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Sample records for chloroplast differentiation annual

  1. Nitrogen control of chloroplast differentiation. Annual progress report

    SciTech Connect

    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.

  2. Nitrogen control of chloroplast development and differentiation. Annual progress report

    SciTech Connect

    Schmidt, G.W.

    1991-12-01

    The growth and development of plants and photosynthetic microorganisms is commonly limited by the availability of nitrogen. Our work concerns understanding the mechanisms by which plants and algae that are subjected to nitrogen deprivation alter the composition of photosynthetic membranes and enzymes involved in photosynthetic carbon metabolism. Toward these ends, we study biosynthetic and gene expression processes in the unicellular green alga Chlamydomonas reinhardtii which is grown in an ammonium-limited continuous culture system. We have found that the expression of nuclear genes, including those encoding for light-harvesting proteins, are severely repressed in nitrogen-limited cells whereas, in general, chloroplast protein synthesis is attenuated primarily at the level of mRNA translation. Conversely, nitrogen deprivation appears to lead to enhanced synthesis of enzymes that are involved in starch and storage lipid deposition. In addition, as a possible means by which photosynthetic electron transport activities and ATP synthesis is sustained during chronic periods of nitrogen deprivation, thylakoid membranes become enriched with components for chlororespiration. Characterization of the chlororespiratory electron transport constituents, including cytochrome complexes and NAD(P)H dehydrogenase is a major current effort. Also, we are striving to isolate the genes encoding chlororespiration proteins toward determining how they and others that are strongly responsive to nutrient availability are regulated.

  3. Dynamics of Chloroplast Translation during Chloroplast Differentiation in Maize

    PubMed Central

    Chotewutmontri, Prakitchai; Barkan, Alice

    2016-01-01

    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

  4. Nitrogen control of chloroplast differentiation

    SciTech Connect

    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.

  5. Nitrogen control of chloroplast differentiation. Final report

    SciTech Connect

    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.

  6. Nitrogen control of chloroplast development and differentiation

    SciTech Connect

    Schmidt, G.W.

    1991-12-01

    The growth and development of plants and photosynthetic microorganisms is commonly limited by the availability of nitrogen. Our work concerns understanding the mechanisms by which plants and algae that are subjected to nitrogen deprivation alter the composition of photosynthetic membranes and enzymes involved in photosynthetic carbon metabolism. Toward these ends, we study biosynthetic and gene expression processes in the unicellular green alga Chlamydomonas reinhardtii which is grown in an ammonium-limited continuous culture system. We have found that the expression of nuclear genes, including those encoding for light-harvesting proteins, are severely repressed in nitrogen-limited cells whereas, in general, chloroplast protein synthesis is attenuated primarily at the level of mRNA translation. Conversely, nitrogen deprivation appears to lead to enhanced synthesis of enzymes that are involved in starch and storage lipid deposition. In addition, as a possible means by which photosynthetic electron transport activities and ATP synthesis is sustained during chronic periods of nitrogen deprivation, thylakoid membranes become enriched with components for chlororespiration. Characterization of the chlororespiratory electron transport constituents, including cytochrome complexes and NAD(P)H dehydrogenase is a major current effort. Also, we are striving to isolate the genes encoding chlororespiration proteins toward determining how they and others that are strongly responsive to nutrient availability are regulated.

  7. The PLASTID DIVISION1 and 2 Components of the Chloroplast Division Machinery Determine the Rate of Chloroplast Division in Land Plant Cell Differentiation[C][W

    PubMed Central

    Okazaki, Kumiko; Kabeya, Yukihiro; Suzuki, Kenji; Mori, Toshiyuki; Ichikawa, Takanari; Matsui, Minami; Nakanishi, Hiromitsu; Miyagishima, Shin-ya

    2009-01-01

    In most algae, the chloroplast division rate is held constant to maintain the proper number of chloroplasts per cell. By contrast, land plants evolved cell and chloroplast differentiation systems in which the size and number of chloroplasts change along with their respective cellular function by regulation of the division rate. Here, we show that PLASTID DIVISION (PDV) proteins, land plant–specific components of the division apparatus, determine the rate of chloroplast division. Overexpression of PDV proteins in the angiosperm Arabidopsis thaliana and the moss Physcomitrella patens increased the number but decreased the size of chloroplasts; reduction of PDV levels resulted in the opposite effect. The level of PDV proteins, but not other division components, decreased during leaf development, during which the chloroplast division rate also decreased. Exogenous cytokinins or overexpression of the cytokinin-responsive transcription factor CYTOKININ RESPONSE FACTOR2 increased the chloroplast division rate, where PDV proteins, but not other components of the division apparatus, were upregulated. These results suggest that the integration of PDV proteins into the division machinery enabled land plant cells to change chloroplast size and number in accord with the fate of cell differentiation. PMID:19567705

