cheryl R.

    2011-03-01

    In the search for new energy sources, lessons can be learned from chloroplast photonics. The nano-architecture of chloroplasts is remarkably well-adapted to mediate sunlight interactions for efficient energy conversion. We carried out experiments with chloroplasts isolated from spinach and leaf lettuce to elucidate the relationship between nano-architecture, biomolecular composition and photonic properties. We obtained high-resolution microscopic images of single chloroplasts to identify geometries of chloroplasts and interior grana. We performed micro-spectroscopy to identify strengths of absorption and fluorescence transitions and related them to broadband reflectance and transmittance spectra of whole leaf structures. Finally, the nonlinear optical properties were investigated with nanolaser spectroscopy by placing chloroplasts into micro-resonators and optically pumping. These spectra reveal chloroplast photonic modes and allow measurement of single chloroplast light scattering cross section, polarizability, and refractive index. The nanolaser spectra recorded at increasing pump powers enabled us to observe non-linear optics, photon dynamics, and stimulated emission from single chloroplasts. All of these experiments provide insight into plant photonics and inspiration of paradigms for synthetic biomaterials to harness sunlight in new ways.

  14. New insights into dynamic actin-based chloroplast photorelocation movement.

    Science.gov (United States)

    Kong, Sam-Geun; Wada, Masamitsu

    2011-09-01

    Chloroplast movement is essential for plants to survive under various environmental light conditions. Phototropins-plant-specific blue-light-activated receptor kinases-mediate the response by perceiving light intensity and direction. Recently, novel chloroplast actin (cp-actin) filaments have been identified as playing a pivotal role in the directional chloroplast photorelocation movement. Encouraging progress has recently been made in this field of research through molecular genetics and cell biological analyses. This review describes factors that have been identified as being involved in chloroplast movement and their roles in the regulation of cp-actin filaments, thus providing a basis for reflection on their biochemical activities and functions.

  15. Novel protein phosphorylation site identification in spinach stroma membranes by titanium dioxide microcolumns and tandem mass spectrometry

    DEFF Research Database (Denmark)

    Rinalducci, Sara; Larsen, Martin Røssel; Mohammed, Shabaz

    2006-01-01

    In this work, spinach stroma membrane, instead of thylakoid, has been investigated for the presence of phosphorylated proteins. We identified seven previously unknown phosphorylation sites by taking advantage of TiO(2) phosphopeptides enrichment coupled to mass spectrometric analysis. Upon illumi...

  16. X-ray structure, thermodynamics, elastic properties and MD simulations of cardiolipin/dimyristoylphosphatidylcholine mixed membranes

    DEFF Research Database (Denmark)

    Boscia, Alexander L.; Treece, Bradley W.; Mohammadyani, Dariush

    2014-01-01

    Cardiolipins (CLs) are important biologically for their unique role in biomembranes that couple phosphorylation and electron transport like bacterial plasma membranes, chromatophores, chloroplasts and mitochondria. CLs are often tightly coupled to proteins involved in oxidative phosphorylation. T...

  17. Light- and metabolism-related regulation of the chloroplast ATP synthase has distinct mechanisms and functions.

    Science.gov (United States)

    Kohzuma, Kaori; Dal Bosco, Cristina; Meurer, Jörg; Kramer, David M

    2013-05-01

    The chloroplast CF0-CF1-ATP synthase (ATP synthase) is activated in the light and inactivated in the dark by thioredoxin-mediated redox modulation of a disulfide bridge on its γ subunit. The activity of the ATP synthase is also fine-tuned during steady-state photosynthesis in response to metabolic changes, e.g. altering CO2 levels to adjust the thylakoid proton gradient and thus the regulation of light harvesting and electron transfer. The mechanism of this fine-tuning is unknown. We test here the possibility that it also involves redox modulation. We found that modifying the Arabidopsis thaliana γ subunit by mutating three highly conserved acidic amino acids, D211V, E212L, and E226L, resulted in a mutant, termed mothra, in which ATP synthase which lacked light-dark regulation had relatively small effects on maximal activity in vivo. In situ equilibrium redox titrations and thiol redox-sensitive labeling studies showed that the γ subunit disulfide/sulfhydryl couple in the modified ATP synthase has a more reducing redox potential and thus remains predominantly oxidized under physiological conditions, implying that the highly conserved acidic residues in the γ subunit influence thiol redox potential. In contrast to its altered light-dark regulation, mothra retained wild-type fine-tuning of ATP synthase activity in response to changes in ambient CO2 concentrations, indicating that the light-dark- and metabolic-related regulation occur through different mechanisms, possibly via small molecule allosteric effectors or covalent modification.

  18. A Novel Thylakoid Ascorbate Peroxidase from Jatrophacurcas Enhances Salt Tolerance in Transgenic Tobacco

    Directory of Open Access Journals (Sweden)

    Zhibin Liu

    2013-12-01

    Full Text Available Ascorbate peroxidase (APX plays an important role in the metabolism of hydrogen peroxide in higher plants. In the present study, a novel APX gene (JctAPX was cloned from Jatropha curcas L. The deduced amino acid sequence was similar to that of APX of some other plant species. JctAPX has a chloroplast transit peptide and was localized to the chloroplasts by analysis with a JctAPX-green fluorescent protein (GFP fusion protein. Quantitative polymerase chain reaction (qPCR analysis showed that JctAPX was constitutively expressed in different tissues from J. curcas and was upregulated by NaCl stress. To characterize its function in salt tolerance, the construct p35S: JctAPX was created and successfully introduced into tobacco by Agrobacterium-mediated transformation. Compared with wild type (WT, the transgenic plants exhibited no morphological abnormalities in the no-stress condition. However, under 200 mM NaCl treatment, JctAPX over-expressing plants showed increased tolerance to salt during seedling establishment and growth. In addition, the transgenic lines showed higher chlorophyll content and APX activity, which resulted in lower H2O2 content than WT when subjected to 400 mM NaCl stress. These results suggest that the increased APX activity in the chloroplasts from transformed plants increased salt tolerance by enhancing reactive oxygen species (ROS-scavenging capacity under short-term NaCl stress conditions.

  19. Border control: selectivity of chloroplast protein import and regulation at the TOC-complex.

    Science.gov (United States)

    Demarsy, Emilie; Lakshmanan, Ashok M; Kessler, Felix

    2014-01-01

    Plants have evolved complex and sophisticated molecular mechanisms to regulate their development and adapt to their surrounding environment. Particularly the development of their specific organelles, chloroplasts and other plastid-types, is finely tuned in accordance with the metabolic needs of the cell. The normal development and functioning of plastids require import of particular subsets of nuclear encoded proteins. Most preproteins contain a cleavable sequence at their N terminal (transit peptide) serving as a signal for targeting to the organelle and recognition by the translocation machinery TOC-TIC (translocon of outer membrane complex-translocon of inner membrane complex) spanning the dual membrane envelope. The plastid proteome needs constant remodeling in response to developmental and environmental factors. Therefore selective regulation of preprotein import plays a crucial role in plant development. In this review we describe the diversity of transit peptides and TOC receptor complexes, and summarize the current knowledge and potential directions for future research concerning regulation of the different Toc isoforms.

  20. Anchoring a plant cytochrome P450 via PsaM to the thylakoids in Synechococcus sp. PCC 7002: evidence for light-driven biosynthesis.

    Directory of Open Access Journals (Sweden)

    Lærke Münter Lassen

    Full Text Available Plants produce an immense variety of specialized metabolites, many of which are of high value as their bioactive properties make them useful as for instance pharmaceuticals. The compounds are often produced at low levels in the plant, and due to their complex structures, chemical synthesis may not be feasible. Here, we take advantage of the reducing equivalents generated in photosynthesis in developing an approach for producing plant bioactive natural compounds in a photosynthetic microorganism by functionally coupling a biosynthetic enzyme to photosystem I. This enables driving of the enzymatic reactions with electrons extracted from the photosynthetic electron transport chain. As a proof of concept, we have genetically fused the soluble catalytic domain of the cytochrome P450 CYP79A1, originating from the endoplasmic reticulum membranes of Sorghum bicolor, to a photosystem I subunit in the cyanobacterium Synechococcus sp. PCC 7002, thereby targeting it to the thylakoids. The engineered enzyme showed light-driven activity both in vivo and in vitro, demonstrating the possibility to achieve light-driven biosynthesis of high-value plant specialized metabolites in cyanobacteria.

  1. Arabidopsis thaliana leaves with altered chloroplast numbers and chloroplast movement exhibit impaired adjustments to both low and high light.

    Science.gov (United States)

    Königer, Martina; Delamaide, Joy A; Marlow, Elizabeth D; Harris, Gary C

    2008-01-01

    The effects of chloroplast number and size on the capacity for blue light-dependent chloroplast movement, the ability to increase light absorption under low light, and the susceptibility to photoinhibition were investigated in Arabidopsis thaliana. Leaves of wild-type and chloroplast number mutants with mean chloroplast numbers ranging from 120 to two per mesophyll cell were analysed. Chloroplast movement was monitored as changes in light transmission through the leaves. Light transmission was used as an indicator of the ability of leaves to optimize light absorption. The ability of leaves to deal with 3 h of high light stress at 10 degrees C and their capacity to recover in low light was determined by measuring photochemical efficiencies of PSII using chlorophyll a fluorescence. Chloroplast movement was comparable in leaves ranging in chloroplast numbers from 120 to 30 per mesophyll cell: the final light transmission levels after exposure to 0.1 (accumulation response) and 100 micromol photons m(-2) s(-1) (avoidance response) were indistinguishable, the chloroplasts responded quickly to small increases in light intensity and the kinetics of movement were similar. However, when chloroplast numbers per mesophyll cell decreased to 18 or below, the accumulation response was significantly reduced. The avoidance response was only impaired in mutants with nine or fewer chloroplasts, both in terms of final transmission levels and the speed of movement. Only mutants lacking both blue light receptors (phot1/phot2) or those with drastically reduced chloroplast numbers and severely impacted avoidance responses showed a reduced ability to recover from high light stress.

  2. Orientation of the pigment molecules in the chloroplast

    NARCIS (Netherlands)

    Goedheer, J.C.

    1955-01-01

    Dichroism, absorption anisotropy, and anomal dispersion of birefringence were measured in the big lamellate chloroplasts of Mougeotia. The results of these measurements indicate a certain orientation of the chlorophyll molecules, and to a smaller extent, of the carotenoids in the chloroplast. In sp

  3. Expression of eukaryotic polypeptides in chloroplasts

    Energy Technology Data Exchange (ETDEWEB)

    Mayfield, Stephen P

    2013-06-04

    The present invention relates to a gene expression system in eukaryotic and prokaryotic cells, preferably plant cells and intact plants. In particular, the invention relates to an expression system having a RB47 binding site upstream of a translation initiation site for regulation of translation mediated by binding of RB47 protein, a member of the poly(A) binding protein family. Regulation is further effected by RB60, a protein disulfide isomerase. The expression system is capable of functioning in the nuclear/cytoplasm of cells and in the chloroplast of plants. Translation regulation of a desired molecule is enhanced approximately 100 fold over that obtained without RB47 binding site activation.

  4. Effect of cationic plastoquinone SkQ1 on electron transfer reactions in chloroplasts and mitochondria from pea seedlings.

    Science.gov (United States)

    Samuilov, V D; Kiselevsky, D B

    2015-04-01

    Plastoquinone bound with decyltriphenylphosphonium cation (SkQ1) penetrating through the membrane in nanomolar concentrations inhibited H2O2 generation in cells of epidermis of pea seedling leaves that was detected by the fluorescence of 2',7'-dichlorofluorescein. Photosynthetic electron transfer in chloroplasts isolated from pea leaves is suppressed by SkQ1 at micromolar concentrations: the electron transfer in chloroplasts under the action of photosystem II or I (with silicomolybdate or methyl viologen as electron acceptors, respectively) is more sensitive to SkQ1 than under the action of photosystem II + I (with ferricyanide or p-benzoquinone as electron acceptors). SkQ1 reduced by borohydride is oxidized by ferricyanide, p-benzoquinone, and, to a lesser extent, by silicomolybdate, but not by methyl viologen. SkQ1 is not effective as an electron acceptor supporting O2 evolution from water in illuminated chloroplasts. The data on suppression of photosynthetic O2 evolution or consumption show that SkQ1, similarly to phenazine methosulfate, causes conversion of the chloroplast redox-chain from non-cyclic electron transfer mode to the cyclic mode without O2 evolution. Oxidation of NADH or succinate in mitochondria isolated from pea roots is stimulated by SkQ1.

  5. Exploring ligand recognition, selectivity and dynamics of TPR domains of chloroplast Toc64 and mitochondria Om64 from Arabidopsis thaliana.

    Science.gov (United States)

    Panigrahi, Rashmi; Whelan, James; Vrielink, Alice

    2014-06-01

    The study aims to gain insight into the mode of ligand recognition by tetratricopeptide repeat (TPR) domains of chloroplast translocon at the outer envelope of chloroplast (Toc64) and mitochondrial Om64, two paralogous proteins that mediate import of proteins into chloroplast and mitochondria, respectively. Chaperone proteins associate with precursor proteins in the cytosol to maintain them in a translocation competent conformation and are recognized by Toc64 and Om64 that are located on the outer membrane of the target organelle. Heat shock proteins (Hsp70) and Hsp90 are two chaperones, which are known to play import roles in protein import. The C-termini of these chaperones are known to interact with the TPR domain of chloroplast Toc64 and mitochondrial Om64 in Arabidopsis thaliana (At). Using a molecular dynamics approach and binding energy calculations, we identify important residues involved in the interactions. Our findings suggest that the TPR domain from AtToc64 has higher affinity towards C-terminal residues of Hsp70. The interaction occurs as the terminal helices move towards each other enclosing the cradle on interaction of AtHsp70 with the TPR domain. In contrast, the TPR domain from AtOm64 does not discriminate between the C-termini of Hsp70 and Hsp90. These binding affinities are discussed with respect to our knowledge of protein targeting and specificity of protein import into endosymbiotic organelles in plant cells.

  6. Non-contact intracellular binding of chloroplasts in vivo

    Science.gov (United States)

    Li, Yuchao; Xin, Hongbao; Liu, Xiaoshuai; Li, Baojun

    2015-06-01

    Non-contact intracellular binding and controllable manipulation of chloroplasts in vivo was demonstrated using an optical fiber probe. Launching a 980-nm laser beam into a fiber, which was placed about 3 μm above the surface of a living plant (Hydrilla verticillata) leaf, enabled stable binding of different numbers of chloroplasts, as well as their arrangement into one-dimensional chains and two-dimensional arrays inside the leaf without damaging the chloroplasts. Additionally, the formed chloroplast chains were controllably transported inside the living cells. The optical force exerted on the chloroplasts was calculated to explain the experimental results. This method provides a flexible method for studying intracellular organelle interaction with highly organized organelle-organelle contact in vivo in a non-contact manner.

  7. Chloroplast division during leaf development of Xanthium pensylvanicum Wallr. (Compositae

    Directory of Open Access Journals (Sweden)

    Roman Maksymowych

    2014-02-01

    Full Text Available Division and growth of chloroplasts was studied during leaf development of Xanthium pensylvanicum at various stages of development represented by the leaf plastochron index.Between leaf plastochron indices -1.00 and 2.56 chloroplast division was observed with little enlargement. Between 2.50 and 5.00 chloroplasts enlarged in diameter with an average rate of 0.21 µm per day. At leaf plastochron index 5.00 chloroplasts attained their mature size of 6.12 µm. No chloroplast division was found after leaf plastochron index 2.50. A change in shape of plastids from spherical proplastids to discoidal accompanied their growth during stages 2.50 and 5.00.

  8. Laser Stimulation of the Chloroplast/Endoplasmic Reticulum Nexus in Tobacco Transiently Produces Protein Aggregates (Boluses) within the Endoplasmic Reticulum and Stimulates Local ER Remodeling

    Institute of Scientific and Technical Information of China (English)

    Lawrence R. Griffing

    2011-01-01

    Does the ER subdomain that associates with the chloroplast in vivo,hereafter referred to as the chloroplast/ER nexus,play a role in protein flow within the ER? In studies of tobacco cells either constitutively or transiently expressing ER-retained luminal,GFP-HDEL,or trans-membrane,YFP-RHD3,fluorescent fusion proteins,brief 405-nm (3-6-mW) laser stimulation of the nexus causes a qualitative difference in the movement and behavior of proteins in the ER.Photostimulating the nexus produces fluorescent protein punctate aggregates (boluses) within the lumen and membrane of the ER.The aggregation propagates through the membrane network throughout the cell,but within minutes can revert to normal,with disaggregation propagating back toward the originally photostimulated nexus.In the meantime,the ER grows and anastomoses around the chloroplast,forming a dense cisternal and tubular network.If this network is again photostimulated,bolus formation does not recur and,if the photostimulation results in photobleaching,fluorescence recovery after photobleaching occurs as it would typically in areas away from the nexus.Bolus propagation is not mediated by the actin cytoskeleton,but can be reversed by pre-conditioning the cells for 30 min with high,40-45℃,temperature (heat stress).Because it is not reversed with heat stress,the reorganization of the ER at the nexus following photostimulation is a separate event.

  9. The chloroplast permease PIC1 regulates plant growth and development by directing homeostasis and transport of iron.

    Science.gov (United States)

    Duy, Daniela; Stübe, Roland; Wanner, Gerhard; Philippar, Katrin

    2011-04-01

    The membrane-spanning protein PIC1 (for permease in chloroplasts 1) in Arabidopsis (Arabidopsis thaliana) was previously described to mediate iron transport across the inner envelope membrane of chloroplasts. The albino phenotype of pic1 knockout mutants was reminiscent of iron-deficiency symptoms and characterized by severely impaired plastid development and plant growth. In addition, plants lacking PIC1 showed a striking increase in chloroplast ferritin clusters, which function in protection from oxidative stress by sequestering highly reactive free iron in their spherical protein shell. In contrast, PIC1-overexpressing lines (PIC1ox) in this study rather resembled ferritin loss-of-function plants. PIC1ox plants suffered from oxidative stress and leaf chlorosis, most likely originating from iron overload in chloroplasts. Later during growth, plants were characterized by reduced biomass as well as severely defective flower and seed development. As a result of PIC1 protein increase in the inner envelope membrane of plastids, flower tissue showed elevated levels of iron, while the content of other transition metals (copper, zinc, manganese) remained unchanged. Seeds, however, specifically revealed iron deficiency, suggesting that PIC1 overexpression sequestered iron in flower plastids, thereby becoming unavailable for seed iron loading. In addition, expression of genes associated with metal transport and homeostasis as well as photosynthesis was deregulated in PIC1ox plants. Thus, PIC1 function in plastid iron transport is closely linked to ferritin and plastid iron homeostasis. In consequence, PIC1 is crucial for balancing plant iron metabolism in general, thereby regulating plant growth and in particular fruit development.

  10. 干旱胁迫对3种不同光合类型荒漠植物叶绿体和线粒体超微结构的影响%Effect of Soil Drought Stress on the Ultramicrostructures of Chloroplasts and Mitochondria in Three Desert Plants with Different Photosynthetic Types

    Institute of Scientific and Technical Information of China (English)

    闻志彬; 莱孜提·库里库; 张明理

    2016-01-01

    clear and the thylakoids expanded,but the differences between these two plants were that mitochondria of S.junatovii firstly appeared degradation,and its inclusions were partly lost.For S.laricifolia,the outer membrane of mitochondria was deformed and the ridges were re-duced.The mitochondria of S.arbuscula remained normal,but the chloroplasts slightly expanded.(4 ) Under the severe drought stress,the chloroplasts of S.junatovii and S.laricifolia were damaged and mi-tochondria were degraded.For S.arbuscula,the chloroplasts expanded,the outer membrane of mitochon-dria was deformed and the ridges were reduced.These results have showed that the damage degree of S. arbuscula in chloroplasts and mitochondria under different degree of drought stress was the lowest;For S. junatovii and S.laricifolia,the damage degree of chloroplasts under drought stress was similar;the mi-tochondria had better tolerance to drought stress than chloroplasts in S.laricifolia and S.arbuscula.

  11. Chloroplasts move towards the nearest anticlinal walls under dark condition.

    Science.gov (United States)

    Tsuboi, Hidenori; Wada, Masamitsu

    2012-03-01

    Chloroplasts change their intracellular positions in response to their light environment. Under darkness, chloroplasts assume special positions that are different from those under light conditions. Here, we analyzed chloroplast dark positioning using Adiantum capillus-veneris gametophyte cells. When chloroplasts were transferred into darkness, during the first 1-5 h, they moved towards the anticlinal cell walls bordering the adjacent cells rather rapidly. Then, they slowed down and accumulated at the anticlinal walls gradually over the following 24-36 h. The chloroplast movements could be roughly classified into two different categories: initial rapid straight movement and later, slow staggering movement. When the chloroplast accumulation response was induced in dark-adapted cells by partial cell irradiation with a microbeam targeted to the center of the cells, chloroplasts moved towards the beam spot from the anticlinal walls. However, when the microbeam was switched off, they moved to the nearest anticlinal walls and not to their original positions if they were not the closest, indicating that they know the direction of the nearest anticlinal wall and do not have particular areas that they migrate to during dark positioning.

  12. Transposon-induced nuclear mutations that alter chloroplast gene expression

    Energy Technology Data Exchange (ETDEWEB)

    Barkan, A.

    1992-01-01

    The goal of this project is to use mutant phenotypes as a guide to nuclear genes that determine the timing and localization of chloroplast development The immediate goals are to identify nuclear mutants with defects in chloroplast gene expression from maize lines harboring active Mu transposons; characterize their phenotypes to determine the precise defect in gene expression; clone several of the most interesting mutations by exploiting the transposon tag; and use the clones to further define the roles of these genes in modulating chloroplast gene expression. Three mutants were described earlier that had global defects in chloroplast gene expression. We have found that two of these mutations are allelic. Both alleles have global defects in chloroplast translation initiation, as revealed by the failure to assemble chloroplast mRNAs into polysomes. We have isolated and characterized three new mutants from Mu lines that have novel defects in chloroplast RNA metabolism. We are now ready to begin the task of cloning several of these genes, by using the Mu transposon tag.

  13. Chloroplast: The Trojan Horse in Plant-Virus Interaction.

    Science.gov (United States)

    Bhattacharyya, Dhriti; Chakraborty, Supriya

    2017-01-05

    Chloroplast is one of the most dynamic organelle of a plant cell. It carries out photosynthesis, synthesizes major phytohormones, takes active part in defence response, and is crucial for inter-organelle signaling. Viruses, on the other hand, are extremely strategic in manipulating the internal environment of the host cell. Chloroplast, a prime target for viruses, undergoes enormous structural and functional damage during viral infection. In fact, large proportions of affected gene products in a virus infected plant are closely associated to chloroplast and photosynthesis process. Although chloroplast is deficient in gene-silencing machinery, it elicits effector-triggered immune response against viral pathogens. Virus infection induces the organelle to produce extensive network of stromules which are involved in both viral propagation and anti-viral defence. From last few decades' study, involvement of chloroplast in regulating plant-virus interaction has become increasingly evident. Current review presents an exhaustive account of these facts, with their implication in pathogenicity. We have attempted to highlight the intricacies of chloroplast-virus interaction and explained the existing gaps in current knowledge, which will promote the virologists to utilize the chloroplast genome-based antiviral resistance in economically important crops. This article is protected by copyright. All rights reserved.

  14. Identification of an Atypical Membrane Protein Involved in the Formation of Protein Disulfide Bonds in Oxygenic Photosynthetic Organisms*S⃞

    OpenAIRE

    Singh, Abhay K.; Bhattacharyya-Pakrasi, Maitrayee; Pakrasi, Himadri B.

    2008-01-01

    The evolution of oxygenic photosynthesis in cyanobacteria nearly three billion years ago provided abundant reducing power and facilitated the elaboration of numerous oxygen-dependent reactions in our biosphere. Cyanobacteria contain an internal thylakoid membrane system, the site of photosynthesis, and a typical Gram-negative envelope membrane system. Like other organisms, the extracytoplasmic space in cyanobacteria houses numerous cysteine-containing proteins. However, the ...