  8. Functional Differentiation of Bundle Sheath and Mesophyll Maize Chloroplasts Determined by Comparative ProteomicsW⃞

    PubMed Central

    Majeran, Wojciech; Cai, Yang; Sun, Qi; van Wijk, Klaas J.

    2005-01-01

    Chloroplasts of maize (Zea mays) leaves differentiate into specific bundle sheath (BS) and mesophyll (M) types to accommodate C4 photosynthesis. Consequences for other plastid functions are not well understood but are addressed here through a quantitative comparative proteome analysis of purified M and BS chloroplast stroma. Three independent techniques were used, including cleavable stable isotope coded affinity tags. Enzymes involved in lipid biosynthesis, nitrogen import, and tetrapyrrole and isoprenoid biosynthesis are preferentially located in the M chloroplasts. By contrast, enzymes involved in starch synthesis and sulfur import preferentially accumulate in BS chloroplasts. The different soluble antioxidative systems, in particular peroxiredoxins, accumulate at higher levels in M chloroplasts. We also observed differential accumulation of proteins involved in expression of plastid-encoded proteins (e.g., EF-Tu, EF-G, and mRNA binding proteins) and thylakoid formation (VIPP1), whereas others were equally distributed. Enzymes related to the C4 shuttle, the carboxylation and regeneration phase of the Calvin cycle, and several regulators (e.g., CP12) distributed as expected. However, enzymes involved in triose phosphate reduction and triose phosphate isomerase are primarily located in the M chloroplasts, indicating that the M-localized triose phosphate shuttle should be viewed as part of the BS-localized Calvin cycle, rather than a parallel pathway. PMID:16243905

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

    PubMed Central

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

    1993-01-01

    ripened either in the dark or in the light. These results suggest that the lesion in gf may alleviate conditions associated with chloroplast deterioration during the chloroplast-chromoplast transition in tomato ripening but has no direct effect on chromoplast differentiation per se. The ultrastructure of gf provides unequivocal evidence that, in ripening tomato, chromoplasts indeed differentiate from preexisting chloroplasts; on the other hand, chromoplast differentiation in the dark-matured and -ripened tomato fruits indicates that chromoplast development can be a process entirely independent of the chloroplasts. PMID:12231777

  10. Differential positioning of C(4) mesophyll and bundle sheath chloroplasts: recovery of chloroplast positioning requires the actomyosin system.

    PubMed

    Kobayashi, Hiroaki; Yamada, Masahiro; Taniguchi, Mitsutaka; Kawasaki, Michio; Sugiyama, Tatsuo; Miyake, Hiroshi

    2009-01-01

    In C(4) plants, bundle sheath (BS) chloroplasts are arranged in the centripetal position or in the centrifugal position, although mesophyll (M) chloroplasts are evenly distributed along cell membranes. To examine the molecular mechanism for the intracellular disposition of these chloroplasts, we observed the distribution of actin filaments in BS and M cells of the C(4) plants finger millet (Eleusine coracana) and maize (Zea mays) using immunofluorescence. Fine actin filaments encircled chloroplasts in both cell types, and an actin network was observed adjacent to plasma membranes. The intracellular disposition of both chloroplasts in finger millet was disrupted by centrifugal force but recovered within 2 h in the dark. Actin filaments remained associated with chloroplasts during recovery. We also examined the effects of inhibitors on the rearrangement of chloroplasts. Inhibitors of actin polymerization, myosin-based activities and cytosolic protein synthesis blocked migration of chloroplasts. In contrast, a microtubule-depolymerizing drug had no effect. These results show that C(4) plants possess a mechanism for keeping chloroplasts in the home position which is dependent on the actomyosin system and cytosolic protein synthesis but not tubulin or light. PMID:19022806

  11. Exploring mechanisms linked to differentiation and function of dimorphic chloroplasts in the single cell C4 species Bienertia sinuspersici