  15. Effects of plant density on the photosynthetic and chloroplast characteristics of maize under high-yielding conditions

    Science.gov (United States)

    Ren, Baizhao; Liu, Wei; Zhang, Jiwang; Dong, Shuting; Liu, Peng; Zhao, Bin

    2017-04-01

    Plant density has been recognized as a major factor determining the grain yield. The photosynthetic performance changes as the density increases. The main objective of this research was to evaluate responses of photosynthetic performance and chloroplast ultrastructure to planting densities in two summer maize ( Zea mays L.) hybrids Denghai661 (DH661) and Nongda108 (ND108). DH661 was planted at densities of 30,000, 45,000, 60,000, 75,000, 90,000, 105,000, 120,000, or 135,000 plants ha-1. ND108 was planted at densities of 30,000, 45,000, 60,000, 75,000, or 90,000 plants ha-1. Research variables included leaf area, grain yield, chlorophyll content, leaf gas exchange parameters, number of chloroplasts, and chloroplast ultrastructure. As plant density increased, chlorophyll a and b content significantly decreased; carotenoids initially decreased and then increased; the net photosynthetic rate during each growth period significantly decreased; the membrane structure of mesophyll cells was gradually damaged; the number of chloroplasts significantly decreased; the external form of chloroplasts shifted from long and oval to elliptical or circular; the number of grana significantly decreased, while the number of grana lamellae increased; grana gradually became hypogenetic and eventually dissolved; plot yield increased; and yield per plant significantly decreased. The yield per plant of DH661 at 135,000 plants ha-1 and that of ND108 at 90,000 plants ha-1 decreased by 65.8 and 42.5%, respectively, compared with those at 30,000 plants ha-1.

  16. 2012 MITOCHONDRIA AND CHLOROPLASTS GORDON RESEARCH CONFERENCE & GORDON RESEARCH SEMINAR, JULY 29 - AUGUST 3, 2012

    Energy Technology Data Exchange (ETDEWEB)

    Barkan, Alice

    2012-08-03

    The 2012 Gordon Research Conference on Mitochondria and Chloroplasts will assemble an international group of scientists investigating fundamental properties of these organelles, and their integration into broader physiological processes. The conference will emphasize the many commonalities between mitochondria and chloroplasts: their evolution from bacterial endosymbionts, their genomes and gene expression systems, their energy transducing membranes whose proteins derive from both nuclear and organellar genes, the challenge of maintaining organelle integrity in the presence of the reactive oxygen species that are generated during energy transduction, their incorporation into organismal signaling pathways, and more. The conference will bring together investigators working in animal, plant, fungal and protozoan systems who specialize in cell biology, genetics, biochemistry, physiology, proteomics, genomics, and structural biology. As such, this conference will provide a unique forum that engenders cross-disciplinary discussions concerning the biogenesis, dynamics, and regulation of these key cellular structures. By fostering interactions among mammalian, fungal and plant organellar biologists, this conference also provides a conduit for the transmission of mechanistic insights obtained in model organisms to applications in medicine and agriculture. The 2012 conference will highlight areas that are moving rapidly and emerging themes. These include new insights into the ultrastructure and organization of the energy transducing membranes, the coupling of organellar gene expression with the assembly of photosynthetic and respiratory complexes, the regulatory networks that couple organelle biogenesis with developmental and physiological signals, the signaling events through which organellar physiology influences nuclear gene expression, and the roles of organelles in disease and development.

  17. Dimerization of TOC receptor GTPases and its implementation for the control of protein import into chloroplasts.

    Science.gov (United States)

    Aronsson, Henrik; Jarvis, Paul

    2011-06-01

    Pre-protein import into chloroplasts is facilitated by multiprotein translocon complexes in the envelope membranes. Major components of the TOC (translocon at the outer envelope membrane of chloroplasts) complex are the receptor proteins Toc33 and Toc159. These two receptors are related GTPases, and they are predicted to engage in homodimerization and/or heterodimerization. Although such dimerization has been studied extensively, its exact function in vivo remains elusive. In this issue of the Biochemical Journal, Oreb et al. present evidence that homodimerization of Toc33 prevents nucleotide exchange, thereby locking the receptor in the GDP-loaded state and preventing further activity. Pre-protein arrival is proposed to release this lock, through disruption of the dimer and subsequent nucleotide exchange. The Toc33-bound pre-protein is then able to progress to downstream steps in the translocation mechanism, with GTP hydrolysis defining another important control point as well as preparing the receptor for the next pre-protein client. These new results are discussed in the context of previous findings pertaining to TOC receptor dimerization and function.

  18. Two kinesin-like proteins mediate actin-based chloroplast movement in Arabidopsis thaliana

    OpenAIRE

    Suetsugu, Noriyuki; Yamada, Noboru; Kagawa, Takatoshi; Yonekura, Hisashi; Uyeda, Taro Q. P.; Kadota, Akeo; Wada, Masamitsu

    2010-01-01

    Organelle movement is essential for efficient cellular function in eukaryotes. Chloroplast photorelocation movement is important for plant survival as well as for efficient photosynthesis. Chloroplast movement generally is actin dependent and mediated by blue light receptor phototropins. In Arabidopsis thaliana, phototropins mediate chloroplast movement by regulating short actin filaments on chloroplasts (cp-actin filaments), and the chloroplast outer envelope protein CHUP1 is necessary for c...

  19. Origins of prokaryotes, eukaryotes, mitochondria, and chloroplasts

    Science.gov (United States)

    Schwartz, R. M.; Dayhoff, M. O.

    1978-01-01

    A computer branching model is used to analyze cellular evolution. Attention is given to certain key amino acids and nucleotide residues (ferredoxin, 5s ribosomal RNA, and c-type cytochromes) because of their commonality over a wide variety of cell types. Each amino acid or nucleotide residue is a sequence in an inherited biological trait; and the branching method is employed to align sequences so that changes reflect substitution of one residue for another. Based on the computer analysis, the symbiotic theory of cellular evolution is considered the most probable. This theory holds that organelles, e.g., mitochondria and chloroplasts invaded larger bodies, e.g., bacteria, and combined functions to form eucaryotic cells.

  20. PSII-LHCII supercomplex organizations in photosynthetic membrane by coarse-grained simulation.

    Science.gov (United States)

    Lee, Cheng-Kuang; Pao, Chun-Wei; Smit, Berend

    2015-03-12

    Green plant photosystem II (PSII) and light-harvesting complex II (LHCII) in the stacked grana regions of thylakoid membranes can self-organize into various PSII-LHCII supercomplexes with crystalline or fluid-like supramolecular structures to adjust themselves with external stimuli such as high/low light and temperatures, rendering tunable solar light absorption spectrum and photosynthesis efficiencies. However, the mechanisms controlling the PSII-LHCII supercomplex organizations remain elusive. In this work, we constructed a coarse-grained (CG) model of the thylakoid membrane including lipid molecules and a PSII-LHCII supercomplex considering association/dissociation of moderately bound-LHCIIs. The CG interaction between CG beads were constructed based on electron microscope (EM) experimental results, and we were able to simulate the PSII-LHCII supramolecular organization of a 500 × 500 nm(2) thylakoid membrane, which is compatible with experiments. Our CGMD simulations can successfully reproduce order structures of PSII-LHCII supercomplexes under various protein packing fractions, free-LHCII:PSII ratios, and temperatures, thereby providing insights into mechanisms leading to PSII-LHCII supercomplex organizations in photosynthetic membranes.

  1. New Insights into Dynamic Actin-Based Chloroplast Photorelocation Movement

    Institute of Scientific and Technical Information of China (English)

    Sam-Geun Kong; Masamitsu Wada

    2011-01-01

    Chloroplast movement is essential for plants to survive under various environmental light conditions.Phototropins-plant-specific blue-light-activated receptor kinases-mediate the response by perceiving light intensity and direction.Recently,novel chloroplast actin (cp-actin) filaments have been identified as playing a pivotal role in the directional chloroplast photorelocation movement.Encouraging progress has recently been made in this field of research through molecular genetics and cell biological analyses.This review describes factors that have been identified as being involved in chloroplast movement and their roles in the regulation of cp-actin filaments,thus providing a basis for reflection on their biochemical activities and functions.

  2. Complete Chloroplast Genome of Tanaecium tetragonolobum: The First Bignoniaceae Plastome.

    Directory of Open Access Journals (Sweden)

    Alison Gonçalves Nazareno

    Full Text Available Bignoniaceae is a Pantropical plant family that is especially abundant in the Neotropics. Members of the Bignoniaceae are diverse in many ecosystems and represent key components of the Tropical flora. Despite the ecological importance of the Bignoniaceae and all the efforts to reconstruct the phylogeny of this group, whole chloroplast genome information has not yet been reported for any members of the family. Here, we report the complete chloroplast genome sequence of Tanaecium tetragonolobum (Jacq. L.G. Lohmann, which was reconstructed using de novo and referenced-based assembly of single-end reads generated by shotgun sequencing of total genomic DNA in an Illumina platform. The gene order and organization of the chloroplast genome of T. tetragonolobum exhibits the general structure of flowering plants, and is similar to other Lamiales chloroplast genomes. The chloroplast genome of T. tetragonolobum is a circular molecule of 153,776 base pairs (bp with a quadripartite structure containing two single copy regions, a large single copy region (LSC, 84,612 bp and a small single copy region (SSC, 17,586 bp separated by inverted repeat regions (IRs, 25,789 bp. In addition, the chloroplast genome of T. tetragonolobum has 38.3% GC content and includes 121 genes, of which 86 are protein-coding, 31 are transfer RNA, and four are ribosomal RNA. The chloroplast genome of T. tetragonolobum presents a total of 47 tandem repeats and 347 simple sequence repeats (SSRs with mononucleotides being the most common and di-, tri-, tetra-, and hexanucleotides occurring with less frequency. The results obtained here were compared to other chloroplast genomes of Lamiales available to date, providing new insight into the evolution of chloroplast genomes within Lamiales. Overall, the evolutionary rates of genes in Lamiales are lineage-, locus-, and region-specific, indicating that the evolutionary pattern of nucleotide substitution in chloroplast genomes of flowering

  3. Chloroplastic NADPH oxidase-like activity-mediated perpetual hydrogen peroxide generation in the chloroplast induces apoptotic-like death of Brassica napus leaf protoplasts.

    Science.gov (United States)

    Tewari, Rajesh Kumar; Watanabe, Daisuke; Watanabe, Masami

    2012-01-01

    Despite extensive research over the past years, regeneration from protoplasts has been observed in only a limited number of plant species. Protoplasts undergo complex metabolic modification during their isolation. The isolation of protoplasts induces reactive oxygen species (ROS) generation in Brassica napus leaf protoplasts. The present study was conducted to provide new insight into the mechanism of ROS generation in B. napus leaf protoplasts. In vivo localization of H(2)O(2) and enzymes involved in H(2)O(2) generation and detoxification, molecular antioxidant-ascorbate and its redox state and lipid peroxidation were investigated in the leaf and isolated protoplasts. Incubating leaf strips in the macerating enzyme (ME) for different duration (3, 6, and 12 h) induced accumulation of H(2)O(2) and malondialdehyde (lipid peroxidation, an index of membrane damage) in protoplasts. The level of H(2)O(2) was highest just after protoplast isolation and subsequently decreased during culture. Superoxide generating NADPH oxidase (NOX)-like activity was enhanced, whereas superoxide dismutase (SOD) and ascorbate peroxidase (APX) decreased in the protoplasts compared to leaves. Diaminobenzidine peroxidase (DAB-POD) activity was also lower in the protoplasts compared to leaves. Total ascorbate content, ascorbate to dehydroascorbate ratio (redox state), were enhanced in the protoplasts compared to leaves. Higher activity of NOX-like enzyme and weakening in the activity of antioxidant enzymes (SOD, APX, and DAB-POD) in protoplasts resulted in excessive accumulation of H(2)O(2) in chloroplasts of protoplasts. Chloroplastic NADPH oxidase-like activity mediated perpetual H(2)O(2) generation probably induced apoptotic-like cell death of B. napus leaf protoplasts as indicated by parallel DNA laddering and decreased mitochondrial membrane potential.

  4. Speed of signal transfer in the chloroplast accumulation response.

    Science.gov (United States)

    Tsuboi, Hidenori; Wada, Masamitsu

    2010-05-01

    Chloroplast photorelocation movement is important for plants to perform efficient photosynthesis. Phototropins were identified as blue-light receptors for chloroplast movement in Arabidopsis thaliana and in the fern Adiantum capillus-veneris, whereas neochrome functions as a dual red/blue light receptor in the latter. However, the signal transduction pathways involved in chloroplast movement remain to be clarified. To investigate the kinetic properties of signalling from these photoreceptors to the chloroplasts, we deduced the speed of signal transfer using Adiantum capillus-veneris gametophytes. When a region of dark-adapted gametophyte cells was subjected to microbeam irradiation, chloroplasts moved towards the irradiated area even in subsequent darkness. We therefore recorded the movement and calculated the speeds of signal transfer by time-lapse imaging. Movement speeds under red or blue light were similar, e.g., about 1.0 microm min(-1) in prothallial cells. However, speeds varied according to cell polarity in protonemal cells. The speed of signal transfer from the protonemal apex to the base was approximately 0.7 microm min(-1), but roughly 2.3 microm min(-1) in the opposite direction. The speed of signal transfer in Arabidopsis thaliana mesophyll cells was approximately 0.8 microm min(-1) by comparison. Surprisingly, chloroplasts located farthest away from the microbeam were found to move faster than those in close proximity to the site of irradiation both in Adiantum capillus-veneris and A. thaliana.

  5. Chloroplasts continuously monitor photoreceptor signals during accumulation movement.

    Science.gov (United States)

    Tsuboi, Hidenori; Wada, Masamitsu

    2013-07-01

    Under low light conditions, chloroplasts gather at a cell surface to maximize light absorption for efficient photosynthesis, which is called the accumulation response. Phototropin1 (phot1) and phototropin2 (phot2) were identified as blue light photoreceptors in the accumulation response that occurs in Arabidopsis thaliana and Adiantum capillus-veneris with neochrome1 (neo1) as a red light photoreceptor in A. capillus-veneris. However, the signal molecule that is emitted from the photoreceptors and transmitted to the chloroplasts is not known. To investigate this topic, the accumulation response was induced by partial cell irradiation with a microbeam of red, blue and far-red light in A. capillus-veneris gametophyte cells. Chloroplasts moved towards the irradiated region and were able to sense the signal as long as its signal flowed. The signal from neo1 had a longer life than the signal that came from phototropins. When two microbeams with the same wavelength and the same fluence rate were placed 20 μm apart from each other and were applied to a dark-adapted cell, chloroplasts at an equidistant position always moved towards the center (midpoint) of the two microbeams, but not towards either one. This result indicates that chloroplasts are detecting the concentration of the signal but not the direction of signal flow. Chloroplasts repeatedly move and stop at roughly 10 s intervals during the accumulation response, suggesting that they monitor the intermittent signal waves from photoreceptors.

  6. Expression of human soluble TRAIL in Chlamydomonas reinhardtii chloroplast

    Institute of Scientific and Technical Information of China (English)

    YANG Zongqi; LI yinü; CHEN Feng; LI Dong; ZHANG Zhifang; LIU Yanxin; ZHENG Dexian; WANG Yong; SHEN Guifang

    2006-01-01

    Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces selectively apoptosis in various tumor cells and virus-infected cells, but rarely in normal cells. A chloroplast expression vector, p64TRAIL, containing the cDNA coding for the soluble TRAIL (sTRAIL), was constructed with clpP-trnL-petB-chlL-rpl23-rpl2 as Chlamydomonas reinhardtii plastid homologous recombinant fragments and spectinomycin-resistant aadA gene as a select marker. The plasmid p64TRAIL was transferred into the chloroplast genome of C. reinhardtii by the biolistic method. Three independently transformed lines were obtained by 100 mg/L spectinomycin selection. PCR amplification, Southern blot analysis of the sTRAIL coding region DNA and cultivation cells in the dark all showed that the exogenous DNA had been integrated into chloroplast genome of C. reinhardtii. Western blot analysis showed that human soluble TRAIL was expressed in C. reinhardtii chloroplast. The densitometric analysis of Western blot indicated that the expressed human sTRAIL protein in the chloroplasts of C. reinhardtii accounted for about 0.43%-0.67% of the total soluble proteins.These experimental results demonstrated the possibility of using transgenic chloroplasts of green alga as bioreactors for production of biopharmaceuticals.

  7. Induction of Apoptosis in Purified Nuclei from Tobacco-Suspension Cells by Cytochrome b6/f Complex

    Institute of Scientific and Technical Information of China (English)

    张贵友; 李萍; 朱瑞宇; 田瑞华; 戴尧仁

    2004-01-01

    An apoptotic cell-free system containing cytosol and nuclei from normally cultured tobacco suspension cells was used to show that a spinach chloroplast preparation can induce apoptosis in nuclei,evidenced by DNA electrophoresis and fluorescence microscopy observations.Further study showed that the chloroplast preparation or its pellet (thylakoid membrane) after hypoosmotic or supersonic treatment still exhibited the apoptosis-inducing activity,but the supernatant had no effect,which indicates that the apoptosis-inducing effector in the chloroplast preparation is water-insoluble.The induction of apoptosis by chloroplast preparation could be attenuated by Ac-DEVD-CHO,the specific inhibitor of Caspase-3,implying involvement of a Caspase-3-like protease during the process.Furthermore,extensive apoptosis in nuclei was induced by cytochrome b6/f on the thylakoid membrane,indicating that this important cytochrome complex may have an important role in the chloroplast-related apoptotic pathway.

  8. Ferns, mosses and liverworts as model systems for light-mediated chloroplast movements.

    Science.gov (United States)

    Suetsugu, Noriyuki; Higa, Takeshi; Wada, Masamitsu

    2016-11-17

    Light-induced chloroplast movement is found in most plant species, including algae and land plants. In land plants with multiple small chloroplasts, under weak light conditions, the chloroplasts move towards the light and accumulate on the periclinal cell walls to efficiently perceive light for photosynthesis (the accumulation response). Under strong light conditions, chloroplasts escape from light to avoid photodamage (the avoidance response). In most plant species, blue light induces chloroplast movement, and phototropin receptor kinases are the blue light receptors. Molecular mechanisms for photoreceptors, signal transduction and chloroplast motility systems are being studied using the model plant Arabidopsis thaliana. However, to further understand the molecular mechanisms and evolutionary history of chloroplast movement in green plants, analyses using other plant systems are required. Here, we review recent works on chloroplast movement in green algae, liverwort, mosses and ferns that provide new insights on chloroplast movement.

  9. Regulation of photosynthesis by ion channels in cyanobacteria and higher plants.

    Science.gov (United States)

    Checchetto, Vanessa; Teardo, Enrico; Carraretto, Luca; Formentin, Elide; Bergantino, Elisabetta; Giacometti, Giorgio Mario; Szabo, Ildiko

    2013-12-01

    Photosynthesis converts light energy into chemical energy, and supplies ATP and NADPH for CO2 fixation into carbohydrates and for the synthesis of several compounds which are essential for autotrophic growth. Oxygenic photosynthesis takes place in thylakoid membranes of chloroplasts and photosynthetic prokaryote cyanobacteria. An ancestral photoautotrophic prokaryote related to cyanobacteria has been proposed to give rise to chloroplasts of plants and algae through an endosymbiotic event. Indeed, photosynthetic complexes involved in the electron transport coupled to H(+) translocation and ATP synthesis are similar in higher plants and cyanobacteria. Furthermore, some of the protein and solute/ion conducting machineries also share common structure and function. Electrophysiological and biochemical evidence support the existence of ion channels in the thylakoid membrane in both types of organisms. By allowing specific ion fluxes across thylakoid membranes, ion channels have been hypothesized to either directly or indirectly regulate photosynthesis, by modulating the proton motive force. Recent molecular identification of some of the thylakoid-located channels allowed to obtain genetic proof in favor of such hypothesis. Furthermore, some ion channels of the envelope membrane in chloroplasts have also been shown to impact on this light-driven process. Here we give an overview of thylakoid/chloroplast located ion channels of higher plants and of cyanobacterium Synechocystis sp. PCC 6803. We focus on channels shown to be implicated in the regulation of photosynthesis and discuss the possible mechanisms of action.

  10. Rapid severing and motility of chloroplast-actin filaments are required for the chloroplast avoidance response in Arabidopsis.

    Science.gov (United States)

    Kong, Sam-Geun; Arai, Yoshiyuki; Suetsugu, Noriyuki; Yanagida, Toshio; Wada, Masamitsu

    2013-02-01

    Phototropins (phot1 and phot2 in Arabidopsis thaliana) relay blue light intensity information to the chloroplasts, which move toward weak light (the accumulation response) and away from strong light (the avoidance response). Chloroplast-actin (cp-actin) filaments are vital for mediating these chloroplast photorelocation movements. In this report, we examine in detail the cp-actin filament dynamics by which the chloroplast avoidance response is regulated. Although stochastic dynamics of cortical actin fragments are observed on the chloroplasts, the basic mechanisms underlying the disappearance (including severing and turnover) of the cp-actin filaments are regulated differently from those of cortical actin filaments. phot2 plays a pivotal role in the strong blue light-induced severing and random motility of cp-actin filaments, processes that are therefore essential for asymmetric cp-actin formation for the avoidance response. In addition, phot2 functions in the bundling of cp-actin filaments that is induced by dark incubation. By contrast, the function of phot1 is dispensable for these responses. Our findings suggest that phot2 is the primary photoreceptor involved in the rapid reorganization of cp-actin filaments that allows chloroplasts to change direction rapidly and control the velocity of the avoidance movement according to the light's intensity and position.

  11. CURE-Chloroplast: A chloroplast C-to-U RNA editing predictor for seed plants

    OpenAIRE

    2009-01-01

    Abstract Background RNA editing is a type of post-transcriptional modification of RNA and belongs to the class of mechanisms that contribute to the complexity of transcriptomes. C-to-U RNA editing is commonly observed in plant mitochondria and chloroplasts. The in vivo mechanism of recognizing C-to-U RNA editing sites is still unknown. In recent years, many efforts have been made to computationally predict C-to-U RNA editing sites in the mitochondria of seed plants, but there is still no algo...

  12. Organization of chlorophyll biosynthesis and insertion of chlorophyll into the chlorophyll-binding proteins in chloroplasts.

    Science.gov (United States)

    Wang, Peng; Grimm, Bernhard

    2015-12-01

    Oxygenic photosynthesis requires chlorophyll (Chl) for the absorption of light energy, and charge separation in the reaction center of photosystem I and II, to feed electrons into the photosynthetic electron transfer chain. Chl is bound to different Chl-binding proteins assembled in the core complexes of the two photosystems and their peripheral light-harvesting antenna complexes. The structure of the photosynthetic protein complexes has been elucidated, but mechanisms of their biogenesis are in most instances unknown. These processes involve not only the assembly of interacting proteins, but also the functional integration of pigments and other cofactors. As a precondition for the association of Chl with the Chl-binding proteins in both photosystems, the synthesis of the apoproteins is synchronized with Chl biosynthesis. This review aims to summarize the present knowledge on the posttranslational organization of Chl biosynthesis and current attempts to envision the proceedings of the successive synthesis and integration of Chl into Chl-binding proteins in the thylakoid membrane. Potential auxiliary factors, contributing to the control and organization of Chl biosynthesis and the association of Chl with the Chl-binding proteins during their integration into photosynthetic complexes, are discussed in this review.

  13. Effects of Short-Term Chilling Stress on the Photosystems and Chloroplast Ultrastructure in Sweet Pepper

    Institute of Scientific and Technical Information of China (English)

    LI Xin-guo; BI Yu-ping; ZHAO Shi-jie; MENG Qing-wei; HE Qi-wei; ZOU Qi

    2005-01-01

    By measuring chlorophyll fluorescence parameters, composition of fatty acids, active oxygen species and activities of some antioxidant enzymes, effects of chilling stress (4℃) in the low light (100 μmol m-2 s-1) on chilling-sensitive plants were studied. After 6 h chilling stress (4℃) in the low light, the maximal photochemical efficiency of PSⅡ (Fv/Fm) of sweet pepper leaves decreased by 35.6%, and the oxidizable P700 decreased by 60%. However, chilling stress in the dark had no effect on both of them. Unsaturation of fatty acids in thylakoid membrane was accelerated, which might be helpful to stabilize photosynthetic apparatus. Distortion and swelling of grana caused by chilling in the dark probably decreased activities of antioxidant enzymes, which resulted in the accumulation of active oxygen species. On the contrary,photooxidation might be related to the disintegration and unstacking of grana. Chilling stress induced photoinhibition of PSⅡ and PSⅠ, and active oxygen species might be one of the factors causing the decrease of the oxidizable P700. PSⅠseemed to be more sensitive to chilling stress in the low light than PSⅡ.