    PubMed Central

    2014-01-01

    Background In the model single-cell C4 plant Bienertia sinuspersici, chloroplast- and nuclear-encoded photosynthetic enzymes, characteristically confined to either bundle sheath or mesophyll cells in Kranz-type C4 leaves, all occur together within individual leaf chlorenchyma cells. Intracellular separation of dimorphic chloroplasts and key enzymes within central and peripheral compartments allow for C4 carbon fixation analogous to NAD-malic enzyme (NAD-ME) Kranz type species. Several methods were used to investigate dimorphic chloroplast differentiation in B. sinuspersici. Results Confocal analysis revealed that Rubisco-containing chloroplasts in the central compartment chloroplasts (CCC) contained more photosystem II proteins than the peripheral compartment chloroplasts (PCC) which contain pyruvate,Pi dikinase (PPDK), a pattern analogous to the cell type-specific chloroplasts of many Kranz type NAD-ME species. Transient expression analysis using GFP fusion constructs containing various lengths of a B. sinuspersici Rubisco small subunit (RbcS) gene and the transit peptide of PPDK revealed that their import was not specific to either chloroplast type. Immunolocalization showed the rbcL-specific mRNA binding protein RLSB to be selectively localized to the CCC in B. sinuspersici, and to Rubisco-containing BS chloroplasts in the closely related Kranz species Suaeda taxifolia. Comparative fluorescence analyses were made using redox-sensitive and insensitive GFP forms, as well comparative staining using the peroxidase indicator 3,3-diaminobenzidine (DAB), which demonstrated differences in stromal redox potential, with the CCC having a more negative potential than the PCC. Conclusions Both CCC RLSB localization and the differential chloroplast redox state are suggested to have a role in post-transcriptional rbcL expression. PMID:24443986

  12. Consequences of C4 Differentiation for Chloroplast Membrane Proteomes in Maize Mesophyll and Bundle Sheath Cells *S⃞

    PubMed Central

    Majeran, Wojciech; Zybailov, Boris; Ytterberg, A. Jimmy; Dunsmore, Jason; Sun, Qi; van Wijk, Klaas J.

    2008-01-01

    Chloroplasts of maize leaves differentiate into specific bundle sheath (BS) and mesophyll (M) types to accommodate C4 photosynthesis. Chloroplasts contain thylakoid and envelope membranes that contain the photosynthetic machineries and transporters but also proteins involved in e.g. protein homeostasis. These chloroplast membranes must be specialized within each cell type to accommodate C4 photosynthesis and regulate metabolic fluxes and activities. This quantitative study determined the differentiated state of BS and M chloroplast thylakoid and envelope membrane proteomes and their oligomeric states using innovative gel-based and mass spectrometry-based protein quantifications. This included native gels, iTRAQ, and label-free quantification using an LTQ-Orbitrap. Subunits of Photosystems I and II, the cytochrome b6f, and ATP synthase complexes showed average BS/M accumulation ratios of 1.6, 0.45, 1.0, and 1.33, respectively, whereas ratios for the light-harvesting complex I and II families were 1.72 and 0.68, respectively. A 1000-kDa BS-specific NAD(P)H dehydrogenase complex with associated proteins of unknown function containing more than 15 proteins was observed; we speculate that this novel complex possibly functions in inorganic carbon concentration when carboxylation rates by ribulose-bisphosphate carboxylase/oxygenase are lower than decarboxylation rates by malic enzyme. Differential accumulation of thylakoid proteases (Egy and DegP), state transition kinases (STN7,8), and Photosystem I and II assembly factors was observed, suggesting that cell-specific photosynthetic electron transport depends on post-translational regulatory mechanisms. BS/M ratios for inner envelope transporters phosphoenolpyruvate/Pi translocator, Dit1, Dit2, and Mex1 were determined and reflect metabolic fluxes in carbon metabolism. A wide variety of hundreds of other proteins showed differential BS/M accumulation. Mass spectral information and functional annotations are available

  13. Phosphoinositides Play Differential Roles in Regulating Phototropin1- and Phototropin2-Mediated Chloroplast Movements in Arabidopsis