  14. Progress in surface and membrane science

    CERN Document Server

    Cadenhead, D A

    1979-01-01

    Progress in Surface and Membrane Science, Volume 12 covers the advances in the study of surface and membrane science. The book discusses the topographical differentiation of the cell surface; the NMR studies of model biological membrane system; and an irreversible thermodynamic approach to energy coupling in mitochondria and chloroplasts. The text also describes water at surfaces; the nature of microemulsions; and the energy principle in the stability of interfaces. Biochemists, physicists, chemical engineers, and people involved in surface and coatings research will find the book invaluable.

  15. Optimization of ATP synthase function in mitochondria and chloroplasts via the adenylate kinase equilibrium

    Directory of Open Access Journals (Sweden)

    Abir U Igamberdiev

    2015-01-01

    Full Text Available The bulk of ATP synthesis in plants is performed by ATP synthase, the main bioenergetics engine of cells, operating both in mitochondria and in chloroplasts. The reaction mechanism of ATP synthase has been studied in detail for over half a century; however, its optimal performance depends also on the steady delivery of ATP synthase substrates and the removal of its products. For mitochondrial ATP synthase, we analyze here the provision of stable conditions for (i the supply of ADP and Mg2+, supported by adenylate kinase (AK equilibrium in the intermembrane space, (ii the supply of phosphate via membrane transporter in symport with H+, and (iii the conditions of outflow of ATP by adenylate transporter carrying out the exchange of free adenylates. We also show that, in chloroplasts, AK equilibrates adenylates and governs Mg2+ contents in the stroma, optimizing ATP synthase and Calvin cycle operation, and affecting the import of inorganic phosphate in exchange with triose phosphates. It is argued that chemiosmosis is not the sole component of ATP synthase performance, which also depends on AK-mediated equilibrium of adenylates and Mg2+, adenylate transport and phosphate release and supply.

  16. Localization of Adenosine Triphosphatase Activity on the Chloroplast Envelope in Tendrils of Pisum sativum1

    Science.gov (United States)

    Sabnis, Dinkar D.; Gordon, Mildred; Galston, Arthur W.

    1970-01-01

    When samples of pea tendril tissue were incubated in the Wachstein-Meisel medium for the demonstration of adenosine triphosphatases, deposits of lead reaction product were localized between the membranes of the chloroplast envelope. The presence of Mg2+ was necessary for adenosine triphosphatase activity, and Ca2+ could not substitute for this requirement. Varying the pH of incubation to 5.5 or 9.4 inhibited enzyme activity, as did the addition of p-chloromercuribenzoic acid or N-ethylmaleimide. The adenosine triphosphatase was apparently inactivated or degraded when the plants were grown in the dark for 24 hours prior to incubation. The enzyme was substrate-specific for adenosine triphosphate; no reaction was obtained with adenosine diphosphate, uridine triphosphate, inosine triphosphate, p-nitrophenyl phosphate, and sodium β-glycerophosphate. Sites of nonspecific depositions of lead are described. The adenosine triphosphatase on the chloroplast envelope may be involved in the light-induced contraction of this organelle. Images PMID:4245003

  17. Preprotein import into chloroplasts via the Toc and Tic complexes is regulated by redox signals in Pisum sativum.

    Science.gov (United States)

    Stengel, Anna; Benz, J Philipp; Buchanan, Bob B; Soll, Jürgen; Bölter, Bettina

    2009-11-01

    The import of nuclear-encoded preproteins is necessary to maintain chloroplast function. The recognition and transfer of most precursor proteins across the chloroplast envelopes are facilitated by two membrane-inserted protein complexes, the translocons of the chloroplast outer and inner envelope (Toc and Tic complexes, respectively). Several signals have been invoked to regulate the import of preproteins. In our study, we were interested in redox-based import regulation mediated by two signals: regulation based on thiols and on the metabolic NADP+/NADPH ratio. We sought to identify the proteins participating in the regulation of these transport pathways and to characterize the preprotein subgroups whose import is redox-dependent. Our results provide evidence that the formation and reduction of disulfide bridges in the Toc receptors and Toc translocation channel have a strong influence on import yield of all tested preproteins that depend on the Toc complex for translocation. Furthermore, the metabolic NADP+/NADPH ratio influences not only the composition of the Tic complex, but also the import efficiency of most, but not all, preproteins tested. Thus, several Tic subcomplexes appear to participate in the translocation of different preprotein subgroups, and the redox-active components of these complexes likely play a role in regulating transport.

  18. Influence of drought stress on leaf anatomical structure and micro-morphology traits and chloroplast ultrastructure of three Malus species%干旱胁迫对3种苹果属植物叶片解剖结构、微形态特征及叶绿体超微结构的影响

    Institute of Scientific and Technical Information of China (English)

    王顺才; 邹养军; 马锋旺

    2014-01-01

    genotypes , compared with well watered plants ,the leaf mesophyll contents and palisade tissue thickness as well as cell tightness rate (CTR) in drought-stressed plants significantly decreased ( P<0 .05 ) ,while the sponge tissue thickness and scattered rate (SR ) significantly in-creased ( P<0 .05 ) .SEM photos indicated that the stomatal density in young leaves of drought-stressed plants increased ( P<0 .05 ) ,while the stomata width ,stomatal opening rate ,and the relative opening degree of stomata decreased .TEM analysis showed that the upper and lower cuticle thickness of M . prunifolia and M . sieversii increased ,whereas those of M . hupehensis increased first and then decreased with prolonged drought treatment ,swelling chloroplast ,decreased the number of larger starch granules ,and loosing grana and thylakoid were the typical leaf ultrastructure for medium water stress .Under severe water deficit ,the chloroplasts were round in a shape ,with more damaged structure of membranes , and an extensive vacuolization and disorganization of thylakoid . However , the degree of drought-induced damage was smaller in M . prunifolia and M . sieversii plants as compared to M . hupehensis plants ,duo to the ability to maintain cell integrity .

  19. Red light, Phot1 and JAC1 modulate Phot2-dependent reorganization of chloroplast actin filaments and chloroplast avoidance movement.

    Science.gov (United States)

    Ichikawa, Satoshi; Yamada, Noboru; Suetsugu, Noriyuki; Wada, Masamitsu; Kadota, Akeo

    2011-08-01

    The phototropin (phot)-dependent intracellular relocation of chloroplasts is a ubiquitous phenomenon in plants. We have previously revealed the involvement of a short cp-actin (chloroplast actin) filament-based mechanism in this movement. Here, the reorganization of cp-actin filaments during the avoidance movement of chloroplasts was analyzed in higher time resolution under blue GFP (green fluorescent protein) excitation light in an actin filament-visualized line of Arabidopsis thaliana. Under standard background red light of 89 μmol m(-2) s(-1), cp-actin filaments transiently disappeared at approximately 30 s and reappeared in a biased configuration on chloroplasts approximately 70 s after blue excitation light irradiation. The timing of biased cp-actin reappearance was delayed under the background of strong red light or in the absence of red light. Consistently, chloroplast movement was delayed under these conditions. In phot1 mutants, acceleration of both the disappearance and reappearance of cp-actin filaments occurred, indicating an inhibitory action of phot1 on reorganization of cp-actin filaments. Avoidance movements began sooner in phot1 than in wild-type plants. No reorganization of cp-actin filaments was seen in phot2 or phot1phot2 mutants lacking phot2, which is responsible for avoidance movements. Surprisingly, jac1 (j-domain protein required for chloroplast accumulation response 1) mutants, lacking the accumulation response, showed no avoidance movements under the whole-cell irradiation condition for GFP observation. Cp-actin filaments in jac1 did not show a biased distribution, with a small or almost no transient decrease in the number. These results indicate a close association between the biased distribution of cp-actin filaments and chloroplast movement. Further, JAC1 is suggested to function in the biased cp-actin filament distribution by regulating their appearance and disappearance.

  20. Production and scavenging of reactive oxygen species in photosynthesis of chloroplasts%叶绿体光合代谢中活性氧的产生与清除

    Institute of Scientific and Technical Information of China (English)

    张有福; 陈春艳; 孙会忠; 陈应武

    2011-01-01

    有氧代谢不可避免产生活性氧(ROS),叶绿体的PSⅠ和PSⅡ反应中心均是ROS产生的主要位点.叶绿体产生的ROS主要有超氧阴离子(O2-)、过氧化氢(H2O2)、羟自由基(·OH)和单线氧(1O1),其中在PSI产生的O2-将进一步产生H2O2和·OH,而1O2产生在PSⅡ.正常生理代谢条件下,叶绿体内抗氧化系统和光能吸收利用的调节保持活性氧产生和消灭的平衡,不会影响植物的正常生理功能.%Aerobic metabolism inevitably products the reactive oxygen species ( ROS) in chloroplasts , and the reaction centers of PSⅠ and PSⅡ in chloroplast thylakoids are the major generation sites of ROS. ROS from chloroplasts mainly include superoxide anion ( O2- ) , hydrogen peroxide ( H2O2) , singlet state oxygen (1O2) and hydroxy radical ( · OH) . Both · OH and H2O2origin from O2- in PSⅠ, however, 1O2generate in PSⅡ. In normal physiological conditions, the production and scavenging of ROS maintains balance, and will not affect the plant's physiological function by regulating antioxidant system, absorption and elimination of light.

  1. Chloroplast DNA Diversity of Oak Species in Eastern Romania

    Directory of Open Access Journals (Sweden)

    Ioan Calin MOLDOVAN

    2010-12-01

    Full Text Available The chloroplast DNA of 34 sessile oak (Quercus petraea and 27 pedunculate oak (Q. robur populations covering the entire natural distribution of the two oak species in Eastern Romania was investigated using four large regions of the chloroplast genome by PCR and RFLP technique. A total of seven chloroplast DNA haplotypes sensu lato have been observed by analysing 305 mature trees. However, due to the high resolution of the electrophoresis method a total of 22 chloroplast variants could have been detected, with new mutations and fragment combinations in two of the amplified regions: psbC/trnD and trnT/trnF. All of the haplotypes belong to the phylogenetic lineages A and E, which originate from the Balkan Peninsula. Most of genetic diversity is distributed among populations (GST=0.779. The chloroplast DNA haplotypes are shared by the two oak species. Different dispersal abilities may explain the higher value of genetic differentiation among populations in sessile oak than in pedunculate oak.

  2. Comparison of intraspecific, interspecific and intergeneric chloroplast diversity in Cycads

    Science.gov (United States)

    Jiang, Guo-Feng; Hinsinger, Damien Daniel; Strijk, Joeri Sergej

    2016-01-01

    Cycads are among the most threatened plant species. Increasing the availability of genomic information by adding whole chloroplast data is a fundamental step in supporting phylogenetic studies and conservation efforts. Here, we assemble a dataset encompassing three taxonomic levels in cycads, including ten genera, three species in the genus Cycas and two individuals of C. debaoensis. Repeated sequences, SSRs and variations of the chloroplast were analyzed at the intraspecific, interspecific and intergeneric scale, and using our sequence data, we reconstruct a phylogenomic tree for cycads. The chloroplast was 162,094 bp in length, with 133 genes annotated, including 87 protein-coding, 37 tRNA and 8 rRNA genes. We found 7 repeated sequences and 39 SSRs. Seven loci showed promising levels of variations for application in DNA-barcoding. The chloroplast phylogeny confirmed the division of Cycadales in two suborders, each of them being monophyletic, revealing a contradiction with the current family circumscription and its evolution. Finally, 10 intraspecific SNPs were found. Our results showed that despite the extremely restricted distribution range of C. debaoensis, using complete chloroplast data is useful not only in intraspecific studies, but also to improve our understanding of cycad evolution and in defining conservation strategies for this emblematic group. PMID:27558458

  3. Arabidopsis chloroplast chaperonin 10 is a calmodulin-binding protein

    Science.gov (United States)

    Yang, T.; Poovaiah, B. W.

    2000-01-01

    Calcium regulates diverse cellular activities in plants through the action of calmodulin (CaM). By using (35)S-labeled CaM to screen an Arabidopsis seedling cDNA expression library, a cDNA designated as AtCh-CPN10 (Arabidopsis thaliana chloroplast chaperonin 10) was cloned. Chloroplast CPN10, a nuclear-encoded protein, is a functional homolog of E. coli GroES. It is believed that CPN60 and CPN10 are involved in the assembly of Rubisco, a key enzyme involved in the photosynthetic pathway. Northern analysis revealed that AtCh-CPN10 is highly expressed in green tissues. The recombinant AtCh-CPN10 binds to CaM in a calcium-dependent manner. Deletion mutants revealed that there is only one CaM-binding site in the last 31 amino acids of the AtCh-CPN10 at the C-terminal end. The CaM-binding region in AtCh-CPN10 has higher homology to other chloroplast CPN10s in comparison to GroES and mitochondrial CPN10s, suggesting that CaM may only bind to chloroplast CPN10s. Furthermore, the results also suggest that the calcium/CaM messenger system is involved in regulating Rubisco assembly in the chloroplast, thereby influencing photosynthesis. Copyright 2000 Academic Press.

  4. Two types of chloroplast gene promoters in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Klein, U; De Camp, J D; Bogorad, L

    1992-04-15

    Structures of the promoters of Chlamydomonas reinhardtii plastid atpB and 16S rRNA-encoding genes were analyzed in vivo. Chimeric constructs, containing the Chlamydomonas chloroplast atpB or 16S rRNA-encoding gene promoter coupled to the Escherichia coli uidA (beta-glucuronidase, GUS) reporter gene and bordered by C. reinhardtii chloroplast sequences, were stably introduced into the chloroplast of Chlamydomonas by microprojectile bombardment. Activity of the promoters in the chloroplast of GUS gene-positive transformants was assayed by measuring the abundance of GUS transcripts and determining the relative rates of GUS transcription in vivo. Deletion analyses of the 16S rRNA gene and atpB promoter fragments showed that the two promoters differ structurally. The 16S rRNA gene promoter resembles the bacterial sigma 70 type with typical -10 and -35 elements. The atpB promoter, on the other hand, lacks a conserved motif in the -35 region but contains, in the -10 region, a characteristic octameric palindrome (TATAATAT) that is conserved in the promoter sequences of some other C. reinhardtii chloroplast genes. For maximum activity, the atpB promoter requires sequences of approximately 22 base pairs upstream and approximately 60 base pairs downstream of the transcription start site.

  5. Oryza sativa Chloroplast Signal Recognition Particle 43 (OscpSRP43) Is Required for Chloroplast Development and Photosynthesis.

    Science.gov (United States)

    Lv, Xiang-guang; Shi, Yong-feng; Xu, Xia; Wei, Yan-lin; Wang, Hui-mei; Zhang, Xiao-bo; Wu, Jian-li

    2015-01-01

    A rice chlorophyll-deficient mutant w67 was isolated from an ethyl methane sulfonate (EMS)-induced IR64 (Oryza sativa L. ssp. indica) mutant bank. The mutant exhibited a distinct yellow-green leaf phenotype in the whole plant growth duration with significantly reduced levels of chlorophyll and carotenoid, impaired chloroplast development and lowered capacity of photosynthesis compared with the wild-type IR64. Expression of a number of genes associated with chlorophyll metabolism, chloroplast biogenesis and photosynthesis was significantly altered in the mutant. Genetic analysis indicated that the yellow-green phenotype was controlled by a single recessive nuclear gene located on the short arm of chromosome 3. Using map-based strategy, the mutation was isolated and predicted to encode a chloroplast signal recognition particle 43 KD protein (cpSRP43) with 388 amino acid residuals. A single base substitution from A to T at position 160 resulted in a premature stop codon. OscpSRP43 was constitutively expressed in various organs with the highest level in the leaf. Functional complementation could rescue the mutant phenotype and subcellular localization showed that the cpSRP43:GFP fusion protein was targeted to the chloroplast. The data suggested that Oryza sativa cpSRP43 (OscpSRP43) was required for the normal development of chloroplasts and photosynthesis in rice.

  6. Oryza sativa Chloroplast Signal Recognition Particle 43 (OscpSRP43 Is Required for Chloroplast Development and Photosynthesis.

    Directory of Open Access Journals (Sweden)

    Xiang-guang Lv

    Full Text Available A rice chlorophyll-deficient mutant w67 was isolated from an ethyl methane sulfonate (EMS-induced IR64 (Oryza sativa L. ssp. indica mutant bank. The mutant exhibited a distinct yellow-green leaf phenotype in the whole plant growth duration with significantly reduced levels of chlorophyll and carotenoid, impaired chloroplast development and lowered capacity of photosynthesis compared with the wild-type IR64. Expression of a number of genes associated with chlorophyll metabolism, chloroplast biogenesis and photosynthesis was significantly altered in the mutant. Genetic analysis indicated that the yellow-green phenotype was controlled by a single recessive nuclear gene located on the short arm of chromosome 3. Using map-based strategy, the mutation was isolated and predicted to encode a chloroplast signal recognition particle 43 KD protein (cpSRP43 with 388 amino acid residuals. A single base substitution from A to T at position 160 resulted in a premature stop codon. OscpSRP43 was constitutively expressed in various organs with the highest level in the leaf. Functional complementation could rescue the mutant phenotype and subcellular localization showed that the cpSRP43:GFP fusion protein was targeted to the chloroplast. The data suggested that Oryza sativa cpSRP43 (OscpSRP43 was required for the normal development of chloroplasts and photosynthesis in rice.

  7. Chloroplastic and stomatal aspects of ozone-induced reduction of net photosynthesis in plants

    Energy Technology Data Exchange (ETDEWEB)

    Torsethaugen, Gro

    1998-09-01

    The present thesis relates to ozone-induced reduction of photosynthesis in plants. As a photochemical oxidant O{sub 3} is formed by the interaction of hydrocarbons, nitrogen oxides and oxygen in sunlight. Ozone (O{sub 3}) is the most phytotoxic of all the air pollutants and is known to reduce plant growth and net photosynthesis, cause stomatal closure, induce visible injury, accelerate senescence and induce or inhibit transcription of a variety of genes with a corresponding increase/decrease in protein products. The underlying cellular mechanisms for many of these changes are unknown. Following fields are investigated: Ozone-induced reduction of net photosynthesis; ozone and the photosynthetic apparatus in the chloroplasts; ozone and stomata; ozone effects on plant membranes; protection against ozone injury in plants. 249 refs., 22 figs., 4 tabs.

  8. Extending the biosynthetic repertoires of cyanobacteria and chloroplasts

    DEFF Research Database (Denmark)

    Nielsen, Agnieszka Janina Zygadlo; Mellor, Silas Busck; Vavitsas, Konstantinos

    2016-01-01

    The chloroplasts found in plants and algae, and photosynthetic microorganisms such as cyanobacteria, are emerging hosts for sustainable production of valuable biochemicals, using only inorganic nutrients, water, CO2 and light as inputs. In the past decade, many bioengineering efforts have focused...... on metabolic engineering and synthetic biology in the chloroplast or in cyanobacteria for the production of fuels, chemicals, as well as complex, high-value bioactive molecules. Biosynthesis of all these compounds can be performed in photosynthetic organelles/organisms by heterologous expression...... of chloroplasts and cyanobacteria as biosynthetic compartments and hosts, and we estimate the production levels to be expected from photosynthetic hosts in light of the fraction of electrons and carbon that can potentially be diverted from photosynthesis. The supply of reducing power, in the form of electrons...

  9. X-ray absorption spectroscopy and EPR studies of oriented spinach thylakoid preparations

    Energy Technology Data Exchange (ETDEWEB)

    Andrews, J.C. [Univ. of California, Berkeley, CA (United States). Dept. of Chemistry]|[Lawrence Berkeley Lab., CA (United States). Structural Biology Div.

    1995-08-01

    In this study, oriented Photosystem II (PS II) particles from spinach chloroplasts are studied with electron paramagnetic resonance (EPR) and x-ray absorption spectroscopy (XAS) to determine more details of the structure of the oxygen evolving complex (OEC). The nature of halide binding to Mn is also studied with Cl K-edge and Mn EXAFS (extended x-ray absorption fine structure) of Mn-Cl model compounds, and with Mn EXAFS of oriented PS II in which Br has replaced Cl. Attention is focused on the following: photosynthesis and the oxygen evolving complex; determination of mosaic spread in oriented photosystem II particles from signal II EPR measurement; oriented EXAFS--studies of PS II in the S{sub 2} state; structural changes in PS II as a result of treatment with ammonia: EPR and XAS studies; studies of halide binding to Mn: Cl K-edge and Mn EXAFS of Mn-Cl model compounds and Mn EXAFS of oriented Br-treated photosystem II.

  10. Mutation of a chloroplast-targeting signal in Alternanthera mosaic virus TGB3 impairs cell-to-cell movement and eliminates long-distance virus movement.

    Science.gov (United States)

    Lim, Hyoun-Sub; Vaira, Anna Maria; Bae, Hanhong; Bragg, Jennifer N; Ruzin, Steven E; Bauchan, Gary R; Dienelt, Margaret M; Owens, Robert A; Hammond, John

    2010-08-01

    Cell-to-cell movement of potexviruses requires coordinated action of the coat protein and triple gene block (TGB) proteins. The structural properties of Alternanthera mosaic virus (AltMV) TGB3 were examined by methods differentiating between signal peptides and transmembrane domains, and its subcellular localization was studied by Agrobacterium-mediated transient expression and confocal microscopy. Unlike potato virus X (PVX) TGB3, AltMV TGB3 was not associated with the endoplasmic reticulum, and accumulated preferentially in mesophyll cells. Deletion and site-specific mutagenesis revealed an internal signal VL(17,18) of TGB3 essential for chloroplast localization, and either deletion of the TGB3 start codon or alteration of the chloroplast-localization signal limited cell-to-cell movement to the epidermis, yielding a virus that was unable to move into the mesophyll layer. Overexpression of AltMV TGB3 from either AltMV or PVX infectious clones resulted in veinal necrosis and vesiculation at the chloroplast membrane, a cytopathology not observed in wild-type infections. The distinctive mesophyll and chloroplast localization of AltMV TGB3 highlights the critical role played by mesophyll targeting in virus long-distance movement within plants.

  11. [Response of reactive oxygen metabolism in melon chloroplasts to short-term salinity-alkalinity stress regulated by exogenous γ-aminobutyric acid].

    Science.gov (United States)

    Xiang, Li-xia; Hu, Li-pan; Hu, Xiao-hui; Pan, Xiong-bo; Ren, Wen-qi

    2015-12-01

    The regulatory effect of exogenous γ-aminobutyric acid (GABA) on metabolism of reactive oxygen species (ROS) in melon chloroplasts under short-term salinity-alkalinity stress were investigated in melon variety 'Jinhui No. 1', which was cultured with deep flow hydroponics. The result showed that under salinity-alkalinity stress, the photosynthetic pigment content, MDA content, superoxide anion (O₂·) production rate and hydrogen peroxide (H₂O₂) content in chloroplast increased significantly, the contents of antioxidants ascorbic acid (AsA) and glutathione (GSH) increased, and the activities of H⁺-ATPase and H⁺-PPiase were inhibited obviously. With exogenous GABA application, the accumulations of O₂·, MDA and H₂O₂ induced by salinity-alkalinity stress were inhibited. Exogenous GABA alleviated the increase of photosynthetic pigment content, improved the activity of SOD, enzymes of AsA-GSH cycle, total AsA and total GSH while decreased the AsA/DHA ratio and GSH/GSSH ratio. Foliar GABA could enhance the H⁺-ATPase and H⁺-PPiase activities. Our results suggested that the exogenous GABA could accelerate the ROS metabolism in chloroplast, promote the recycle of AsA-GSH, and maintain the permeability of cell membrane to improve the ability of melon chloroplast against salinity-alkalinity stress.

  12. Actin-dependence of the chloroplast cold positioning response in the liverwort Marchantia polymorpha L.

    Directory of Open Access Journals (Sweden)

    Shun Kimura

    2016-09-01

    Full Text Available The subcellular positioning of chloroplasts can be changed by alterations in the environment such as light and temperature. For example, in leaf mesophyll cells, chloroplasts localize along anticlinal cell walls under high-intensity light, and along periclinal cell walls under low-intensity light. These types of positioning responses are involved in photosynthetic optimization. In light-mediated chloroplast positioning responses, chloroplasts move to the appropriate positions in an actin-dependent manner, although some exceptions also depend on microtubule. Even under low-intensity light, at low temperature (e.g., 5°C, chloroplasts localize along anticlinal cell walls; this phenomenon is termed chloroplast cold positioning. In this study, we analyzed whether chloroplast cold positioning is dependent on actin filaments and/or microtubules in the liverwort Marchantia polymorpha L. When liverwort cells were treated with drugs for the de-polymerization of actin filaments, chloroplast cold positioning was completely inhibited. In contrast, chloroplast cold positioning was not affected by treatment with a drug for the de-polymerization of microtubules. These observations indicate the actin-dependence of chloroplast cold positioning in M. polymorpha. Actin filaments during the chloroplast cold positioning response were visualized by using fluorescent probes based on fluorescent proteins in living liverwort cells, and thus, their behavior during the chloroplast cold positioning response was documented.

  13. Possible association of actin filaments with chloroplasts of spinach mesophyll cells in vivo and in vitro.

    Science.gov (United States)

    Kumatani, T; Sakurai-Ozato, N; Miyawaki, N; Yokota, E; Shimmen, T; Terashima, I; Takagi, S

    2006-11-01

    In palisade mesophyll cells of spinach (Spinacia oleracea L.) kept under low-intensity white light, chloroplasts were apparently immobile and seemed to be surrounded by fine bundles of actin filaments. High-intensity blue light induced actin-dependent chloroplast movement concomitant with the appearance of a couple of long, straight bundles of actin filaments in each cell, whereas high-intensity red light was essentially ineffective in inducing these responses. The actin organization observed under low-intensity white light has been postulated to function in anchoring chloroplasts at proper intracellular positions through direct interaction with the chloroplasts. Intact chloroplasts, which retained their outer envelopes, were isolated after homogenization of leaves and Percoll centrifugation. No endogenous actin was detected by immunoblotting in the final intact-chloroplast fraction prepared from the leaves kept under low-intensity white light or in darkness. In cosedimentation assays with exogenously added skeletal muscle filamentous actin, however, actin was detected in the intact-chloroplast fraction precipitated after low-speed centrifugation. The association of actin with chloroplasts was apparently dependent on incubation time and chloroplast density. After partial disruption of the outer envelope of isolated chloroplasts by treatment with trypsin, actin was no longer coprecipitated. The results suggest that chloroplasts in spinach leaves can directly interact with actin, and that this interaction may be involved in the regulation of intracellular positioning of chloroplasts.

  14. Robust expression of a bioactive mammalian protein in Chlamydomonas chloroplast

    Energy Technology Data Exchange (ETDEWEB)

    Mayfield, Stephen P

    2015-01-13

    Methods and compositions are disclosed to engineer chloroplast comprising heterologous mammalian genes via a direct replacement of chloroplast Photosystem II (PSII) reaction center protein coding regions to achieve expression of recombinant protein above 5% of total protein. When algae is used, algal expressed protein is produced predominantly as a soluble protein where the functional activity of the peptide is intact. As the host algae is edible, production of biologics in this organism for oral delivery of proteins/peptides, especially gut active proteins, without purification is disclosed.

  15. The chloroplast-to-chromoplast transition in tomato fruit

    OpenAIRE

    Bian, Wanping

    2012-01-01

    L'un des phénomènes les plus importants survenus pendant la maturation du fruit de tomate est le changement de couleur du vert au rouge. Ce changement a lieu dans les plastes et correspond à la différenciation des plastes photosynthétiques, les chloroplastes, en plastes non-photosynthétiques qui accumulent des caroténoïdes, les chromoplastes. Dans cette thèse, nous présentons d'abord une introduction bibliographique sur le domaine de la transition chloroplaste-chromoplaste, en décrivant les m...

  16. Photosynthetic electron transport inhibition by 2-substituted 4-alkyl-6-benzylamino-1,3,5-triazines with thylakoids from wild- type and atrazine-resistant Chenopodium album

    NARCIS (Netherlands)

    Okano, R.; Ohki, A.; Ohki, S.; Kohno, H.; Rensen, van J.J.S.; Böger, P.; Wakabayashi, K.

    2002-01-01

    The effect of 2-benzylamino-1,3,5-triazines on photosynthetic electron transport (PET) was measured with thylakoids isolated from atrazine-resistant, wild-type Chenopodium album, and spinach to find novel 1,3,5-triazine herbicides bearing a strong PET inhibition. The PET inhibition assay with Chenop

  17. Age dependent alterations in photosystem II acceptor side in Cucumis sativus cotyledonary leaf thylakoids: analysis of binding characteristics of herbicide [14C]-atrazine.

    Science.gov (United States)

    Prakash, J S; Baig, M A; Mohanty, P

    1999-02-01

    Senescence induced temporal changes in photosystems can be conveniently studied in cotyledonary leaves. We monitored the protein, chlorophyll and electron transport activities in Cucumis sativus cv Poinsette cotyledonary leaves and observed that by 20th day, there was a 50%, 41% and 30-33% decline in the chlorophyll, protein and photosystem II activity respectively when compared to 6th day cotyledonary leaves taken as control. We investigated the changes in photosystem II activity (O2 evolution) as a function of light intensity. The photosystem II functional antenna decreased by 27% and the functional photosystem II units decreased by 30% in 20-day old cotyledonary leaf thylakoids. The herbicide [14C]-atrazine binding assay to monitor specific binding of the herbicide to the acceptor side of photosystem II reaction centre protein, D1, showed an increase in the affinity for atrazine towards D1 protein and decrease in the QB binding sites in 20th day leaf thylakoids when compared to 6th day leaf thylakoids. The western blot analysis also suggested a decrease in steady state levels of D1 protein in 20th day cotyledonary leaf thylakoids as compared to 6th day sample which is in agreement with [14C]-atrazine binding assay and light saturation kinetics.

  18. Myosin inhibitors block accumulation movement of chloroplasts in Arabidopsis thaliana leaf cells.

    Science.gov (United States)

    Paves, H; Truve, E

    2007-01-01

    Chloroplasts alter their distribution within plant cells depending on the external light conditions. Myosin inhibitors 2,3-butanedione monoxime (BDM), N-ethylmaleimide (NEM), and 1-(5-iodonaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine hydrochloride (ML-7) were used to study the possible role of myosins in chloroplast photorelocation in Arabidopsis thaliana mesophyll cells. None of these agents had an effect on the chloroplast high-fluence-rate avoidance movement but all of the three myosin inhibitors blocked the accumulation movement of chloroplasts after a high-fluence-rate irradiation of the leaves. The results suggest that myosins have a role in A. thaliana chloroplast photorelocation.

  19. Isolation of dimorphic chloroplasts from the single-cell C4 species Bienertia sinuspersici

    Directory of Open Access Journals (Sweden)

    Lung Shiu-Cheung

    2012-03-01

    Full Text Available Abstract Three terrestrial plants are known to perform C4 photosynthesis without the dual-cell system by partitioning two distinct types of chloroplasts in separate cytoplasmic compartments. We report herein a protocol for isolating the dimorphic chloroplasts from Bienertia sinuspersici. Hypo-osmotically lysed protoplasts under our defined conditions released intact compartments containing the central chloroplasts and intact vacuoles with adhering peripheral chloroplasts. Following Percoll step gradient purification both chloroplast preparations demonstrated high homogeneities as evaluated from the relative abundance of respective protein markers. This protocol will open novel research directions toward understanding the mechanism of single-cell C4 photosynthesis.

  20. Complete Chloroplast Genome Sequence of Dendrobium nobile from Northeastern India

    Science.gov (United States)

    Parameswaran, Sriram; Sundar, Durai

    2016-01-01

    The orchid species Dendrobium nobile belonging to the family Orchidaceae and genus Dendrobium (a vast genus that encompasses nearly 1,200 species) has an herbal medicinal history of about 2000 years in east and south Asian countries. Here, we report the complete chloroplast genome sequence of D. nobile from northeastern India for the first time.

  1. Chloroplast DNA phylogeography and cytotype geography in autopolyploid Plantago media

    NARCIS (Netherlands)

    Van Dijk, P.J.; Bakx-Schotman, Tanja

    1997-01-01

    In order to gain insight into the causes of parapatric diploid and tetraploid distributions in Plantago media chloroplast DNA (cpDNA) restriction site polymorphism was studied in 36 European populations. Parapatric distributions are often explained by adaptive differences between cytotypes to an und

  2. Protein disorder in plants: a view from the chloroplast

    Directory of Open Access Journals (Sweden)

    Yruela Inmaculada

    2012-09-01

    Full Text Available Abstract Background The intrinsically unstructured state of some proteins, observed in all living organisms, is essential for basic cellular functions. In this field the available information from plants is limited but it has been reached a point where these proteins can be comprehensively classified on the basis of disorder, function and evolution. Results Our analysis of plant genomes confirms that nuclear-encoded proteins follow the same trend than other multi-cellular eukaryotes; however, chloroplast- and mitochondria- encoded proteins conserve the patterns of Archaea and Bacteria, in agreement with their phylogenetic origin. Based on current knowledge about gene transference from the chloroplast to the nucleus, we report a strong correlation between the rate of disorder of transferred and nuclear-encoded proteins, even for polypeptides that play functional roles back in the chloroplast. We further investigate this trend by reviewing the set of chloroplast ribosomal proteins, one of the most representative transferred gene clusters, finding that the ribosomal large subunit, assembled from a majority of nuclear-encoded proteins, is clearly more unstructured than the small one, which integrates mostly plastid-encoded proteins. Conclusions Our observations suggest that the evolutionary dynamics of the plant nucleus adds disordered segments to genes alike, regardless of their origin, with the notable exception of proteins currently encoded in both genomes, probably due to functional constraints.

  3. Mitochondrial and chloroplast DNA based phylogeny of Pelargonium (Geraniaceae)

    NARCIS (Netherlands)

    Bakker, F.T.; Culham, A.; Pankhurst, C.E.; Gibby, M.

    2000-01-01

    Overall phylogenetic relationships within the genus Pelargonium (Geraniaceae) were inferred based on DNA sequences from mitochondrial(mt)-encoded nad1 b/c exons and from chloroplast(cp)-encoded trnL (UAA) 5' exon-trnF (GAA) exon regions using two species of Geranium and Sarcocaulon vanderetiae as ou

  4. Functional characterization of the chloroplast ferric chelate oxidoreductase enzyme.

    Science.gov (United States)

    Solti, Adám; Müller, Brigitta; Czech, Viktória; Sárvári, Éva; Fodor, Ferenc

    2014-05-01

    Iron (Fe) has an essential role in the biosynthesis of chlorophylls and redox cofactors, and thus chloroplast iron uptake is a process of special importance. The chloroplast ferric chelate oxidoreductase (cFRO) has a crucial role in this process but it is poorly characterized. To study the localization and mechanism of action of cFRO, sugar beet (Beta vulgaris cv Orbis) chloroplast envelope fractions were isolated by gradient ultracentrifugation, and their purity was tested by western blotting against different marker proteins. The ferric chelate reductase (FCR) activity of envelope fractions was studied in the presence of NAD(P)H (reductants) and FAD coenzymes. Reduction of Fe(III)-ethylenediaminetetraacetic acid was monitored spectrophotometrically by the Fe(II)-bathophenanthroline disulfonate complex formation. FCR activity, that is production of free Fe(II) for Fe uptake, showed biphasic saturation kinetics, and was clearly associated only to chloroplast inner envelope (cIE) vesicles. The reaction rate was > 2.5 times higher with NADPH than with NADH, which indicates the natural coenzyme preference of cFRO activity and its dependence on photosynthesis. FCR activity of cIE vesicles isolated from Fe-deficient plants also showed clear biphasic kinetics, where the KM of the low affinity component was elevated, and thus this component was down-regulated.

  5. Chloroplast EF-Tu and thermal aggregation of Rubisco activase

    Science.gov (United States)

    Chloroplast protein synthesis elongation factor, EF-Tu, has been implicated in heat tolerance in maize. The recombinant precursor of this protein, pre-EF-Tu, has been found to exhibit chaperone activity and protect heat-labile proteins, such as citrate synthase and malate dehydrogenase, from therma...

  6. Chloroplast microsatellite markers for Artocarpus (Moraceae) developed from transcriptome sequences

    Science.gov (United States)

    Premise of the study: Chloroplast microsatellite loci were characterized from transcriptomes of Artocarpus (A.) altilis (breadfruit) and A. camansi (breadnut). They were tested in A. odoratissimus (terap) and A. altilis and evaluated in silico for two congeners. Methods and Results: 15 simple seque...

  7. Distribution pattern changes of actin filaments during chloroplast movement in Adiantum capillus-veneris.

    Science.gov (United States)

    Tsuboi, Hidenori; Wada, Masamitsu

    2012-05-01

    Chloroplasts change their positions in a cell in response to light intensities. The photoreceptors involved in chloroplast photo-relocation movements and the behavior of chloroplasts during their migration were identified in our previous studies, but the mechanism of movement has yet to be clarified. In this study, the behavior of actin filaments under various light conditions was observed in Adiantum capillus-veneris gametophytes. In chloroplasts staying in one place under a weak light condition and not moving, circular structures composed of actin filaments were observed around the chloroplast periphery. In contrast, short actin filaments were observed at the leading edge of moving chloroplasts induced by partial cell irradiation. In the dark, the circular structures found under the weak light condition disappeared and then reappeared around the moving chloroplasts. Mutant analyses revealed that the disappearance of the circular actin structure was mediated by the blue light photoreceptor, phototropin2.

  8. Chloroplast movement: dissection of events downstream of photo- and mechano-perception.

    Science.gov (United States)

    Sato, Yoshikatsu; Kadota, Akeo; Wada, Masamitsu

    2003-02-01

    The study of chloroplast photorelocation movement is progressing rapidly now that mutants for chloroplast movement have become available in Arabidopsis thaliana. However, mechanistic approaches in cell biology still stand to elucidate the mechanisms and regulations of such movement. The fern Adiantum capillus-veneris and the moss Physcomitrella patens are particularly suitable materials for analyzing the kinetics of intracellular chloroplast movement. In these plants, chloroplast movement is induced by red light as well as blue light, mediated by phytochrome and blue light receptor, respectively. In this paper, we review the unique force-generating system for chloroplast motility in P. patens. In addition to light-induced chloroplast movement, we also summarize mechanically induced chloroplast movement in these plants and the motility systems involved. Finally, the different dependency of mechano- and photo-relocation movement on external Ca(2+) is discussed.

  9. Differential positioning of chloroplasts in C4 mesophyll and bundle sheath cells.

    Science.gov (United States)

    Maai, Eri; Miyake, Hiroshi; Taniguchi, Mitsutaka

    2011-08-01

    Chloroplast photorelocation movement is extensively studied in C3 but not C4 plants. C4 plants have 2 types of photosynthetic cells: mesophyll and bundle sheath cells. Mesophyll chloroplasts are randomly distributed along cell walls, whereas bundle sheath chloroplasts are located close to the vascular tissues or mesophyll cells depending on the plant species. The cell-specific C 4 chloroplast arrangement is established during cell maturation, and is maintained throughout the life of the cell. However, only mesophyll chloroplasts can change their positions in response to environmental stresses. The migration pattern is unique to C4 plants and differs from that of C3 chloroplasts. In this mini-review, we highlight the cell-specific disposition of chloroplasts in C4 plants and discuss the possible physiological significances.

  10. Evolution of the Cp-Actin-based Motility System of Chloroplasts in Green Plants.

    Science.gov (United States)

    Suetsugu, Noriyuki; Wada, Masamitsu

    2016-01-01

    During the course of green plant evolution, numerous light responses have arisen that optimize their growth under fluctuating light conditions. The blue light receptor phototropin mediates several photomovement responses at the tissue, cellular and organelle levels. Chloroplast photorelocation movement is one such photomovement response, and is found not only in most green plants, but also in some red algae and photosynthetic stramenopiles. In general, chloroplasts move toward weak light to maximally capture photosynthetically active radiation (the chloroplast accumulation response), and they move away from strong light to avoid photodamage (the avoidance response). In land plants, chloroplast movement is dependent on specialized actin filaments, chloroplast-actin filaments (cp-actin filaments). Through molecular genetic analysis using Arabidopsis thaliana, many molecular factors that regulate chloroplast photorelocation were identified. In this Perspective, we discuss the evolutionary history of the molecular mechanism for chloroplast photorelocation movement in green plants in view of cp-actin filaments.

  11. Broad host range plasmid-based gene transfer system in the cyanobacterium Gloeobacter violaceus which lacks thylakoids

    Institute of Scientific and Technical Information of China (English)

    GUO Haitao; XU Xudong

    2004-01-01

    Gloeobacter violaceus, a cyanobacterium lack of thylakoids, is refractory to genetic manipulations because its cells are enveloped by a thick gelatinous sheath and in colonial form.In this study, a large number of single cells were obtained by repeated pumping with a syringe with the gelatinous sheath removed.And an exogenous broad host range plasmid pKT210 was conjugatively transferred into G.violaceus.Analyses with dot-blot hybridization and restriction mapping showed that the exogenous plasmid pKT210 had been introduced into G.violaceus and stably maintained with no alteration in its structure.pKT210 extracted from G.violaceus exconjugants could be transformed into the mcr- mrr- E.coli strain DH10B but not the mcr+ mrr+ strain DH5α, which suggests that a methylase system may be present in G.violaceus.

  12. Excitation energy transfer between Light-harvesting complex II and Photosystem I in reconstituted membranes.

    Science.gov (United States)

    Akhtar, Parveen; Lingvay, Mónika; Kiss, Teréz; Deák, Róbert; Bóta, Attila; Ughy, Bettina; Garab, Győző; Lambrev, Petar H

    2016-04-01

    Light-harvesting complex II (LHCII), the major peripheral antenna of Photosystem II in plants, participates in several concerted mechanisms for regulation of the excitation energy and electron fluxes in thylakoid membranes. In part, these include interaction of LHCII with Photosystem I (PSI) enhancing the latter's absorption cross-section - for example in the well-known state 1 - state 2 transitions or as a long-term acclimation to high light. In this work we examined the capability of LHCII to deliver excitations to PSI in reconstituted membranes in vitro. Proteoliposomes with native plant thylakoid membrane lipids and different stoichiometric ratios of LHCII:PSI were reconstituted and studied by steady-state and time-resolved fluorescence spectroscopy. Fluorescence emission from LHCII was strongly decreased in PSI-LHCII membranes due to trapping of excitations by PSI. Kinetic modelling of the time-resolved fluorescence data revealed the existence of separate pools of LHCII distinguished by the time scale of energy transfer. A strongly coupled pool, equivalent to one LHCII trimer per PSI, transferred excitations to PSI with near-unity efficiency on a time scale of less than 10ps but extra LHCIIs also contributed significantly to the effective antenna size of PSI, which could be increased by up to 47% in membranes containing 3 LHCII trimers per PSI. The results demonstrate a remarkable competence of LHCII to increase the absorption cross-section of PSI, given the opportunity that the two types of complexes interact in the membrane.

  13. Live imaging of chloroplast FtsZ1 filaments, rings, spirals, and motile dot structures in the AtMinE1 mutant and overexpressor of Arabidopsis thaliana.

    Science.gov (United States)

    Fujiwara, Makoto T; Sekine, Kohsuke; Yamamoto, Yoshiharu Y; Abe, Tomoko; Sato, Naoki; Itoh, Ryuuichi D

    2009-06-01

    Chloroplast division involves the tubulin-related GTPase FtsZ that assembles into a ring structure (Z-ring) at the mid-chloroplast division site, which is where invagination and constriction of the envelope membranes occur. Z-ring assembly is usually confined to the mid-chloroplast site by a well balanced counteraction of the stromal proteins MinD and MinE. The in vivo mechanisms by which FtsZ nucleates at specific sites, polymerises into a protofilament and organizes a closed ring of filament bundles remain largely unknown. To clarify the dynamic aspects of FtsZ, we developed a living cell system for simultaneous visualisation of various FtsZ configurations, utilising the Arabidopsis thaliana overexpressor and mutant of the MinE (AtMinE1) gene, which were modified to weakly express green fluorescent protein (GFP) fused to AtFtsZ1-1. Time-lapse observation in the chloroplasts of both plants revealed disorderly movement of the dots and short filaments of FtsZ. The short filaments often appeared to emanate from the dots and to converge with a long filament, producing a thick cable. In the AtMinE1 overexpressor, we also observed spirals along the longitudinal axis of the organelle that often rolled the closed rings together. In the atminE1 mutant, we visualised the 'isolated' rings with a maximum diameter of approximately 2 mum that did not encircle the organelle periphery, but appeared to be suspended in the stroma. Our observations further demonstrated heterogeneity in chloroplast shapes and concurrently altered configurations of FtsZ in the mutant.

  14. Mutants, Overexpressors, and Interactors of Arabidopsis Plastocyanin Isoforms: Revised Roles of Plastocyanin in Photosynthetic Electron Flow and Thylakoid Redox State

    Institute of Scientific and Technical Information of China (English)

    Paolo Pesaresi; Michael Scharfenberg; Martin Weigel; Irene Granlund; Wolfgang P. Schr(o)der; Giovanni Finazzi; Fabrice Rappaport; Simona Masiero; Antonella Furini; Peter Jahns; Dario Leister

    2009-01-01

    Two homologous plastocyanin isoforms are encoded by the genes PETE1 and PETE2 in the nuclear genome of Arabidopsis thaliana. The PETE2 transcript is expressed at considerably higher levels and the PETE2 protein is the more abundant isoform. Null mutations in the PETE genes resulted in plants, designated pete1 and pete2, with decreased plas-tocyanin contents. However, despite reducing plastocyanin levels by over~90%, a pete2 null mutation on its own affects rates of photosynthesis and growth only slightly, whereas pete1 knockout plants, with about 60-80% of the wild-type plastocyanin level, did not show any alteration. Hence, plastocyanin concentration is not limiting for photosynthetic elec-tron flow under optimal growth conditions, perhaps implying other possible physiological roles for the protein. Indeed, plastocyanin has been proposed previously to cooperate with cytochrome C6A (Cyt C6A) in thylakoid redox reactions, but we find no evidence for a physical interaction between the two proteins, using interaction assays in yeast. We observed homodimerization of Cyt C6A in yeast interaction assays, but also Cyt C6A homodimers failed to interact with plastocyanin. Moreover, phenotypic analysis of atc6-1 pete1 and atc6-1 pete2 double mutants, each lacking Cyt C6A and one of the two plastocyanin-encoding genes, failed to reveal any genetic interaction. Overexpression of either PETE1 or PETE2 in the pete1 pete2 double knockout mutant background results in essentially wild-type photosynthetic performance, excluding the possibility that the two plastocyanin isoforms could have distinct functions in thylakoid electron flow.

  15. Observation and Comparison of Chloroplast Structure in Hybrid and Different Cytoplasmic Male-sterile Wheat Lines%不同细胞质小麦雄性不育系及杂种F1叶绿体的观察与比较

    Institute of Scientific and Technical Information of China (English)

    袁凯; 高庆荣; 张保雷; 于松; 李楠楠; 张磊; 刘桓; 杨志远; 付修义

    2012-01-01

    chloroplast micro-structural characters in CMS male-sterile lines. [Method] The experiment was conducted with cultivars of the K, V, T-type CMS lines, the common maintainer Ji 5418, the F1 hybrids and the cytoplasmic donors under field conditions. The transmission electron microscope was used to investigate the features and expression differences of the chloroplast microstructure of the flag leaf. [Result] The chloroplast structure of K, V, T-type CMS lines were anomalous and showed that the boundaries of grana lamella of the choloraplast of were fuzzy, even disappeared, and also lag behind normal development. Meanwhile, the thylakoids among grana lamella were under developed and even ruptured, with the whole lamella disordered. But the maintainer line(B) Ji 5418's chloroplast was in good condition and close to the intima, the grana lamella clear and in order, and the thylakoid among grana was clearly visible. The number of chloroplasts per cell of the maintainer line was significantly different from K- and V-type (F=40.47, Pr<0.0001), with 19.8 in K-type and 18.4 in T-type, respectively. Moreover, the K-and T-type CMS lines had round chloroplast. The number of chloroplasts per cell in CMS V-type is 24.8, which was observably higher than that of CMS K and T-type (F=40.47, iV<0.0001), but did not significantly differ from the maintainer line (F=40.47, Pr<0.0001), with the number of chloroplasts of 24.1. Besides, the oblong chloroplast was found in CMS V-type CMS line and maintainer line. The donors of Aegilops kotschyi and Triticum timopheevii had round chloroplast, and oblong in Aegilops ventricos. Their boundaries of grana lamella of the choloraplast were fuzzy, the thylakoids among grana lamella were under developed and even ruptured. That is similar to K-, V-, T-type CMS lines, and they have the same source. The three kinds of chloroplasts of (AXR) F1 were oblong and close to the cell intimal. Their bi-layer epicyte were clear and cytoplasm were dense. Meanwhile, the

  16. Arabidopsis VARIEGATED 3 encodes a chloroplast-targeted, zinc-finger protein required for chloroplast and palisade cell development

    DEFF Research Database (Denmark)

    Næsted, Henrik; Holm, Agnethe; Jenkins, Tom

    2004-01-01

    protein containing novel repeats and zinc fingers described as protein interaction domains. VAR3 interacts specifically in yeast and in vitro with NCED4, a putative polyene chain or carotenoid dioxygenase, and both VAR3 and NCED4 accumulate in the chloroplast stroma. Metabolic profiling demonstrates...