    PubMed Central

    Aggarwal, Chhavi; Łabuz, Justyna; Gabryś, Halina

    2013-01-01

    Phototropins are UVA/blue-light receptors involved in controlling the light-dependent physiological responses which serve to optimize the photosynthetic activity of plants and promote growth. The phototropin-induced phosphoinositide (PI) metabolism has been shown to be essential for stomatal opening and phototropism. However, the role of PIs in phototropin-induced chloroplast movements remains poorly understood. The aim of this work is to determine which PI species are involved in the control of chloroplast movements in Arabidopsis and the nature of their involvement. We present the effects of the inactivation of phospholipase C (PLC), PI3-kinase (PI3K) and PI4-kinase (PI4K) on chloroplast relocations in Arabidopsis. The inhibition of the phosphatidylinositol 4,5-bisphospahte [PI(4,5)P2]-PLC pathway, using neomycin and U73122, suppressed the phot2-mediated chloroplast accumulation and avoidance responses, without affecting movement responses controlled by phot1. On the other hand, PI3K and PI4K activities are more restricted to phot1- and phot2-induced weak-light responses. The inactivation of PI3K and PI4K by wortmannin and LY294002 severely affected the weak blue-light-activated accumulation response but had little effect on the strong blue-light-activated avoidance response. The inhibitory effect observed with PI metabolism inhibitors is, at least partly, due to a disturbance in Ca2+(c) signaling. Using the transgenic aequorin system, we show that the application of these inhibitors suppresses the blue-light-induced transient Ca2+(c) rise. These results demonstrate the importance of PIs in chloroplast movements, with the PI(4,5)P2-PLC pathway involved in phot2 signaling while PI3K and PI4K are required for the phot1- and phot2-induced accumulation response. Our results suggest that these PIs modulate cytosolic Ca2+ signaling during movements. PMID:23405144

  14. Differential Subplastidial Localization and Turnover of Enzymes Involved in Isoprenoid Biosynthesis in Chloroplasts

    PubMed Central

    Perello, Catalina; Llamas, Ernesto; Burlat, Vincent; Ortiz-Alcaide, Miriam; Phillips, Michael A.; Pulido, Pablo; Rodriguez-Concepcion, Manuel

    2016-01-01

    Plastidial isoprenoids are a diverse group of metabolites with roles in photosynthesis, growth regulation, and interaction with the environment. The methylerythritol 4-phosphate (MEP) pathway produces the metabolic precursors of all types of plastidial isoprenoids. Proteomics studies in Arabidopsis thaliana have shown that all the enzymes of the MEP pathway are localized in the plastid stroma. However, immunoblot analysis of chloroplast subfractions showed that the first two enzymes of the pathway, deoxyxylulose 5-phosphate synthase (DXS) and reductoisomerase (DXR), can also be found in non-stromal fractions. Both transient and stable expression of GFP-tagged DXS and DXR proteins confirmed the presence of the fusion proteins in distinct subplastidial compartments. In particular, DXR-GFP was found to accumulate in relatively large vesicles that could eventually be released from chloroplasts, presumably to be degraded by an autophagy-independent process. Together, we propose that protein-specific mechanisms control the localization and turnover of the first two enzymes of the MEP pathway in Arabidopsis chloroplasts. PMID:26919668

  15. Differential Subplastidial Localization and Turnover of Enzymes Involved in Isoprenoid Biosynthesis in Chloroplasts.

    PubMed

    Perello, Catalina; Llamas, Ernesto; Burlat, Vincent; Ortiz-Alcaide, Miriam; Phillips, Michael A; Pulido, Pablo; Rodriguez-Concepcion, Manuel

    2016-01-01

    Plastidial isoprenoids are a diverse group of metabolites with roles in photosynthesis, growth regulation, and interaction with the environment. The methylerythritol 4-phosphate (MEP) pathway produces the metabolic precursors of all types of plastidial isoprenoids. Proteomics studies in Arabidopsis thaliana have shown that all the enzymes of the MEP pathway are localized in the plastid stroma. However, immunoblot analysis of chloroplast subfractions showed that the first two enzymes of the pathway, deoxyxylulose 5-phosphate synthase (DXS) and reductoisomerase (DXR), can also be found in non-stromal fractions. Both transient and stable expression of GFP-tagged DXS and DXR proteins confirmed the presence of the fusion proteins in distinct subplastidial compartments. In particular, DXR-GFP was found to accumulate in relatively large vesicles that could eventually be released from chloroplasts, presumably to be degraded by an autophagy-independent process. Together, we propose that protein-specific mechanisms control the localization and turnover of the first two enzymes of the MEP pathway in Arabidopsis chloroplasts. PMID:26919668

  16. Transposon-induced nuclear mutations that alter chloroplast gene expression. Annual report, September 1, 1991--August 31, 1992

    SciTech Connect

    Barkan, A.