  17. Differential positioning of C4 mesophyll and bundle sheath chloroplasts: aggregative movement of C4 mesophyll chloroplasts in response to environmental stresses.

    Science.gov (United States)

    Yamada, Masahiro; Kawasaki, Michio; Sugiyama, Tatsuo; Miyake, Hiroshi; Taniguchi, Mitsutaka

    2009-10-01

    In C(4) plants, mesophyll (M) chloroplasts are randomly distributed along the cell walls, while bundle sheath (BS) chloroplasts are typically located in either a centripetal or centrifugal position. We investigated whether these intracellular positions are affected by environmental stresses. When mature leaves of finger millet (Eleusine coracana) were exposed to extremely high intensity light, most M chloroplasts aggregatively re-distributed to the BS side, whereas the intracellular arrangement of BS chloroplasts was unaffected. Compared with the homologous light-avoidance movement of M chloroplasts in C(3) plants, it requires extremely high light (3,000-4,000 micromol m(-2) s(-1)) and responds more slowly (distinctive movement observed in 1 h). The high light-induced movement of M chloroplasts was also observed in maize (Zea mays), another C(4) species, but with a distinct pattern of redistribution along the sides of anticlinal walls, analogous to C(3) plants. The aggregative movement of M chloroplasts occurred at normal light intensities (250-500 micromol m(-2) s(-1)) in response to environmental stresses, such as drought, salinity and hyperosmosis. Moreover, the re-arrangement of M chloroplasts was observed in field-grown C(4) plants when exposed to mid-day sunlight, but also under midsummer drought conditions. The migration of M chloroplasts was controlled by actin filaments and also induced in a light-dependent fashion upon incubation with ABA, which may be the physiological signal transducer. Together these results suggest that M and BS cells of C(4) plants have different mechanisms controlling intracellular chloroplast positioning, and that the aggregative movement of C(4) M chloroplasts is thought to be a protective response under environmental stress conditions.

  18. The Tat system of Gram-positive bacteria

    NARCIS (Netherlands)

    Goosens, Vivianne J.; Monteferrante, Carmine G.; van Dijl, Jan Maarten

    2014-01-01

    The twin-arginine protein translocation (Tat) system has a unique ability to translocate folded and co-factor-containing proteins across lipid bilayers. The Tat pathway is present in bacteria, archaea and in the thylakoid membranes of chloroplasts and, depending on the organism and environmental con

  19. Fluorescence Image Analysis for Quantification of Active Oxygen Induced by Photochemical Reaction

    Science.gov (United States)

    2007-11-02

    state 1 Zn CP-III excited triplet state 3 Zn CP-III ground state ZnCP-III movement of energy absorption intersystem crossing...Takahashi and K. Asada, “Superoxide Anion Permeability of Phospholipid Membranes and Chloroplast Thylakoids,” Arc. Biochem. Biolhys., vol. 226, no23, pp.558–566, 1983.

  20. Effects of elevated CO[sub 2] on growth and chloroplast proteins in Prunus avium

    Energy Technology Data Exchange (ETDEWEB)

    Wilkins, D.; Oosten, J.-J. van; Besford, R.T. (Horticulture Research International, Littlehampton, Sussex (United Kingdom))

    1994-01-01

    A study was conducted of the growth response of Prunus avium L. Stella (wild cherry) to elevated CO[sub 2]. The associated changes in photosynthetic machinery of the leaf tissue were characterized. Self-pollinated seedlings and mature cuttings (clones) from the same parent plant of P. avium were grown for two consecutive growing seasons (about 60 days each) in ambient or elevated CO[sub 2] with high or low nutrient supply. The degree of acclimation of leaf biochemistry and growth response to elevated CO[sub 2] depended on the plant material (seedling or mature cutting) and nutrient supply. There was little or no growth response to elevated CO[sub 2] in seedlings or cuttings in the low nutrient supply treatments, whereas in both seasons, there was a strongly positive growth response to elevated CO[sub 2] in seedlings and cuttings in the high nutrient supply regimes, resulting in increases in the root/shoot ratio and in carbon allocation to the roots. In contrast, the protein content and activity of ribulose-1,5-biophosphate carboxylase-oxygenase (Rubisco, EC 4.1.1.39) were down regulated in elevated CO[sub 2]. The loss of Rubisco on an area basis in plants in the elevated CO[sub 2] treatments was compensated for at the canopy level by increased leaf area. The loss of Rubisco protein was accompanied by decreases in the contents of chlorophyll and the thylakoid membrane proteins D[sub 1], D[sub 2] and cytochrome f, which are involved in light harvesting and photo-electron transport. It is concluded that in the medium- to long-term, the initial stimulation of biomass production by elevated CO[sub 2] may be increasingly offset by a lower photosynthetic capacity per unit leaf area in perennial plants. 27 refs., 2 figs., 3 tabs.

  1. Impacts of high ATP supply from chloroplasts and mitochondria on the leaf metabolism of Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Chao eLiang

    2015-10-01

    Full Text Available Chloroplasts and mitochondria are the major ATP producing organelles in plant leaves. Arabidopsis thaliana purple acid phosphatase 2 (AtPAP2 is a phosphatase dually targeted to the outer membranes of both organelles and it plays a role in the import of selected nuclear-encoded proteins into these two organelles. Overexpression (OE of AtPAP2 in Arabidopsis thaliana accelerates plant growth and promotes flowering, seed yield and biomass at maturity. Measurement of ADP/ATP/NADP+/NADPH contents in the leaves of 20-day-old OE and wild-type lines at the end of night and at 1 and 8 h following illumination in a 16/8 h photoperiod revealed that the ATP levels and ATP/NADPH ratios were significantly increased in the OE line at all three time points. The AtPAP2 OE line is therefore a good model to investigate the impact of high energy on the global molecular status of Arabidopsis. In this study, transcriptome, proteome and metabolome profiles of the high ATP transgenic line were examined and compared with those of wild-type plants. A comparison of OE and WT at the end of the night provide valuable information on the impact of higher ATP output from mitochondria on plant physiology, as mitochondrial respiration is the major source of ATP in the dark in leaves. Similarly, comparison of OE and WT following illumination will provide information on the impact of higher energy output from chloroplasts on plant physiology. Overexpression of AtPAP2 was found to significantly affect the transcript and protein abundances of genes encoded by the two organellar genomes. For example, the protein abundances of many ribosomal proteins encoded by the chloroplast genome were higher in the AtPAP2 OE line under both light and dark conditions, while the protein abundances of multiple components of the photosynthetic complexes were lower. RNA-seq data also showed that the transcription of the mitochondrial genome is greatly affected by the availability of energy. These data

  2. Stochastic dynamics of actin filaments in guard cells regulating chloroplast localization during stomatal movement.

    Science.gov (United States)

    Wang, Xiu-Ling; Gao, Xin-Qi; Wang, Xue-Chen

    2011-08-01

    Actin filaments and chloroplasts in guard cells play roles in stomatal function. However, detailed actin dynamics vary, and the roles that they play in chloroplast localization during stomatal movement remain to be determined. We examined the dynamics of actin filaments and chloroplast localization in transgenic tobacco expressing green fluorescent protein (GFP)-mouse talin in guard cells by time-lapse imaging. Actin filaments showed sliding, bundling and branching dynamics in moving guard cells. During stomatal movement, long filaments can be severed into small fragments, which can form longer filaments by end-joining activities. With chloroplast movement, actin filaments near chloroplasts showed severing and elongation activity in guard cells during stomatal movement. Cytochalasin B treatment abolished elongation, bundling and branching activities of actin filaments in guard cells, and these changes of actin filaments, and as a result, more chloroplasts were localized at the centre of guard cells. However, chloroplast turning to avoid high light, and sliding of actin fragments near the chloroplast, was unaffected following cytochalasin B treatment in guard cells. We suggest that the sliding dynamics of actin may play roles in chloroplast turning in guard cells. Our results indicate that the stochastic dynamics of actin filaments in guard cells regulate chloroplast localization during stomatal movement.

  3. Effects and mechanism of acid rain on plant chloroplast ATP synthase.

    Science.gov (United States)

    Sun, Jingwen; Hu, Huiqing; Li, Yueli; Wang, Lihong; Zhou, Qing; Huang, Xiaohua

    2016-09-01

    Acid rain can directly or indirectly affect plant physiological functions, especially photosynthesis. The enzyme ATP synthase is the key in photosynthetic energy conversion, and thus, it affects plant photosynthesis. To clarify the mechanism by which acid rain affects photosynthesis, we studied the effects of acid rain on plant growth, photosynthesis, chloroplast ATP synthase activity and gene expression, chloroplast ultrastructure, intracellular H(+) level, and water content of rice seedlings. Acid rain at pH 4.5 remained the chloroplast structure unchanged but increased the expression of six chloroplast ATP synthase subunits, promoted chloroplast ATP synthase activity, and increased photosynthesis and plant growth. Acid rain at pH 4.0 or less decreased leaf water content, destroyed chloroplast structure, inhibited the expression of six chloroplast ATP synthase subunits, decreased chloroplast ATP synthase activity, and reduced photosynthesis and plant growth. In conclusion, acid rain affected the chloroplast ultrastructure, chloroplast ATPase transcription and activity, and P n by changing the acidity in the cells, and thus influencing the plant growth and development. Finally, the effects of simulated acid rain on the test indices were found to be dose-dependent.

  4. Analysis of chlorophyll fluorescence reveals stage specific patterns of chloroplast-containing cells during Arabidopsis embryogenesis.

    Science.gov (United States)

    Tejos, Ricardo I; Mercado, Ana V; Meisel, Lee A

    2010-01-01

    The basic body plan of a plant is established early in embryogenesis when cells differentiate, giving rise to the apical and basal regions of the embryo. Using chlorophyll fluorescence as a marker for chloroplasts, we have detected specific patterns of chloroplast-containing cells at specific stages of embryogenesis. Non-randomly distributed chloroplast-containing cells are seen as early as the globular stage of embryogenesis in Arabidopsis. In the heart stage of embryogenesis, chloroplast containing cells are detected in epidermal cells as well as a central region of the heart stage embryo, forming a triangular septum of chloroplast-containing cells that divides the embryo into three equal sectors. Torpedo stage embryos have chloroplast-containing epidermal cells and a central band of chloroplast-containing cells in the cortex layer, just below the shoot apical meristem. In the walking-stick stage of embryogenesis, chloroplasts are present in the epidermal, cortex and endodermal cells. The chloroplasts appear reduced or absent from the provascular and columella cells of walking-stick stage embryos. These results suggest that there is a tight regulation of plastid differentiation during embryogenesis that generates specific patterns of chloroplast-containing cells in specific cell layers at specific stages of embryogenesis.

  5. Pb-induced avoidance-like chloroplast movements in fronds of Lemna trisulca L.

    Directory of Open Access Journals (Sweden)

    Sławomir Samardakiewicz

    Full Text Available Lead ions are particularly dangerous to the photosynthetic apparatus, but little is known about the effects of trace metals, including Pb, on regulation of chloroplast redistribution. In this study a new effect of lead on chloroplast distribution patterns and movements was demonstrated in mesophyll cells of a small-sized aquatic angiosperm Lemna trisulca L. (star duckweed. An analysis of confocal microscopy images of L. trisulca fronds treated with lead (15 μM Pb2+, 24 h in darkness or in weak white light revealed an enhanced accumulation of chloroplasts in the profile position along the anticlinal cell walls, in comparison to untreated plants. The rearrangement of chloroplasts in their response to lead ions in darkness was similar to the avoidance response of chloroplasts in plants treated with strong white light. Transmission electron microscopy X-ray microanalysis showed that intracellular chloroplast arrangement was independent of the location of Pb deposits, suggesting that lead causes redistribution of chloroplasts, which looks like a light-induced avoidance response, but is not a real avoidance response to the metal. Furthermore, a similar redistribution of chloroplasts in L. trisulca cells in darkness was observed also under the influence of exogenously applied hydrogen peroxide (H2O2. In addition, we detected an enhanced accumulation of endogenous H2O2 after treatment of plants with lead. Interestingly, H2O2-specific scavenger catalase partly abolished the Pb-induced chloroplast response. These results suggest that H2O2 can be involved in the avoidance-like movement of chloroplasts induced by lead. Analysis of photometric measurements revealed also strong inhibition (but not complete of blue-light-induced chloroplast movements by lead. This inhibition may result from disturbances in the actin cytoskeleton, as we observed fragmentation and disappearance of actin filaments around chloroplasts. Results of this study show that the

  6. Pb-induced avoidance-like chloroplast movements in fronds of Lemna trisulca L.

    Science.gov (United States)

    Samardakiewicz, Sławomir; Krzeszowiec-Jeleń, Weronika; Bednarski, Waldemar; Jankowski, Artur; Suski, Szymon; Gabryś, Halina; Woźny, Adam

    2015-01-01

    Lead ions are particularly dangerous to the photosynthetic apparatus, but little is known about the effects of trace metals, including Pb, on regulation of chloroplast redistribution. In this study a new effect of lead on chloroplast distribution patterns and movements was demonstrated in mesophyll cells of a small-sized aquatic angiosperm Lemna trisulca L. (star duckweed). An analysis of confocal microscopy images of L. trisulca fronds treated with lead (15 μM Pb2+, 24 h) in darkness or in weak white light revealed an enhanced accumulation of chloroplasts in the profile position along the anticlinal cell walls, in comparison to untreated plants. The rearrangement of chloroplasts in their response to lead ions in darkness was similar to the avoidance response of chloroplasts in plants treated with strong white light. Transmission electron microscopy X-ray microanalysis showed that intracellular chloroplast arrangement was independent of the location of Pb deposits, suggesting that lead causes redistribution of chloroplasts, which looks like a light-induced avoidance response, but is not a real avoidance response to the metal. Furthermore, a similar redistribution of chloroplasts in L. trisulca cells in darkness was observed also under the influence of exogenously applied hydrogen peroxide (H2O2). In addition, we detected an enhanced accumulation of endogenous H2O2 after treatment of plants with lead. Interestingly, H2O2-specific scavenger catalase partly abolished the Pb-induced chloroplast response. These results suggest that H2O2 can be involved in the avoidance-like movement of chloroplasts induced by lead. Analysis of photometric measurements revealed also strong inhibition (but not complete) of blue-light-induced chloroplast movements by lead. This inhibition may result from disturbances in the actin cytoskeleton, as we observed fragmentation and disappearance of actin filaments around chloroplasts. Results of this study show that the mechanisms of the toxic

  7. Chloroplast genome sequence of the moss Tortula ruralis: gene content, polymorphism, and structural arrangement relative to other green plant chloroplast genomes

    OpenAIRE

    Wolf Paul G; Everett Karin DE; Mandoli Dina F; Boore Jeffrey L; Kuehl Jennifer V; Mishler Brent D; Murdock Andrew G; Oliver Melvin J; Duffy Aaron M; Karol Kenneth G

    2010-01-01

    Abstract Background Tortula ruralis, a widely distributed species in the moss family Pottiaceae, is increasingly used as a model organism for the study of desiccation tolerance and mechanisms of cellular repair. In this paper, we present the chloroplast genome sequence of T. ruralis, only the second published chloroplast genome for a moss, and the first for a vegetatively desiccation-t...

  8. Membrane dynamics

    DEFF Research Database (Denmark)

    Bendix, Pól Martin

    2015-01-01

    Current topics include membrane-protein interactions with regard to membrane deformation or curvature sensing by BAR domains. Also, we study the dynamics of membrane tubes of both cells and simple model membrane tubes. Finally, we study membrane phase behavior which has important implications...... for the lateral organization of membranes as wells as for physical properties like bending, permeability and elasticity...

  9. Actin-based mechanisms for light-dependent intracellular positioning of nuclei and chloroplasts in Arabidopsis.

    Science.gov (United States)

    Iwabuchi, Kosei; Takagi, Shingo

    2010-08-01

    The plant organelles, chloroplast and nucleus, change their position in response to light. In Arabidopsis thaliana leaf cells, chloroplasts and nuclei are distributed along the inner periclinal wall in darkness. In strong blue light, they become positioned along the anticlinal wall, while in weak blue light, only chloroplasts are accumulated along the inner and outer periclinal walls. Blue-light dependent positioning of both organelles is mediated by the blue-light receptor phototropin and controlled by the actin cytoskeleton. Interestingly, however, it seems that chloroplast movement requires short, fine actin filaments organized at the chloroplast edge, whereas nuclear movement does cytoplasmic, thick actin bundles intimately associated with the nucleus. Although there are many similarities between photo-relocation movements of chloroplasts and nuclei, plant cells appear to have evolved distinct mechanisms to regulate actin organization required for driving the movements of these organelles.

  10. Photoprotective function of chloroplast avoidance movement: in vivo chlorophyll fluorescence study.

    Science.gov (United States)

    Sztatelman, Olga; Waloszek, Andrzej; Banaś, Agnieszka Katarzyna; Gabryś, Halina

    2010-06-15

    Light-induced chloroplast avoidance movement has long been considered to be a photoprotective mechanism. Here, we present an experimental model in which this function can be shown for wild type Arabidopsis thaliana. We used blue light of different fluence rates for chloroplast positioning, and strong red light inactive in chloroplast positioning as a stressing light. The performance of photosystem II was measured by means of chlorophyll fluorescence. After stressing light treatment, a smaller decrease in photosystem II quantum yield was observed for leaves with chloroplasts in profile position as compared with leaves with chloroplasts in face position. Three Arabidopsis mutants, phot2 (no avoidance response), npq1 (impaired zeaxanhtin accumulation) and stn7 (no state transition), were examined for their chloroplast positioning and chlorophyll fluorescence parameters under identical experimental conditions. The results obtained for these mutants revealed additional stressing effects of blue light as compared with red light.

  11. Study on Chloroplast Ultrastructure in Different Color Period of Euphorbia pulcherrima

    Institute of Scientific and Technical Information of China (English)

    FU Jia; NIU De; WANG Lijuan

    2008-01-01

    By the observation of chloroplast ultrastructure in different period of bract colors of Euphorbia pulcherrima,the paper studied the change of chloroplast ultrastructrural in the transition process of bract colors, identified the rehtionship between E.pulcherrima color change and the chloroplast ultrastructure to provide theorical bases for the cultivation management and further study of E.pulcherrima.Ultrastructural study showed that in the process of change from green to red,the chloroplast of bracts disintegrated gradually,lamellar structure was destroyed gradually,and the content of chloroplasts in mesophyll cells was also reduced gradually. When bracts color resumed to turn green gradually,the content of chloroplasts in mesophyll cells was also increased gradually.

  12. Glycolate oxidation in A. thaliana chloroplasts improves biomass production

    Directory of Open Access Journals (Sweden)

    Alexandra eMaier

    2012-02-01

    Full Text Available A complete glycolate catabolic cycle was established in chloroplasts of the C3-model plant Arabidopsis thaliana by which one molecule of glycolate is completely oxidized within the chloroplast to two molecules of CO2. Genes coding for glycolate oxidase, malate synthase, and catalase were introduced into the nuclear genome of A. thaliana by step-wise transformation. Other genes required for a fully operational pathway are the endogenous NADP-malic enzyme and pyruvate dehydrogenase. Transgenic lines expressing the complete novel pathway produced rossettes with more leaves and higher fresh and dry weight but individual leaves were flatter and thinner than the wild type. The photosynthetic rates of the transgenic plants were higher on a dry weight and chlorophyll basis, but there were no differences in the compensation point. In addition, transgenic plants showed a lower glycine/serine ratio than the wild type indicating a reduction of the flux through the photorespiratory pathway. In this way, due to the increased oxidation of glycolate inside the chloroplasts, a photorespiratory bypass was created, which resulted in higher CO2 assimilation and enhanced biomass production.

  13. Photoinduction of cyclosis-mediated interactions between distant chloroplasts.

    Science.gov (United States)

    Bulychev, Alexander A; Komarova, Anna V

    2015-01-01

    Communications between chloroplasts and other organelles based on the exchange of metabolites, including redox active substances, are recognized as a part of intracellular regulation, chlororespiration, and defense against oxidative stress. Similar communications may operate between spatially distant chloroplasts in large cells where photosynthetic and respiratory activities are distributed unevenly under fluctuating patterned illumination. Microfluorometry of chlorophyll fluorescence in vivo in internodal cells of the alga Chara corallina revealed that a 30-s pulse of localized light induces a transient increase (~25%) in F' fluorescence of remote cell parts exposed to dim background light at a 1.5-mm distance on the downstream side from the illuminated spot in the plane of unilateral cytoplasmic streaming but has no effect on F' at equal distance on the upstream side. An abrupt arrest of cytoplasmic streaming for about 30s by triggering the action potential extended either the ascending or descending fronts of the F' fluorescence response, depending on the exact moment of streaming cessation. The response of F' fluorescence to localized illumination of a distant cell region was absent in dark-adapted internodes, when the localized light was applied within the first minute after switching on continuous background illumination of the whole cell, but it appeared in full after longer exposures to continuous background light. These results and the elimination of the F' response by methyl viologen known to redirect electron transport pathways beyond photosystem I indicate the importance of photosynthetic induction and the stromal redox state for long-distance communications of chloroplasts in vivo.

  14. Pea amyloplast DNA is qualitatively similar to pea chloroplast DNA

    Science.gov (United States)

    Gaynor, J. J.

    1984-01-01

    Amyloplast DNA (apDNA), when subjected to digestion with restriction endonucleases, yields patterns nearly identical to that of DNA from mature pea chloroplasts (ctDNA). Southern transfers of apDNA and ctDNA, probed with the large subunit (LS) gene of ribulose-1,5-bisphosphate carboxylase (Rubisco), shows hybridization to the expected restriction fragments for both apDNA and ctDNA. However, Northern transfers of total RNA from chloroplasts and amyloplasts, probed again with the LS gene of Rubisco, shows that no detectable LS meggage is found in amyloplasts although LS expression in mature chloroplasts is high. Likewise, two dimensional polyacrylamide gel electrophoresis of etiolated gravisensitive pea tissue shows that both large and small subunits of Rubisco are conspicuously absent; however, in greening tissue these two constitute the major soluble proteins. These findings suggest that although the informational content of these two organelle types is equivalent, gene expression is quite different and is presumably under nuclear control.

  15. Functional analysis and expression characteristics of chloroplastic Prx IIE.

    Science.gov (United States)

    Gama, Filipe; Bréhélin, Claire; Gelhaye, Eric; Meyer, Yves; Jacquot, Jean-Pierre; Rey, Pascal; Rouhier, Nicolas

    2008-07-01

    Peroxiredoxins (Prxs) are ubiquitous thiol-dependent peroxidases capable of eliminating a variety of peroxides through reactive catalytic cysteines, which are regenerated by reducing systems. Based on amino acid sequences and their mode of catalysis, five groups of thiol peroxidases have been distinguished in plants, and type II Prx is one of them with representatives in many sub-cellular compartments. The mature form of poplar chloroplastic Prx IIE was expressed as a recombinant protein in Escherichia coli. The protein is able to reduce H2O2 and tert-butyl hydroperoxide and is regenerated by both glutaredoxin (Grx) and thioredoxin (Trx) systems. Nevertheless, compared with Trxs, Grxs, and more especially chloroplastic Grx S12, are far more efficient reductants towards Prx IIE. The expression of Prx IIE at both the mRNA and protein levels as a function of organ type and abiotic stress conditions was investigated. Western blot analysis revealed that Prx IIE gene is constitutively expressed in Arabidopsis thaliana, mostly in young and mature leaves and in flowers. Under photo-oxidative treatment and water deficit, almost no change was observed in the abundance of Prx IIE in A. thaliana, while the level of Prx Q (one of the two other chloroplastic Prxs with 2-Cys Prx) increased in response to both stresses, indicating that plastidic members of the Prx family exhibit specific patterns of expression under stress.

  16. Longevity of guard cell chloroplasts in falling leaves: implication for stomatal function and cellular aging

    Energy Technology Data Exchange (ETDEWEB)

    Zeiger, E.; Schwartz, A.

    1982-11-12

    Guard cell chloroplasts in senescing leaves from 12 species of perennial trees and three species of annual plants survived considerably longer than their mesophyll counterparts. In Ginkgo biloba, stomata from yellow leaves opened during the day and closed at night; guard cell chloroplasts from these leaves showed fluorescence transients associated with electron transport and photophosphorylation. These findings indicate that guard cell chloroplasts are highly conserved throughout the life-span of the leaf and that leaves retain stomatal control during senescence.

  17. Exploring photosynthesis evolution by comparative analysis of metabolic networks between chloroplasts and photosynthetic bacteria

    Directory of Open Access Journals (Sweden)

    Hou Jing

    2006-04-01

    Full Text Available Abstract Background Chloroplasts descended from cyanobacteria and have a drastically reduced genome following an endosymbiotic event. Many genes of the ancestral cyanobacterial genome have been transferred to the plant nuclear genome by horizontal gene transfer. However, a selective set of metabolism pathways is maintained in chloroplasts using both chloroplast genome encoded and nuclear genome encoded enzymes. As an organelle specialized for carrying out photosynthesis, does the chloroplast metabolic network have properties adapted for higher efficiency of photosynthesis? We compared metabolic network properties of chloroplasts and prokaryotic photosynthetic organisms, mostly cyanobacteria, based on metabolic maps derived from genome data to identify features of chloroplast network properties that are different from cyanobacteria and to analyze possible functional significance of those features. Results The properties of the entire metabolic network and the sub-network that consists of reactions directly connected to the Calvin Cycle have been analyzed using hypergraph representation. Results showed that the whole metabolic networks in chloroplast and cyanobacteria both possess small-world network properties. Although the number of compounds and reactions in chloroplasts is less than that in cyanobacteria, the chloroplast's metabolic network has longer average path length, a larger diameter, and is Calvin Cycle -centered, indicating an overall less-dense network structure with specific and local high density areas in chloroplasts. Moreover, chloroplast metabolic network exhibits a better modular organization than cyanobacterial ones. Enzymes involved in the same metabolic processes tend to cluster into the same module in chloroplasts. Conclusion In summary, the differences in metabolic network properties may reflect the evolutionary changes during endosymbiosis that led to the improvement of the photosynthesis efficiency in higher plants. Our

  18. CDP1, a novel component of chloroplast division site positioning system in Arabidopsis

    Institute of Scientific and Technical Information of China (English)

    Min Zhang; Yong Hu; Jingjing Jia; Dapeng Li; Runjie Zhang; Hongbo Gao; Yikun He

    2009-01-01

    Chloroplasts are plant-specific organelles that evolved from endosymbiotic cyanobacteria. They divide through binary fission. Selection of the chloroplast division site is pivotal for the symmetric chloroplast division. In E. coli, positioning of the division site at the midpoint of the cell is regulated by dynamic oscillation of the Min system, which includes MinC, MinD and MinE. Homologs of Mind and MinE in plants are involved in chloroplast division. The homolog of MinC still has not been identified in higher plants. However, an FtsZ-like protein, ARC3, was found to be involved in chloroplast division site positioning. Here, we report that chloroplast division site positioning 1 (AtCDP1) is a novel chloroplast division protein involved in chloroplast division site placement in Arabidopsis. AtCDP1 was dis-covered by screening an Arabidopsis cDNA expression library in bacteria for colonies with a cell division phenotype. AtCDP1 is exclusively expressed in young green tissues in Arabidopsis. Elongated chloroplasts with multiple division sites were observed in the loss-of-function cdpl mutant. Overexpression of AtCDPI caused a chloroplast division phe-notype too. Protein interaction assays suggested that AtCDP1 may mediate the chloroplast division site positioning through the interaction with ARC3. Overall, our results indicate that AtCDP1 is a novel component of the chloroplast division site positioning system, and the working mechanism of this system is different from that of the traditional MinCDE system in prokaryotic cells.

  19. OEP80, an essential protein paralogous to the chloroplast protein translocation channel Toc75, exists as a 70-kD protein in the Arabidopsis thaliana chloroplast outer envelope.

    Science.gov (United States)

    Hsu, Shih-Chi; Nafati, Mehdi; Inoue, Kentaro

    2012-01-01

    Toc75 and OEP80 are paralogous proteins found in the Viridiplantae lineages, and appear to have evolved from a protein in the outer membrane of an ancient cyanobacterium. Toc75 is known to act as a protein translocation channel at the outer membrane of the chloroplast envelope, whereas the exact function of OEP80 is not understood. In Arabidopsis thaliana, each protein is encoded by a single gene, and both are essential for plant viability from embryonic stages onward. Sequence annotation and immunoblotting data with an antibody against its internal sequence (αOEP80(325-337)) indicated that the molecular weight of OEP80 is ca. 80 kD. Here we present multiple data to show that the size of A. thaliana OEP80 is smaller than previously estimated. First, we prepared the antibody against a recombinant protein consisting of annotated full-length A. thaliana OEP80 with an N-terminal hexahistidine tag (αOEP80(1-732)). This antibody recognized a 70-kD protein in the A. thaliana chloroplast membrane fraction which migrated faster than the His-tagged antigen and the protein recognized by the αOEP80(325-337) antibody on SDS-PAGE. Immunoprecipitation followed by LC-MS/MS analysis confirmed that the 70-kD protein was encoded by the OEP80 cDNA. Next, we performed a genetic complementation assay using embryo-lethal oep80-null plants and constructs encoding OEP80 and its variants. The results revealed that the nucleotide sequence encoding the 52 N-terminal amino acids was not required for functional expression of OEP80 and accumulation of the 70-kD protein. The data also indicated that an additional C-terminal T7 tag remained intact without disrupting the functionality of OEP80, and was not exposed to the cytoplasmic surface of the chloroplast envelope. Finally, OEP80-T7 and Toc75 showed distinct migration patterns on blue native-PAGE. This study provides molecular tools to investigate the function of OEP80, and also calls for caution in using an anti-peptide antibody.

  20. Sonication-based isolation and enrichment of Chlorella protothecoides chloroplasts for illumina genome sequencing

    Energy Technology Data Exchange (ETDEWEB)

    Angelova, Angelina [University of Arizona; Park, Sang-Hycuk [University of Arizona; Kyndt, John [Bellevue University; Fitzsimmons, Kevin [University of Arizona; Brown, Judith K [University of Arizona

    2013-09-01

    With the increasing world demand for biofuel, a number of oleaginous algal species are being considered as renewable sources of oil. Chlorella protothecoides Krüger synthesizes triacylglycerols (TAGs) as storage compounds that can be converted into renewable fuel utilizing an anabolic pathway that is poorly understood. The paucity of algal chloroplast genome sequences has been an important constraint to chloroplast transformation and for studying gene expression in TAGs pathways. In this study, the intact chloroplasts were released from algal cells using sonication followed by sucrose gradient centrifugation, resulting in a 2.36-fold enrichment of chloroplasts from C. protothecoides, based on qPCR analysis. The C. protothecoides chloroplast genome (cpDNA) was determined using the Illumina HiSeq 2000 sequencing platform and found to be 84,576 Kb in size (8.57 Kb) in size, with a GC content of 30.8 %. This is the first report of an optimized protocol that uses a sonication step, followed by sucrose gradient centrifugation, to release and enrich intact chloroplasts from a microalga (C. prototheocoides) of sufficient quality to permit chloroplast genome sequencing with high coverage, while minimizing nuclear genome contamination. The approach is expected to guide chloroplast isolation from other oleaginous algal species for a variety of uses that benefit from enrichment of chloroplasts, ranging from biochemical analysis to genomics studies.

  1. A plant-specific protein essential for blue-light-induced chloroplast movements.

    Science.gov (United States)

    DeBlasio, Stacy L; Luesse, Darron L; Hangarter, Roger P

    2005-09-01

    In Arabidopsis (Arabidopsis thaliana), light-dependent chloroplast movements are induced by blue light. When exposed to low fluence rates of light, chloroplasts accumulate in periclinal layers perpendicular to the direction of light, presumably to optimize light absorption by exposing more chloroplast area to the light. Under high light conditions, chloroplasts become positioned parallel to the incoming light in a response that can reduce exposure to light intensities that may damage the photosynthetic machinery. To identify components of the pathway downstream of the photoreceptors that mediate chloroplast movements (i.e. phototropins), we conducted a mutant screen that has led to the isolation of several Arabidopsis mutants displaying altered chloroplast movements. The plastid movement impaired1 (pmi1) mutant exhibits severely attenuated chloroplast movements under all tested fluence rates of light, suggesting that it is a necessary component for both the low- and high-light-dependant chloroplast movement responses. Analysis of pmi1 leaf cross sections revealed that regardless of the light condition, chloroplasts are more evenly distributed in leaf mesophyll cells than in the wild type. The pmi1-1 mutant was found to contain a single nonsense mutation within the open reading frame of At1g42550. This gene encodes a plant-specific protein of unknown function that appears to be conserved among angiosperms. Sequence analysis of the protein suggests that it may be involved in calcium-mediated signal transduction, possibly through protein-protein interactions.

  2. ppGpp inhibits peptide elongation cycle of chloroplast translation system in vitro.

    Science.gov (United States)

    Nomura, Yuhta; Takabayashi, Taito; Kuroda, Hiroshi; Yukawa, Yasushi; Sattasuk, Kwanchanok; Akita, Mitsuru; Nozawa, Akira; Tozawa, Yuzuru

    2012-01-01

    Chloroplasts possess common biosynthetic pathways for generating guanosine 3',5'-(bis)pyrophosphate (ppGpp) from GDP and ATP by RelA-SpoT homolog enzymes. To date, several hypothetical targets of ppGpp in chloroplasts have been suggested, but they remain largely unverified. In this study, we have investigated effects of ppGpp on translation apparatus in chloroplasts by developing in vitro protein synthesis system based on an extract of chloroplasts isolated from pea (Pisum sativum). The chloroplast extracts showed stable protein synthesis activity in vitro, and the activity was sensitive to various types of antibiotics. We have demonstrated that ppGpp inhibits the activity of chloroplast translation in dose-effective manner, as does the toxic nonhydrolyzable GTP analog guanosine 5'-(β,γ-imido)triphosphate (GDPNP). We further examined polyuridylic acid-directed polyphenylalanine synthesis as a measure of peptide elongation activity in the pea chloroplast extract. Both ppGpp and GDPNP as well as antibiotics, fusidic acid and thiostrepton, inhibited the peptide elongation cycle of the translation system, but GDP in the similar range of the tested ppGpp concentration did not affect the activity. Our results thus show that ppGpp directly affect the translation system of chloroplasts, as they do that of bacteria. We suggest that the role of the ppGpp signaling system in translation in bacteria is conserved in the translation system of chloroplasts.

  3. Heterologous expression of a chloroplast outer envelope protein from Suaeda salsa confers oxidative stress tolerance and induces chloroplast aggregation in transgenic Arabidopsis plants.

    Science.gov (United States)

    Wang, Fang; Yang, Chun-Lin; Wang, Li-Li; Zhong, Nai-Qin; Wu, Xiao-Min; Han, Li-Bo; Xia, Gui-Xian

    2012-03-01

    Suaeda salsa is a euhalophytic plant that is tolerant to coastal seawater salinity. In this study, we cloned a cDNA encoding an 8.4 kDa chloroplast outer envelope protein (designated as SsOEP8) from S. salsa and characterized its cellular function. Steady-state transcript levels of SsOEP8 in S. salsa were up-regulated in response to oxidative stress. Consistently, ectopic expression of SsOEP8 conferred enhanced oxidative stress tolerance in transgenic Bright Yellow 2 (BY-2) cells and Arabidopsis, in which H(2) O(2) content was reduced significantly in leaf cells. Further studies revealed that chloroplasts aggregated to the sides of mesophyll cells in transgenic Arabidopsis leaves, and this event was accompanied by inhibited expression of genes encoding proteins for chloroplast movements such as AtCHUP1, a protein involved in actin-based chloroplast positioning and movement. Moreover, organization of actin cytoskeleton was found to be altered in transgenic BY-2 cells. Together, these results suggest that SsOEP8 may play a critical role in oxidative stress tolerance by changing actin cytoskeleton-dependent chloroplast distribution, which may consequently lead to the suppressed production of reactive oxygen species (ROS) in chloroplasts. One significantly novel aspect of this study is the finding that the small chloroplast envelope protein is involved in oxidative stress tolerance.

  4. A factor related to pseudouridine synthases is required for chloroplast group II intron trans-splicing in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Perron, K; Goldschmidt-Clermont, M; Rochaix, J D

    1999-11-15

    In Chlamydomonas reinhardtii, the psaA mRNA is assembled by a process involving two steps of trans-splicing that remove two group II introns and give rise to the mature mRNA. The products of at least 14 nuclear genes and one chloroplast gene (tscA) are necessary for this process. We have cloned Maa2, one of the nuclear genes involved in trans-splicing of the second intron. Maa2 encodes a protein with similarity to conserved domains of pseudouridine synthases, but mutagenesis of putative catalytic residues showed that this activity may not be required for trans-splicing of psaA RNA. Although it is not clear whether the pseudouridine synthase activity has been maintained in Maa2, it is possible that this enzyme was recruited during evolution as an RNA chaperone for folding or stabilizing the psaA intron. The Maa2 protein appears to be associated through ionic interactions with a low density membrane system in the chloroplast that also contains RNA-binding proteins involved in translation.

  5. Spontaneous capture of oilseed rape (Brassica napus) chloroplasts by wild B. rapa: implications for the use of chloroplast transformation for biocontainment.

    Science.gov (United States)

    Haider, Nadia; Allainguillaume, Joel; Wilkinson, Mike J

    2009-04-01

    Environmental concerns over the cultivation of Genetically Modified (GM) crops largely centre on the ecological consequences following gene flow to wild relatives. One attractive solution is to deploy biocontainment measures that prevent hybridization. Chloroplast transformation is the most advanced biocontainment method but is compromised by chloroplast capture (hybridization through the maternal lineage). To date, however, there is a paucity of information on the frequency of chloroplast capture in the wild. Oilseed rape (Brassica napus, AACC) frequently hybridises with wild Brassica rapa (AA, as paternal parent) and yields B. rapa-like introgressed individuals after only two generations. In this study we used chloroplast CAPS markers that differentiate between the two species to survey wild and weedy populations of B. rapa for the capture of B. napus chloroplasts. A total of 464 B. rapa plants belonging to 14 populations growing either in close proximity to B. napus (i.e. sympatric 1 km) were assessed for chloroplast capture using PCR (trnL-F) and CAPS (trnT-L-Xba I) markers. The screen revealed that two sympatric B. rapa populations included 53 plants that possessed the chloroplast of B. napus. In order to discount these B. rapa plants as F(1) crop-wild hybrids, we used a C-genome-specific marker and found that 45 out of 53 plants lacked the C-genome and so were at least second generation introgressants. The most plausible explanation is that these individuals represent multiple cases of chloroplast capture following introgressive hybridisation through the female germ line from the crop. The abundance of such plants in sympatric sites thereby questions whether the use of chloroplast transformation would provide a sufficient biocontainment for GM oilseed rape in the United Kingdom.

  6. Antisense-mediated suppression of tomato thylakoidal ascorbate peroxidase influences anti-oxidant network during chilling stress.

    Science.gov (United States)

    Duan, Ming; Ma, Na-Na; Li, Dong; Deng, Yong-Sheng; Kong, Fan-Ying; Lv, Wei; Meng, Qing-Wei

    2012-09-01

    Photosynthesis is a well-established source of reactive oxygen species (ROS) in plants particularly under chilling stress. Ascorbate peroxidase (APXs) plays an important role in the anti-oxidant system by utilizing AsA as specific electron donor to reduce H(2)O(2) to water. In order to investigate the possible mechanisms of ascorbate peroxidsae (APX) in photoprotection under chilling stress, a tomato (Lycopersicon esculentum Mill.) thylakoidal ascorbate peroxidase gene (LetAPX) was isolated and antisense transgenic tomato plants were produced. Under chilling stress, transgenic plants accumulated more H(2)O(2), and showed higher levels of ion leakage and malondialdehyde (MDA), lower net photosynthetic rate (Pn), lower maximal photochemical efficiency of PSII (Fv/Fm) and less content of D1 protein compared with wild type (WT) plants. On the other hand, after chilling stress, transgenic plants showed higher reduced ascorbate (AsA) and activities of catalase (CAT) and superoxide dismutase (SOD) than those in WT plants, and the expression of several known stress-responsive and antioxidative genes was also higher at the end of chilling treatment. These results suggested that the suppression of LetAPX gene induced compensatory anti-oxidant mechanisms in tomato, and inactivation of tAPX may have a regulatory role in facilitating redox signaling pathways under chilling stress. Furthermore, transient increases in ROS levels also have a vital role in stress signaling and thereby in the survival of plants under chilling conditions.

  7. Requirement of the C3HC4 zinc RING finger of the Arabidopsis PEX10 for photorespiration and leaf peroxisome contact with chloroplasts.

    Science.gov (United States)

    Schumann, Uwe; Prestele, Jakob; O'Geen, Henriette; Brueggeman, Robert; Wanner, Gerhard; Gietl, Christine

    2007-01-16

    Plant peroxisomes perform multiple vital metabolic processes including lipid mobilization in oil-storing seeds, photorespiration, and hormone biosynthesis. Peroxisome biogenesis requires the function of peroxin (PEX) proteins, including PEX10, a C(3)HC(4) Zn RING finger peroxisomal membrane protein. Loss of function of PEX10 causes embryo lethality at the heart stage. We investigated the function of PEX10 with conditional sublethal mutants. Four T-DNA insertion lines expressing pex10 with a dysfunctional RING finger were created in an Arabidopsis WT background (DeltaZn plants). They could be normalized by growth in an atmosphere of high CO(2) partial pressure, indicating a defect in photorespiration. beta-Oxidation in mutant glyoxysomes was not affected. However, an abnormal accumulation of the photorespiratory metabolite glyoxylate, a lowered content of carotenoids and chlorophyll a and b, and a decreased quantum yield of photosystem II were detected under normal atmosphere, suggesting impaired leaf peroxisomes. Light and transmission electron microscopy demonstrated leaf peroxisomes of the DeltaZn plants to be more numerous, multilobed, clustered, and not appressed to the chloroplast envelope as in WT. We suggest that inactivation of the RING finger domain in PEX10 has eliminated protein interaction required for attachment of peroxisomes to chloroplasts and movement of metabolites between peroxisomes and chloroplasts.

  8. Molecular Mechanism Involved in Chloroplast Division in Plants%植物叶绿体分裂的分子机制

    Institute of Scientific and Technical Information of China (English)

    谌志伟; 胡勇

    2013-01-01

    叶绿体是植物细胞内一种重要的细胞器.它不仅是光合作用的场所,还是其它多种中间代谢的场所.叶绿体起源于蓝细菌,与其原核祖先类似,通过二分裂方式进行增殖.最近的研究表明,叶绿体的分裂装置包含原核起源和真核起源的蛋白质,它们在叶绿体的内膜内侧和外膜外侧协同作用以完成叶绿体的分裂.在过去十几年里,包括丝状温度敏感蛋白Z(FtsZ)、Min系统蛋白、质体分裂蛋白(PDV)和ARC蛋白等在内的多个叶绿体分裂相关组分被分离鉴定.本文简要介绍了叶绿体分裂装置各成员的发现、叶绿体被膜的收缩和叶绿体分裂位点的选择机制.另外,植物发育过程中叶绿体分裂可能受到细胞的控制,但目前对细胞如何调控叶绿体分裂知之甚少.本文对该领域的最新研究进展也进行了综述.%The chloroplast is a specific organelle in photosynthetic eukaryotes that houses many essential metabolic pathways. It arose from an endosymbiotic event in which a cyanobacterium was engulfed by a heterotrophic eukaryotic host cell. Similar to its prokaryotic ancestor, each chloroplast arises from a preexisting chloroplast by binary division. Recent studies have revealed that chloroplast division is executed by the coordinated action of prokaryote-derived and host-derived proteins at the division site, encompassing both the inside and the outside of the two envelope membranes. Several chloroplast division components such as filamentous temperature-sensitive protein Z ( FtsZ) , Min, plastid division protein (PDV) and ARC have been identified in the past several years. Here we reviewed the progress in identifying the components of the chloroplast division complex to understand the mechanisms of envelope constriction and division-site placement in plants. The chloroplast division process may be controlled and coordinated by the host cell during proliferation and differentiation, but little is known

  9. Membrane associated qualitative differences in cell ultrastructure of chemically and high pressure cryofixed plant cells.

    Science.gov (United States)

    Zechmann, Bernd; Müller, Maria; Zellnig, Günther

    2007-06-01

    Membrane contrast can sometimes be poor in biological samples after high pressure freezing (HPF) and freeze substitution (FS). The addition of water to the FS-medium has been shown to improve membrane contrast in animal tissue and yeast. In the present study we tested the effects of 1% and 5% water added to the FS-medium (2% osmium with 0.2% uranyl acetate in anhydrous acetone) on the quality and visibility of membranes in high pressure frozen leaf samples of Cucurbita pepo L. plants and compared them to chemically fixed cells (3% glutaraldehyde post-fixed with 1% osmium tetroxide). The addition of water to the FS-medium drastically decreased the amounts of well preserved cells and did not significantly improve the quality nor visibility of membranes. In samples that were freeze substituted in FS-media containing 1% and 5% water the width of thylakoid membranes was found to be significantly increased of about 20% and the perinuclear space was up to 76% wider in comparison to what was found in samples which were freeze substituted without water. No differences were found in the thickness of membranes between chemically and cryofixed cells that were freeze substituted in the FS-medium without water. Nevertheless, in chemically fixed cells the intrathylakoidal space was about 120% wider than in cryofixed cells that were freeze substituted with or without water. The present results demonstrate that the addition of water to the FS-medium does not improve membrane contrast but changes the width of thylakoid membranes and the perinuclear space in the present plant material. The addition of water to the FS-medium is therefore not as essential for improved membrane contrast in the investigated plant samples as it was observed in cells of animal tissues and yeast cells.

  10. A split-ubiquitin yeast two-hybrid screen to examine the substrate specificity of atToc159 and atToc132, two Arabidopsis chloroplast preprotein import receptors.

    Directory of Open Access Journals (Sweden)

    Siddhartha Dutta

    Full Text Available Post-translational import of nucleus-encoded chloroplast pre-proteins is critical for chloroplast biogenesis, and the Toc159 family of proteins serve as receptors for the process. Toc159 shares with other members of the family (e.g. Toc132, homologous GTPase (G- and Membrane (M- domains, but a highly dissimilar N-terminal acidic (A- domain. Although there is good evidence that atToc159 and atToc132 from Arabidopsis mediate the initial sorting step, preferentially recognizing photosynthetic and non-photosynthetic preproteins, respectively, relatively few chloroplast preproteins have been assigned as substrates for particular members of the Toc159 family, which has limited the proof for the hypothesis. The current study expands the number of known preprotein substrates for members of the Arabidopsis Toc159 receptor family using a split-ubiquitin membrane-based yeast two-hybrid system using the atToc159 G-domain (Toc159G, atToc132 G-domain (Toc132G and atToc132 A- plus G-domains (Toc132AG as baits. cDNA library screening with all three baits followed by pairwise interaction assays involving the 81 chloroplast preproteins identified show that although G-domains of the Toc159 family are sufficient for preprotein recognition, they alone do not confer specificity for preprotein subclasses. The presence of the A-domain fused to atToc132G (Toc132AG not only positively influences its specificity for non-photosynthetic preproteins, but also negatively regulates the ability of this receptor to interact with a subset of photosynthetic preproteins. Our study not only substantiates the fact that atToc132 can serve as a receptor by directly binding to chloroplast preproteins but also proposes the existence of subsets of preproteins with different but overlapping affinities for more than one member of the Toc159 receptor family.

  11. Precursor binding to an 880-kDa Toc complex as an early step during active import of protein into chloroplasts.

    Science.gov (United States)

    Chen, Kuan-Yu; Li, Hsou-min

    2007-01-01

    The import of protein into chloroplasts is mediated by translocon components located in the chloroplast outer (the Toc proteins) and inner (the Tic proteins) envelope membranes. To identify intermediate steps during active import, we used sucrose density gradient centrifugation and blue-native polyacrylamide gel electrophoresis (BN-PAGE) to identify complexes of translocon components associated with precursor proteins under active import conditions instead of arrested binding conditions. Importing precursor proteins in solubilized chloroplast membranes formed a two-peak distribution in the sucrose density gradient. The heavier peak was in a similar position as the previously reported Tic/Toc supercomplex and was too large to be analyzed by BN-PAGE. The BN-PAGE analyses of the lighter peak revealed that precursors accumulated in at least two complexes. The first complex migrated at a position close to the ferritin dimer (approximately 880 kDa) and contained only the Toc components. Kinetic analyses suggested that this Toc complex represented an earlier step in the import process than the Tic/Toc supercomplex. The second complex in the lighter peak migrated at the position of the ferritin trimer (approximately 1320 kDa). It contained, in addition to the Toc components, Tic110, Hsp93, and an hsp70 homolog, but not Tic40. Two different precursor proteins were shown to associate with the same complexes. Processed mature proteins first appeared in the membranes at the same fractions as the Tic/Toc supercomplex, suggesting that processing of transit peptides occurs while precursors are still associated with the supercomplex.

  12. Evolutionary conservation of dual Sec translocases in the cyanelles of Cyanophora paradoxa

    Directory of Open Access Journals (Sweden)

    Löffelhardt Wolfgang

    2008-11-01

    Full Text Available Abstract Background Cyanelles, the peptidoglycan-armored plastids of glaucocystophytes, occupy a unique bridge position in between free-living cyanobacteria and chloroplasts. In some respects they side with cyanobacteria whereas other features are clearly shared with chloroplasts. The Sec translocase, an example for "conservative sorting" in the course of evolution, is found in the plasma membrane of all prokaryotes, in the thylakoid membrane of chloroplasts and in both these membrane types of cyanobacteria. Results In this paper we present evidence for a dual location of the Sec translocon in the thylakoid as well as inner envelope membranes of the cyanelles from Cyanophora paradoxa, i. e. conservative sorting sensu stricto. The prerequisite was the generation of specific antisera directed against cyanelle SecY that allowed immunodetection of the protein on SDS gels from both membrane types separated by sucrose density gradient floatation centrifugation. Immunoblotting of blue-native gels yielded positive but differential results for both the thylakoid and envelope Sec complexes, respectively. In addition, heterologous antisera directed against components of the Toc/Tic translocons and binding of a labeled precursor protein were used to discriminate between inner and outer envelope membranes. Conclusion The envelope translocase can be envisaged as a prokaryotic feature missing in higher plant chloroplasts but retained in cyanelles, likely for protein transport to the periplasm. Candidate passengers are cytochrome c6 and enzymes of peptidoglycan metabolism. The minimal set of subunits of the Toc/Tic translocase of a primitive plastid is proposed.

  13. Analysis of Acorus calamus chloroplast genome and its phylogenetic implications.

    Science.gov (United States)

    Goremykin, Vadim V; Holland, Barbara; Hirsch-Ernst, Karen I; Hellwig, Frank H

    2005-09-01

    Determining the phylogenetic relationships among the major lines of angiosperms is a long-standing problem, yet the uncertainty as to the phylogenetic affinity of these lines persists. While a number of studies have suggested that the ANITA (Amborella-Nymphaeales-Illiciales-Trimeniales-Aristolochiales) grade is basal within angiosperms, studies of complete chloroplast genome sequences also suggested an alternative tree, wherein the line leading to the grasses branches first among the angiosperms. To improve taxon sampling in the existing chloroplast genome data, we sequenced the chloroplast genome of the monocot Acorus calamus. We generated a concatenated alignment (89,436 positions for 15 taxa), encompassing almost all sequences usable for phylogeny reconstruction within spermatophytes. The data still contain support for both the ANITA-basal and grasses-basal hypotheses. Using simulations we can show that were the ANITA-basal hypothesis true, parsimony (and distance-based methods with many models) would be expected to fail to recover it. The self-evident explanation for this failure appears to be a long-branch attraction (LBA) between the clade of grasses and the out-group. However, this LBA cannot explain the discrepancies observed between tree topology recovered using the maximum likelihood (ML) method and the topologies recovered using the parsimony and distance-based methods when grasses are deleted. Furthermore, the fact that neither maximum parsimony nor distance methods consistently recover the ML tree, when according to the simulations they would be expected to, when the out-group (Pinus) is deleted, suggests that either the generating tree is not correct or the best symmetric model is misspecified (or both). We demonstrate that the tree recovered under ML is extremely sensitive to model specification and that the best symmetric model is misspecified. Hence, we remain agnostic regarding phylogenetic relationships among basal angiosperm lineages.

  14. Characterization of mango (Mangifera indica L.) transcriptome and chloroplast genome.

    Science.gov (United States)

    Azim, M Kamran; Khan, Ishtaiq A; Zhang, Yong

    2014-05-01

    We characterized mango leaf transcriptome and chloroplast genome using next generation DNA sequencing. The RNA-seq output of mango transcriptome generated >12 million reads (total nucleotides sequenced >1 Gb). De novo transcriptome assembly generated 30,509 unigenes with lengths in the range of 300 to ≥3,000 nt and 67× depth of coverage. Blast searching against nonredundant nucleotide databases and several Viridiplantae genomic datasets annotated 24,593 mango unigenes (80% of total) and identified Citrus sinensis as closest neighbor of mango with 9,141 (37%) matched sequences. The annotation with gene ontology and Clusters of Orthologous Group terms categorized unigene sequences into 57 and 25 classes, respectively. More than 13,500 unigenes were assigned to 293 KEGG pathways. Besides major plant biology related pathways, KEGG based gene annotation pointed out active presence of an array of biochemical pathways involved in (a) biosynthesis of bioactive flavonoids, flavones and flavonols, (b) biosynthesis of terpenoids and lignins and (c) plant hormone signal transduction. The mango transcriptome sequences revealed 235 proteases belonging to five catalytic classes of proteolytic enzymes. The draft genome of mango chloroplast (cp) was obtained by a combination of Sanger and next generation sequencing. The draft mango cp genome size is 151,173 bp with a pair of inverted repeats of 27,093 bp separated by small and large single copy regions, respectively. Out of 139 genes in mango cp genome, 91 found to be protein coding. Sequence analysis revealed cp genome of C. sinensis as closest neighbor of mango. We found 51 short repeats in mango cp genome supposed to be associated with extensive rearrangements. This is the first report of transcriptome and chloroplast genome analysis of any Anacardiaceae family member.

  15. Evolution from the prokaryotic to the higher plant chloroplast signal recognition particle

    DEFF Research Database (Denmark)

    Träger, Chantal; Rosenblad, Magnus Alm; Ziehe, Dominik;

    2012-01-01

    The protein targeting signal recognition particle (SRP) pathway in chloroplasts of higher plants has undergone dramatic evolutionary changes. It disposed of its RNA, which is an essential SRP component in bacteria, and uses a unique chloroplast-specific protein cpSRP43. Nevertheless, homologs of ...

  16. The Unicellular Green Alga Chlamydomonas reinhardtii as an Experimental System to Study Chloroplast RNA Metabolism

    Science.gov (United States)

    Nickelsen, J.; Kück, U.

    Chloroplasts are typical organelles of photoautotrophic eukaryotic cells which drive a variety of functions, including photosynthesis. For many years the unicellular green alga Chlamydomonas reinhardtii has served as an experimental organism for studying photosynthetic processes. The recent development of molecular tools for this organism together with efficient methods of genetic analysis and the availability of many photosynthesis mutants has now made this alga a powerful model system for the analysis of chloroplast biogenesis. For example, techniques have been developed to transfer recombinant DNA into both the nuclear and the chloroplast genome. This allows both complementation tests and analyses of gene functions in vivo. Moreover, site-specific DNA recombinations in the chloroplast allow targeted gene disruption experiments which enable a "reverse genetics" to be performed. The potential of the algal system for the study of chloroplast biogenesis is illustrated in this review by the description of regulatory systems of gene expression involved in organelle biogenesis. One example concerns the regulation of trans-splicing of chloroplast mRNAs, a process which is controlled by both multiple nuclear- and chloroplast-encoded factors. The second example involves the stabilization of chloroplast mRNAs. The available data lead us predict distinct RNA elements, which interact with trans-acting factors to protect the RNA against nucleolytic attacks.

  17. Changes in leaf optical properties associated with light-dependent chloroplast movements.

    Science.gov (United States)

    Davis, Phillip A; Caylor, Steven; Whippo, Craig W; Hangarter, Roger P

    2011-12-01

    We surveyed 24 plant species to examine how leaf anatomy influenced chloroplast movement and how the optical properties of leaves change with chloroplast position. All species examined exhibited light-dependent chloroplast movements but the associated changes in leaf absorptance varied considerably in magnitude. Chloroplast movement-dependent changes in leaf absorptance were greatest in shade species, in which absorptance changes of >10% were observed between high- and low-light treatments. Using the Kubelka-Munk theory, we found that changes in the absorption (k) and chlorophyll a absorption efficiency (k*) associated with chloroplast movement correlated with cell diameter, such that the narrower, more columnar cells found in sun leaves restricted the ability of chloroplasts to move. The broader, more spherical cells of shade leaves allowed greater chloroplast rearrangements and in low-light conditions allowed efficient light capture. Across the species tested, light-dependent chloroplast movements modulated leaf optical properties and light absorption efficiency by manipulating the package (sieve or flattening) effect but not the detour (path lengthening) effect.

  18. Phototropins mediate blue and red light-induced chloroplast movements in Physcomitrella patens.

    Science.gov (United States)

    Kasahara, Masahiro; Kagawa, Takatoshi; Sato, Yoshikatsu; Kiyosue, Tomohiro; Wada, Masamitsu

    2004-07-01

    Phototropin is the blue-light receptor that mediates phototropism, chloroplast movement, and stomatal opening in Arabidopsis. Blue and red light induce chloroplast movement in the moss Physcomitrella patens. To study the photoreceptors for chloroplast movement in P. patens, four phototropin genes (PHOTA1, PHOTA2, PHOTB1, and PHOTB2) were isolated by screening cDNA libraries. These genes were classified into two groups (PHOTA and PHOTB) on the basis of their deduced amino acid sequences. Then phototropin disruptants were generated by homologous recombination and used for analysis of chloroplast movement. Data revealed that blue light-induced chloroplast movement was mediated by phototropins in P. patens. Both photA and photB groups were able to mediate chloroplast avoidance, as has been reported for Arabidopsis phot2, although the photA group contributed more to the response. Red light-induced chloroplast movement was also significantly reduced in photA2photB1photB2 triple disruptants. Because the primary photoreceptor for red light-induced chloroplast movement in P. patens is phytochrome, phototropins may be downstream components of phytochromes in the signaling pathway. To our knowledge, this work is the first to show a function for the phototropin blue-light receptor in a response to wavelengths that it does not absorb.

  19. Chloroplast photorelocation movement mediated by phototropin family proteins in green plants.

    Science.gov (United States)

    Suetsugu, Noriyuki; Wada, Masamitsu

    2007-09-01

    Chloroplasts gather in areas irradiated with weak light to maximize photosynthesis (the accumulation response). They move away from areas irradiated with strong light to minimize damage of the photosynthetic apparatus (the avoidance response). The processes underlying these chloroplast movements can be divided into three parts: photoperception, signal transduction, and chloroplast movement. Photoreceptors for chloroplast movement have been identified recently in various plant species. A blue light receptor phototropin (phot) mediates chloroplast photorelocation movement in the seed plant Arabidopsis thaliana, the fern Adiantum capillus-veneris, the moss Physcomitrella patens and possibly the green alga Mougeotia scalaris. A chimeric photoreceptor between phytochrome and phototropin, neochrome (neo), was found in some advanced ferns and in the green alga M. scalaris. While the mechanism of chloroplast movement is not well understood, it is known that actin filaments play an important role in this process. To understand the molecular mechanisms associated with chloroplast movement, several mutants were isolated in A. thaliana (jac1 and chup1) and the corresponding genes were cloned. In this review, recent progress in photoreceptor research into chloroplast movement in various plant species and the possible factors functioning in signal transduction or the regulation of actin filaments identified in A. thaliana is discussed.

  20. Functional proteomics of barley and barley chloroplasts – strategies, methods and perspectives

    DEFF Research Database (Denmark)

    Petersen, Jørgen; Rogowska-Wrzesinska, Adelina; Jensen, Ole Nørregaard

    2013-01-01

    tolerance, micronutrient utilization, and photosynthesis in barley. In the present review we present the current state of proteomics research for investigations of barley chloroplasts, i.e., the organelle that contain the photosynthetic apparatus in the plant. We describe several different proteomics...... strategies and discuss their applications in characterization of the barley chloroplast as well as future perspectives for functional proteomics in barley research....

  1. Hartmut Lichtenthaler: an authority on chloroplast structure and isoprenoid biochemistry.

    Science.gov (United States)

    Sharkey, Thomas D; Govindjee

    2016-05-01

    We pay tribute to Hartmut Lichtenthaler for making important contributions to the field of photosynthesis research. He was recently recognized for ground-breaking discoveries in chloroplast structure and isoprenoid biochemistry by the Rebeiz Foundation for Basic Research (RFBR; http://vlpbp.org/ ), receiving a 2014 Lifetime Achievement Award for Photosynthesis. The ceremony, held in Champaign, Illinois, was attended by many prominent researchers in the photosynthesis field. We provide below a brief note on his education, and then describe some of the areas in which Hartmut Lichtenthaler has been a pioneer.

  2. Roles of Lipids in Photosynthesis.

    Science.gov (United States)

    Kobayashi, Koichi; Endo, Kaichiro; Wada, Hajime

    2016-01-01

    Thylakoid membranes in cyanobacterial cells and chloroplasts of algae and higher plants are the sites of oxygenic photosynthesis. The lipid composition of the thylakoid membrane is unique and highly conserved among oxygenic photosynthetic organisms. Major lipids in thylakoid membranes are glycolipids, monogalactosyldiacylglycerol, digalactosyldiacylglycerol and sulfoquinovosyldiacylglycerol, and the phospholipid, phosphatidylglycerol. The identification of almost all genes involved in the biosynthesis of each lipid class over the past decade has allowed the generation and isolation of mutants of various photosynthetic organisms incapable of synthesizing specific lipids. Numerous studies using such mutants have revealed that these lipids play important roles not only in the formation of the lipid bilayers of thylakoid membranes but also in the folding and assembly of the protein subunits in photosynthetic complexes. In addition to the studies with the mutants, recent X-ray crystallography studies of photosynthetic complexes in thylakoid membranes have also provided critical information on the association of lipids with photosynthetic complexes and their activities. In this chapter, we summarize our current understanding about the structural and functional involvement of thylakoid lipids in oxygenic photosynthesis.

  3. The complete chloroplast genome sequence of Dendropanax morbifera (Léveillé).

    Science.gov (United States)

    Kim, Kyunghee; Lee, Sang-Choon; Yang, Tae-Jin

    2016-07-01

    The complete chloroplast genome sequence of Dendropanax morbifera, an economically and medicinally important endemic tree species in Korea, was obtained by de novo assembly with whole-genome sequence data and manual correction. A circular 156 366-bp chloroplast genome showed typical chloroplast genome structure comprising a large single copy region of 86 475 bp, a small single copy region of 18 125 bp, and a pair of inverted repeats of 25 883 bp. The chloroplast genome harbored 87 protein-coding genes. Phylogenetic analysis with the chloroplast genome revealed that D. morbifera is most closely related to Dendropanax dentiger, an evergreen tree species in China and Southeastern Asia.

  4. Update on Chloroplast Research: New Tools, New Topics, and New Trends

    Institute of Scientific and Technical Information of China (English)

    Ute Armbruster; Paolo Pesaresi; Mathias Pribil; Alexander Hertle; Dario Leister

    2011-01-01

    Chloroplasts, the green differentiation form of plastids, are the sites of photosynthesis and other important plant functions. Genetic and genomic technologies have greatly boosted the rate of discovery and functional characterization of chloroplast proteins during the past decade. Indeed, data obtained using high-throughput methodologies, in particular proteomics and transcriptomics, are now routinely used to assign functions to chloroplast proteins. Our knowledge of many chloroplast processes, notably photosynthesis and photorespiration, has reached such an advanced state that biotechnological approaches to crop improvement now seem feasible. Meanwhile, efforts to identify the entire complement of chloroplast proteins and their interactions are progressing rapidly, making the organelle a prime target for systems biology research in plants.

  5. Unexpected roles of plastoglobules (plastid lipid droplets) in vitamin K1 and E metabolism.

    Science.gov (United States)

    Spicher, Livia; Kessler, Felix

    2015-06-01

    Tocopherol (vitamin E) and phylloquinone (vitamin K1) are lipid-soluble antioxidants that can only be synthesized by photosynthetic organisms. These compounds function primarily at the thylakoid membrane but are also present in chloroplast lipid droplets, also known as plastoglobules (PG). Depending on environmental conditions and stage of plant development, changes in the content, number and size of PG occur. PG are directly connected to the thylakoid membrane via the outer lipid leaflet. Apart from storage, PG are active in metabolism and likely trafficking of diverse lipid species. This review presents recent advances on how plastoglobules are implicated in the biosynthesis and metabolism of vitamin E and K.

  6. Chloroplast genome sequence of the moss Tortula ruralis: gene content, polymorphism, and structural arrangement relative to other green plant chloroplast genomes

    Directory of Open Access Journals (Sweden)

    Wolf Paul G

    2010-02-01

    Full Text Available Abstract Background Tortula ruralis, a widely distributed species in the moss family Pottiaceae, is increasingly used as a model organism for the study of desiccation tolerance and mechanisms of cellular repair. In this paper, we present the chloroplast genome sequence of T. ruralis, only the second published chloroplast genome for a moss, and the first for a vegetatively desiccation-tolerant plant. Results The Tortula chloroplast genome is ~123,500 bp, and differs in a number of ways from that of Physcomitrella patens, the first published moss chloroplast genome. For example, Tortula lacks the ~71 kb inversion found in the large single copy region of the Physcomitrella genome and other members of the Funariales. Also, the Tortula chloroplast genome lacks petN, a gene found in all known land plant plastid genomes. In addition, an unusual case of nucleotide polymorphism was discovered. Conclusions Although the chloroplast genome of Tortula ruralis differs from that of the only other sequenced moss, Physcomitrella patens, we have yet to determine the biological significance of the differences. The polymorphisms we have uncovered in the sequencing of the genome offer a rare possibility (for mosses of the generation of DNA markers for fine-level phylogenetic studies, or to investigate individual variation within populations.

  7. Mesophyll Chloroplast Investment in C3, C4 and C2 Species of the Genus Flaveria.

    Science.gov (United States)

    Stata, Matt; Sage, Tammy L; Hoffmann, Natalie; Covshoff, Sarah; Ka-Shu Wong, Gane; Sage, Rowan F

    2016-05-01

    The mesophyll (M) cells of C4 plants contain fewer chloroplasts than observed in related C3 plants; however, it is uncertain where along the evolutionary transition from C3 to C4 that the reduction in M chloroplast number occurs. Using 18 species in the genus Flaveria, which contains C3, C4 and a range of C3-C4 intermediate species, we examined changes in chloroplast number and size per M cell, and positioning of chloroplasts relative to the M cell periphery. Chloroplast number and coverage of the M cell periphery declined in proportion to increasing strength of C4 metabolism in Flaveria, while chloroplast size increased with increasing C4 cycle strength. These changes increase cytosolic exposure to the cell periphery which could enhance diffusion of inorganic carbon to phosphenolpyruvate carboxylase (PEPC), a cytosolic enzyme. Analysis of the transcriptome from juvenile leaves of nine Flaveria species showed that the transcript abundance of four genes involved in plastid biogenesis-FtsZ1, FtsZ2, DRP5B and PARC6-was negatively correlated with variation in C4 cycle strength and positively correlated with M chloroplast number per planar cell area. Chloroplast size was negatively correlated with abundance of FtsZ1, FtsZ2 and PARC6 transcripts. These results indicate that natural selection targeted the proteins of the contractile ring assembly to effect the reduction in chloroplast numbers in the M cells of C4 Flaveria species. If so, efforts to engineer the C4 pathway into C3 plants might evaluate whether inducing transcriptome changes similar to those observed in Flaveria could reduce M chloroplast numbers, and thus introduce a trait that appears essential for efficient C4 function.

  8. The complete chloroplast genome provides insight into the evolution and polymorphism of Panax ginseng

    Directory of Open Access Journals (Sweden)

    Yongbing eZhao

    2015-01-01

    Full Text Available Panax ginseng C.A. Meyer (P. ginseng is an important medicinal plant and is often used in traditional Chinese medicine. With next generation sequencing (NGS technology, we determined the complete chloroplast genome sequences for four Chinese P. ginseng strains, which are Damaya (DMY, Ermaya (EMY, Gaolishen (GLS and Yeshanshen (YSS. The total chloroplast genome sequence length for DMY, EMY and GLS was 156,354 bp, while that for YSS was 156,355 bp. Comparative genomic analysis of the chloroplast genome sequences indicate that gene content, GC content, and gene order in DMY are quite similar to its relative species, and nucleotide sequence diversity of inverted repeat region (IR is lower than that of its counterparts, large single copy region (LSC and small single copy region (SSC. A comparison among these four P. ginseng strains revealed that the chloroplast genome sequences of DMY, EMY, and GLS were identical and YSS had a 1-bp insertion at base 5472. To further study the heterogeneity in chloroplast genome during domestication, high-resolution reads were mapped to the genome sequences to investigate the differences at the minor allele level; 208 minor allele sites with minor allele frequencies (MAF of ≥ 0.05 were identified. The polymorphism site numbers per kb of chloroplast genome sequence for DMY, EMY, GLS, and YSS were 0.74, 0.59, 0.97, and 1.23, respectively. All the minor allele sites located in LSC and IR regions, and the four strains showed the same variation types (substitution base or indel at all identified polymorphism sites. Comparison results of heterogeneity in the chloroplast genome sequences showed that the minor allele sites on the chloroplast genome were undergoing purifying selection to adapt to changing environment during domestication process. A study of P. ginseng chloroplast genome with particular focus on minor allele sites would aid in investigating the dynamics on the chloroplast genomes and different P. ginseng

  9. Membrane fusion

    DEFF Research Database (Denmark)

    Bendix, Pól Martin

    2015-01-01

    At Stanford University, Boxer lab, I worked on membrane fusion of small unilamellar lipid vesicles to flat membranes tethered to glass surfaces. This geometry closely resembles biological systems in which liposomes fuse to plasma membranes. The fusion mechanism was studied using DNA zippering...... between complementary strands linked to the two apposing membranes closely mimicking the zippering mechanism of SNARE fusion complexes....

  10. Biobased Membrane

    NARCIS (Netherlands)

    Koenders, E.A.B.; Zlopasa, J.; Picken, S.J.

    2015-01-01

    The present invention is in the field of a composition for forming a bio-compatible membrane applicable to building material, such as concrete, cement, etc., to a meth od of applying said composition for forming a bio-compatible membrane, a biocompatible membrane, use of said membrane for various pu

  11. SEQUENCE POLYMORPHISMS OF FOUR CHLOROPLAST GENES IN FOUR ACACIA SPECIES

    Directory of Open Access Journals (Sweden)

    Anthonius Y.P.B.C. Widyatmoko

    2011-06-01

    Full Text Available Sequence polymorphisms among and within four Acacia species,  A. aulacocarpa, A. auriculiformis, A. crassicarpa, and A. mangium, were investigated using four chloroplast DNA genes (atpA, petA, rbcL, and rpoA. The phylogenetic relationship among these species is discussed in light of the results of the sequence information. No intraspecific sequence variation was found in the four genes of the four species, and a conservative rate of mutation of the chloroplast DNA genes was also confirmed in the Acacia species. In the atpA and petA of the four genes, all four species possessed identical sequences, and no sequence variation was found among the four Acacia species. In the rbcL and rpoA genes, however, sequence polymorphisms were revealed among these species. Acacia aulacocarpa and A. crassicarpa shared an identical sequence, and A. auriculiformis and A. mangium also showed no sequence variation.  The fact that A. mangium and A. auriculiformis shared identical sequences as did A. aulacocarpa and A. crassicarpa indicated that the two respective species were extremely closely related. Although a putative natural hybrid of A. aulacocarpa and A. auriculiformis has been reported, our results suggested that natural hybridization should be further verified using molecular markers.

  12. Chloroplast DNA evolution and phylogenetic relationships in Lycopersicon.

    Science.gov (United States)

    Palmer, J D; Zamir, D

    1982-08-01

    Chloroplast DNA was purified from 12 accessions that represent most of the species diversity in the genus Lycopersicon (family Solanaceae) and from 3 closely related species in the genus Solanum. Fragment patterns produced by digestion of these DNAs with 25 different restriction endonucleases were analyzed by agarose gel electrophoresis. In all 15 DNAs, a total of only 39 restriction site mutations were detected among 484 restriction sites surveyed, representing 2,800 base pairs of sequence information. This low rate of base sequence change is paralleled by an extremely low rate of convergent change in restriction sites; only 1 of the 39 mutations appears to have occurred independently in two different lineages. Parsimony analysis of shared mutations has allowed the construction of a maternal phylogeny for the 15 accessions. This phylogeny is generally consistent with relationships based on morphology and crossability but provides more detailed resolution at several places. All accessions within Lycopersicon form a coherent group, with two of the three species of Solanum as outside reference points. Chloroplast DNA analysis places S. pennellii firmly within Lycopersicon, confirming recent studies that have removed it from Solanum. Red-orange fruit color is shown to be a monophyletic trait in three species of Lycopersicon, including the cultivated tomato, L. esculentum. Analysis of six accessions within L. peruvianum reveals a limited amount of intraspecific polymorphism which, however, encompasses all the variation observed in L. chilense and L. chmielewskii. It is suggested that these latter two accessions be relegated to positions within the L. peruvianum complex.

  13. The roles of toc34 and toc75 in targeting the toc159 preprotein receptor to chloroplasts.

    Science.gov (United States)

    Wallas, Tanya R; Smith, Matthew D; Sanchez-Nieto, Sobeida; Schnell, Danny J

    2003-11-07

    The Toc complex at the outer envelope of chloroplasts initiates the import of nuclear-encoded preproteins from the cytosol into the organelle. The core of the Toc complex is composed of two receptor GTPases, Toc159 and Toc34, as well as Toc75, a beta-barrel membrane channel. Toc159 is equally distributed between a soluble cytoplasmic form and a membrane-inserted form, suggesting that assembly of the Toc complex is dynamic. In the present study, we used the Arabidopsis thaliana orthologs of Toc159 and Toc34, atToc159 and atToc33, respectively, to investigate the requirements for assembly of the trimeric Toc complex. In addition to its intrinsic GTPase activity, we demonstrate that integration of atToc159 into the Toc complex requires atToc33 GTPase activity. Additionally, we show that the interaction of the two GTPase domains stimulates association of the membrane anchor of atToc159 with the translocon. Finally, we employ reconstituted proteoliposomes to demonstrate that proper insertion of the receptor requires both Toc75 and Toc34. Collectively these data suggest that Toc34 and Toc75 act sequentially to mediate docking and insertion of Toc159 resulting in assembly of the functional translocon.

  14. Chloroplast parameters differ in wild type and transgenic poplars overexpressing gsh1 in the cytosol.

    Science.gov (United States)

    Ivanova, L A; Ronzhina, D A; Ivanov, L A; Stroukova, L V; Peuke, A D; Rennenberg, H

    2009-07-01

    Poplar mutants overexpressing the bacterial genes gsh1 or gsh2 encoding the enzymes of glutathione biosynthesis are among the best-characterised transgenic plants. However, this characterisation originates exclusively from laboratory studies, and the performance of these mutants under field conditions is largely unknown. Here, we report a field experiment in which the wild-type poplar hybrid Populus tremula x P. alba and a transgenic line overexpressing the bacterial gene gsh1 encoding gamma-glutamylcysteine synthetase in the cytosol were grown for 3 years at a relatively clean (control) field site and a field site contaminated with heavy metals. Aboveground biomass accumulation was slightly smaller in transgenic compared to wild-type plants; soil contamination significantly decreased biomass accumulation in both wild-type and transgenic plants by more than 40%. Chloroplasts parameters, i.e., maximal diameter, projection area and perimeter, surface area and volume, surface/volume ratio and a two-dimensional form coefficient, were found to depend on plant type, leaf tissue and soil contamination. The greatest differences between wild and transgenic poplars were observed at the control site. Under these conditions, chloroplast sizes in palisade tissue of transgenic poplar significantly exceeded those of the wild type. In contrast to the wild type, palisade chloroplast volume exceeded that of spongy chloroplasts in transgenic poplars at both field sites. Chlorophyll content per chloroplast was the same in wild and transgenic poplars. Apparently, the increase in chloroplast volume was not connected to changes in the photosynthetic centres. Chloroplasts of transgenic poplar at the control site were more elongated in palisade cells and close to spherical in spongy mesophyll chloroplasts. At the contaminated site, palisade and spongy cell chloroplasts of leaves from transgenic trees and the wild type were the same shape. Transgenic poplars also had a smaller chloroplast

  15. Differential Contribution of Endoplasmic Reticulum and Chloroplast ω-3 Fatty Acid Desaturase Genes to the Linolenic Acid Content of Olive (Olea europaea) Fruit.

    Science.gov (United States)

    Hernández, M Luisa; Sicardo, M Dolores; Martínez-Rivas, José M

    2016-01-01

    Linolenic acid is a polyunsaturated fatty acid present in plant lipids, which plays key roles in plant metabolism as a structural component of storage and membrane lipids, and as a precursor of signaling molecules. The synthesis of linolenic acid is catalyzed by two different ω-3 fatty acid desaturases, which correspond to microsomal- (FAD3) and chloroplast- (FAD7 and FAD8) localized enzymes. We have investigated the specific contribution of each enzyme to the linolenic acid content in olive fruit. With that aim, we isolated two different cDNA clones encoding two ω-3 fatty acid desaturases from olive (Olea europaea cv. Picual). Sequence analysis indicates that they code for microsomal (OepFAD3B) and chloroplast (OepFAD7-2) ω-3 fatty acid desaturase enzymes, different from the previously characterized OekFAD3A and OekFAD7-1 genes. Functional expression in yeast of the corresponding OepFAD3A and OepFAD3B cDNAs confirmed that they encode microsomal ω-3 fatty acid desaturases. The linolenic acid content and transcript levels of olive FAD3 and FAD7 genes were measured in different tissues of Picual and Arbequina cultivars, including mesocarp and seed during development and ripening of olive fruit. Gene expression and lipid analysis indicate that FAD3A is the gene mainly responsible for the linolenic acid present in the seed, while FAD7-1 and FAD7-2 contribute mostly to the linolenic acid present in the mesocarp and, therefore, in the olive oil. These results also indicate the relevance of lipid trafficking between the endoplasmic reticulum and chloroplast in determining the linolenic acid content of membrane and storage lipids in oil-accumulating photosynthetic tissues.

  16. The complete chloroplast genome sequence of Helwingia himalaica (Helwingiaceae, Aquifoliales and a chloroplast phylogenomic analysis of the Campanulidae

    Directory of Open Access Journals (Sweden)

    Xin Yao

    2016-11-01

    Full Text Available Complete chloroplast genome sequences have been very useful for understanding phylogenetic relationships in angiosperms at the family level and above, but there are currently large gaps in coverage. We report the chloroplast genome for Helwingia himalaica, the first in the distinctive family Helwingiaceae and only the second genus to be sequenced in the order Aquifoliales. We then combine this with 36 published sequences in the large (c. 35,000 species subclass Campanulidae in order to investigate relationships at the order and family levels. The Helwingia genome consists of 158,362 bp containing a pair of inverted repeat (IR regions of 25,996 bp separated by a large single-copy (LSC region and a small single-copy (SSC region which are 87,810 and 18,560 bp, respectively. There are 142 known genes, including 94 protein-coding genes, eight ribosomal RNA genes, and 40 tRNA genes. The topology of the phylogenetic relationships between Apiales, Asterales, and Dipsacales differed between analyses based on complete genome sequences and on 36 shared protein-coding genes, showing that further studies of campanulid phylogeny are needed.

  17. A high-throughput method for detection of DNA in chloroplasts using flow cytometry

    Directory of Open Access Journals (Sweden)

    Oldenburg Delene J

    2007-03-01

    Full Text Available Abstract Background The amount of DNA in the chloroplasts of some plant species has been shown recently to decline dramatically during leaf development. A high-throughput method of DNA detection in chloroplasts is now needed in order to facilitate the further investigation of this process using large numbers of tissue samples. Results The DNA-binding fluorophores 4',6-diamidino-2-phenylindole (DAPI, SYBR Green I (SG, SYTO 42, and SYTO 45 were assessed for their utility in flow cytometric analysis of DNA in Arabidopsis chloroplasts. Fluorescence microscopy and real-time quantitative PCR (qPCR were used to validate flow cytometry data. We found neither DAPI nor SYTO 45 suitable for flow cytometric analysis of chloroplast DNA (cpDNA content, but did find changes in cpDNA content during development by flow cytometry using SG and SYTO 42. The latter dye provided more sensitive detection, and the results were similar to those from the fluorescence microscopic analysis. Differences in SYTO 42 fluorescence were found to correlate with differences in cpDNA content as determined by qPCR using three primer sets widely spaced across the chloroplast genome, suggesting that the whole genome undergoes copy number reduction during development, rather than selective reduction/degradation of subgenomic regions. Conclusion Flow cytometric analysis of chloroplasts stained with SYTO 42 is a high-throughput method suitable for determining changes in cpDNA content during development and for sorting chloroplasts on the basis of DNA content.

  18. The chloroplast genome of a symbiodinium sp. clade C3 isolate

    KAUST Repository

    Barbrook, Adrian C.

    2014-01-01

    Dinoflagellate algae of the genus Symbiodinium form important symbioses within corals and other benthic marine animals. Dinoflagellates possess an extremely reduced plastid genome relative to those examined in plants and other algae. In dinoflagellates the plastid genes are located on small plasmids, commonly referred to as \\'minicircles\\'. However, the chloroplast genomes of dinoflagellates have only been extensively characterised from a handful of species. There is also evidence of considerable variation in the chloroplast genome organisation across those species that have been examined. We therefore characterised the chloroplast genome from an environmental coral isolate, in this case containing a symbiont belonging to the Symbiodinium sp. clade C3. The gene content of the genome is well conserved with respect to previously characterised genomes. However, unlike previously characterised dinoflagellate chloroplast genomes we did not identify any \\'empty\\' minicircles. The sequences of this chloroplast genome show a high rate of evolution relative to other algal species. Particularly notable was a surprisingly high level of sequence divergence within the core polypeptides of photosystem I, the reasons for which are currently unknown. This chloroplast genome also possesses distinctive codon usage and GC content. These features suggest that chloroplast genomes in Symbiodinium are highly plastic. © 2013 Adrian C. Barbrook.

  19. A plant mitochondrial sequence transcribed in transgenic tobacco chloroplasts is not edited

    Energy Technology Data Exchange (ETDEWEB)

    Sutton, C.A.; Hanson, M.R. [Cornell Univ., Ithaca, NY (United States); Zoubenko, O.V.; Maliga, P. [State Univ. of New Jersey, Piscataway, NJ (United States)

    1995-03-01

    RNA editing occurs in two higher-plant organelles, chloroplasts, and mitochondria. Because chloroplasts and mitochondria exhibit some similarity in editing site selection, we investigated whether mitochondrial RNA sequences could be edited in chloroplasts. We produced transgenic tobacco plants that contained chimeric genes in which the second exon of a Petunia hybrida mitochondrial coxII gene was under the control of chloroplast gene regulatory sequences. coxII transcripts accumulated to low or high levels in transgenic chloroplasts containing chimeric genes with the plastid ribosomal protein gene rps16 or the rRNA operon promoter, respectively. Exon 2 of coxII was chosen because it carries seven editing sites and is edited in petunia mitochondria even when located in an abnormal context in an aberrant recombined gene. When editing of the coxII transcripts in transgenic chloroplasts was examined, no RNA editing at any of the usual sites was detected, nor was there any novel editing at any other sites. These results indicate that the RNA editing mechanisms of chloroplasts and mitochondria are not identical but must have at least some organelle-specific components. 33 refs., 5 figs.

  20. Is chloroplast movement in tobacco plants influenced systemically after local illumination or burning stress?

    Science.gov (United States)

    Naus, Jan; Rolencová, Monika; Hlavácková, Vladimíra

    2008-10-01

    Chloroplast movement has been studied in many plants mainly in relation to the local light, mechanical or stress effects. Here we investigated possible systemic responses of chloroplast movement to local light or burning stress in tobacco plants (Nicotiana tabacum cv. Samsun). Chloroplast movement was measured using two independent methods: one with a SPAD 502 Chlorophyll meter and another by collimated transmittance at a selected wavelength (676 nm). A sensitive periodic movement of chloroplasts was used in high or low (2 000 or 50 micromol/m(2) per s photosynthetically active radiation, respectively) cold white light with periods of 50 or 130 min. Measurements were carried out in the irradiated area, in the non-irradiated area of the same leaf or in the leaf located on the stem below the irradiated or burned one. No significant changes in systemic chloroplast movement in non-irradiated parts of the leaf and in the non-treated leaf were detected. Our data indicate that chloroplast movement in tobacco is dependent dominantly on the intensity and spectral composition of the incident light and on the local stimulation and state of the target tissue. No systemic signal was strong enough to evoke a detectable systemic response in chloroplast movement in distant untreated tissues of tobacco plants.

  1. Abscisic acid and blue light signaling pathways in chloroplast movements in Arabidopsis mesophyll.

    Science.gov (United States)

    Eckstein, Aleksandra; Krzeszowiec, Weronika; Banaś, Agnieszka Katarzyna; Janowiak, Franciszek; Gabryś, Halina

    2016-01-01

    Abscisic acid (ABA) and phototropins act antagonistically to control stomatal movements. Here, we investigated the role of ABA in phototropin-directed chloroplast movements in mesophyll cells of Arabidopsis thaliana. We analyzed the expression of phototropins at mRNA and protein level under the influence of ABA. PHOT1 mRNA level was decreased by ABA in the dark while it was insensitive to ABA in light. PHOT2 mRNA level was independent of the hormone treatment. The levels of phototropin proteins were down-regulated by ABA, both in darkness and light. No impact of exogenous ABA on amplitudes and kinetics of chloroplast movements was detected. Chloroplast responses in wild type Arabidopsis and three mutants, abi4, abi2 (abscisic acid insensitive4, 2) and aba1 (abscisic acid1), were measured to account for endogenous ABA signaling. The chloroplast responses were slightly reduced in abi2 and aba1 mutants in strong light. To further investigate the effect, abi2 and aba1 mutants were supplemented with exogenous ABA. In the aba1 mutant, the reaction was rescued but in abi2 it was unaffected. Our results show that ABA is not directly involved in phototropin-controlled chloroplast responses in mature leaves of Arabidopsis. However, the disturbance of ABA biosynthesis and signaling in mutants affects some elements of the chloroplast movement mechanism. In line with its role as a stress hormone, ABA appears to enhance plant sensitivity to light and promote the chloroplast avoidance response.

  2. Chloroplasts do not have a polarity for light-induced accumulation movement.

    Science.gov (United States)

    Tsuboi, Hidenori; Yamashita, Hiroko; Wada, Masamitsu

    2009-01-01

    Chloroplast photorelocation movement in green plants is generally mediated by blue light. However, in cryptogam plants, including ferns, mosses, and algae, both red light and blue light are effective. Although the photoreceptors required for this phenomenon have been identified, the mechanisms underlying this movement response are not yet known. In order to analyze this response in more detail, chloroplast movement was induced in dark-adapted Adiantum capillus-veneris gametophyte cells by partial cell irradiation with a microbeam of red and/or blue light. In each case, chloroplasts were found to move toward the microbeam-irradiated area. A second microbeam was also applied to the cell at a separate location before the chloroplasts had reached the destination of the first microbeam. Under these conditions, chloroplasts were found to change their direction of movement without turning and move toward the second microbeam-irradiated area after a lag time of a few minutes. These findings indicate that chloroplasts can move in any direction and do not exhibit a polarity for chloroplast accumulation movement. This phenomenon was analyzed in detail in Adiantum and subsequently confirmed in Arabidopsis thaliana palisade cells. Interestingly, the lag time for direction change toward the second microbeam in Adiantum was longer in the red light than in the blue light. However, the reason for this discrepancy is not yet understood.

  3. Is Chloroplast Movement in Tobacco Plants Influenced Systemically after Local Illumination or Burning Stress?

    Institute of Scientific and Technical Information of China (English)

    Jan Naus; Monika Rolencova; Vladimira Hlavackova

    2008-01-01

    Chloroplast movement has been studied In many plants mainly in relation to the local light, mechanical or stress effects. Here we investigated possible systemic responses of chloroplast movement to local light or burning stress in tobacco plants (Nicotiana tabacum cv. Samsun). Chloroplast movement was measured using two independent methods: one with a SPAD 502 Chlorophyll meter and another by collimated transmittance at a selected wavelength (676 nm). A sensitive pedodic movement of chloroplasts was used in high or low (2 000 or 50 μmol/m2 per s photosynthetically active radiation, respectively) cold white light with periods of 50 or 130 min. Measurements were carried out in the irradiated area, in the non-irradiated area of the same leaf or in the leaf located on the stem below the irradiated or burned one. No significant changes in systemic chloroplast movement in non-irradiated parts of the leaf and in the non-treated leaf were detected. Our data indicate that chloroplast movement in tobacco is dependent dominantly on the intensity and spectral composition of the incident light and on the local stimulation and state of the target tissue. No systemic signal was strong enough tovoke a detectable systemic response in chloroplast movement in distant untreated tissues of tobacco plants.

  4. Free radical generation and antioxidant content in chloroplasts from soybean leaves expsoed to ultraviolet-B

    Energy Technology Data Exchange (ETDEWEB)

    Galatro, A.; Simontacchi, M.; Puntarulo, S. [Univ. of Buenos Aires, School of Pharmacy and Biochemistry, Physical Chemistry, Buenos Aires (Argentina)

    2001-07-01

    The aim of this work was to study the effect of ultraviolet-B (UV-B) exposure on oxidative status in chloroplasts isolated from soybean (Glycine max cv. Hood). Chloroplasts were isolated from soybean leaves excised from either control seedlings or those exposed to 30 and 60 kJ m{sup -2} day{sup -1} of UV-B radiation for 4 days. Chloroplastic oxidative conditions were assessed as carbon-centered radical, carbonyl groups and ascorbyl radical content. Treatment with UV-B increased the carbon-centered radical-dependent EPR signal significantly by 55 and 100% in chloroplasts from leaves exposed to 30 and 60 kJ m{sup -2} day{sup -1} UV-B, respectively, compared to radical content in chloroplasts from control leaves. The content of carbonyl groups increased by 37 and 62% in chloroplasts isolated from soybean leaves irradiated for 4 days with 30 and 60 kJ m{sup -2} day{sup -1} UV-B, respectively. The content of soluble metabolites in isolated chloroplasts should not be taken as absolute in vivo values; however, these data are valuable for comparative studies. UV-B exposure did not significantly affect ascorbyl radical content compared to controls. The content of ascorbic acid and thiols in chloroplasts isolated from leaves exposed to 60 kJ m{sup -2} day{sup -1} UV-B was increased by 117 and 20.8%, respectively, compared to controls. Neither the content of total carotene nor that of {beta}-carotene or {alpha}-tocopherol was affected by the irradiation. The results: presented here suggest that the increased content of lipid radicals and oxidized proteins in the chloroplasts isolated from leaves exposed to UV-B could be ascribed to both the lack of antioxidant response in the lipid soluble fraction and the modest increase in the soluble antioxidant content. (au)

  5. Transcriptome analysis of ectopic chloroplast development in green curd cauliflower (Brassica oleracea L. var. botrytis

    Directory of Open Access Journals (Sweden)

    Zhou Xiangjun

    2011-11-01

    Full Text Available Abstract Background Chloroplasts are the green plastids where photosynthesis takes place. The biogenesis of chloroplasts requires the coordinate expression of both nuclear and chloroplast genes and is regulated by developmental and environmental signals. Despite extensive studies of this process, the genetic basis and the regulatory control of chloroplast biogenesis and development remain to be elucidated. Results Green cauliflower mutant causes ectopic development of chloroplasts in the curd tissue of the plant, turning the otherwise white curd green. To investigate the transcriptional control of chloroplast development, we compared gene expression between green and white curds using the RNA-seq approach. Deep sequencing produced over 15 million reads with lengths of 86 base pairs from each cDNA library. A total of 7,155 genes were found to exhibit at least 3-fold changes in expression between green and white curds. These included light-regulated genes, genes encoding chloroplast constituents, and genes involved in chlorophyll biosynthesis. Moreover, we discovered that the cauliflower ELONGATED HYPOCOTYL5 (BoHY5 was expressed higher in green curds than white curds and that 2616 HY5-targeted genes, including 1600 up-regulated genes and 1016 down-regulated genes, were differently expressed in green in comparison to white curd tissue. All these 1600 up-regulated genes were HY5-targeted genes in the light. Conclusions The genome-wide profiling of gene expression by RNA-seq in green curds led to the identification of large numbers of genes associated with chloroplast development, and suggested the role of regulatory genes in the high hierarchy of light signaling pathways in mediating the ectopic chloroplast development in the green curd cauliflower mutant.

  6. Photosynthesis of root chloroplasts developed in Arabidopsis lines overexpressing GOLDEN2-LIKE transcription factors.

    Science.gov (United States)

    Kobayashi, Koichi; Sasaki, Daichi; Noguchi, Ko; Fujinuma, Daiki; Komatsu, Hirohisa; Kobayashi, Masami; Sato, Mayuko; Toyooka, Kiminori; Sugimoto, Keiko; Niyogi, Krishna K; Wada, Hajime; Masuda, Tatsuru

    2013-08-01

    In plants, genes involved in photosynthesis are encoded separately in nuclei and plastids, and tight cooperation between these two genomes is therefore required for the development of functional chloroplasts. Golden2-like (GLK) transcription factors are involved in chloroplast development, directly targeting photosynthesis-associated nuclear genes for up-regulation. Although overexpression of GLKs leads to chloroplast development in non-photosynthetic organs, the mechanisms of coordination between the nuclear gene expression influenced by GLKs and the photosynthetic processes inside chloroplasts are largely unknown. To elucidate the impact of GLK-induced expression of photosynthesis-associated nuclear genes on the construction of photosynthetic systems, chloroplast morphology and photosynthetic characteristics in greenish roots of Arabidopsis thaliana lines overexpressing GLKs were compared with those in wild-type roots and leaves. Overexpression of GLKs caused up-regulation of not only their direct targets but also non-target nuclear and plastid genes, leading to global induction of chloroplast biogenesis in the root. Large antennae relative to reaction centers were observed in wild-type roots and were further enhanced by GLK overexpression due to the increased expression of target genes associated with peripheral light-harvesting antennae. Photochemical efficiency was lower in the root chloroplasts than in leaf chloroplasts, suggesting that the imbalance in the photosynthetic machinery decreases the efficiency of light utilization in root chloroplasts. Despite the low photochemical efficiency, root photosynthesis contributed to carbon assimilation in Arabidopsis. Moreover, GLK overexpression increased CO₂ fixation and promoted phototrophic performance of the root, showing the potential of root photosynthesis to improve effective carbon utilization in plants.