    1992-12-31

    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.

  17. Genetic diversity and differentiation in Prunus species (Rosaceae) using chloroplast and mitochondrial DNA CAPS markers.

    PubMed

    Ben Mustapha, S; Ben Tamarzizt, H; Baraket, G; Abdallah, D; Salhi Hannachi, A

    2015-01-01

    Chloroplast (cpDNA) and mitochondrial DNA (mtDNA) were analyzed to establish genetic relationships among Tunisian plum cultivars using the polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) technique. Two mtDNA regions (nad 1 b/c and nad 4 1/2) and a cpDNA region (trnL-trnF) were amplified and digested using restriction enzymes. Seventy and six polymorphic sites were revealed in cpDNA and mtDNA, respectively. As a consequence, cpDNA appears to be more polymorphic than mtDNA. The unweighted pair group method with arithmetic mean (UPGMA) dendrogram showed that accessions were distributed independently of their geographical origin, and introduced and local cultivars appear to be closely related. Both UPGMA and principal component analysis grouped Tunisian plum accessions into similar clusters. The analysis of the pooled sequences allowed the detection of 17 chlorotypes and 12 mitotypes. The unique haplotypes detected for cultivars are valuable for management and preservation of the plum local resources. From this study, PCR-RFLP analysis appears to be a useful approach to detect and identify cytoplasmic variation in plum trees. Our results also provide useful information for the management of genetic resources and to establish a program to improve the genetic resources available for plums. PMID:25966190

  18. Plastid Transcriptomics and Translatomics of Tomato Fruit Development and Chloroplast-to-Chromoplast Differentiation: Chromoplast Gene Expression Largely Serves the Production of a Single Protein[W][OA

    PubMed Central

    Kahlau, Sabine; Bock, Ralph

    2008-01-01

    Plastid genes are expressed at high levels in photosynthetically active chloroplasts but are generally believed to be drastically downregulated in nongreen plastids. The genome-wide changes in the expression patterns of plastid genes during the development of nongreen plastid types as well as the contributions of transcriptional versus translational regulation are largely unknown. We report here a systematic transcriptomics and translatomics analysis of the tomato (Solanum lycopersicum) plastid genome during fruit development and chloroplast-to-chromoplast conversion. At the level of RNA accumulation, most but not all plastid genes are strongly downregulated in fruits compared with leaves. By contrast, chloroplast-to-chromoplast differentiation during fruit ripening is surprisingly not accompanied by large changes in plastid RNA accumulation. However, most plastid genes are translationally downregulated during chromoplast development. Both transcriptional and translational downregulation are more pronounced for photosynthesis-related genes than for genes involved in gene expression, indicating that some low-level plastid gene expression must be sustained in chromoplasts. High-level expression during chromoplast development identifies accD, the only plastid-encoded gene involved in fatty acid biosynthesis, as the target gene for which gene expression activity in chromoplasts is maintained. In addition, we have determined the developmental patterns of plastid RNA polymerase activities, intron splicing, and RNA editing and report specific developmental changes in the splicing and editing patterns of plastid transcripts. PMID:18441214

  19. Transposon-induced nuclear mutations that alter chloroplast gene expression. Annual report, September 1, 1992--April 15, 1993

    SciTech Connect

    Barkan, A.

    1993-04-20

    The goal of this project is to use mutant phenotypes as a guide to nuclear genes that control the timing and cell-type specificity of chloroplast gene expression. Studies are being conducted with nuclear mutants of maize that are defective in the biogenesis or translation of chloroplast mRNAs. Currently studies are focused on two nuclear mutants with specific and unique lesions in chloroplast RNA processing (crp mutants). Crp1 mutants (formerly called hcf136) fail to accumulate the cytochrome f/b6 complex. The protein loss is due to a defect in the metabolism of transcripts encoding the petB and petD gene products, two subunits of the missing complex. Mutant seedlings lack the monocistronic petB and petD MRNAS, which both arise in nominal plants by endonucleolytic cleavage of the polycistronic primary transcript of the psbB gene cluster. Precursor mRNAs accumulate normally in crp1, indicating that its defect is due either to a failure to cleave the precursors, or a failure to stabilize the fully processed mRNAs. We are interested in both the biochemistry of this site-specific RNA processing and in the role of the processing in generating translatable mRNAs. To address the latter, we are quantifying the rates of synthesis of the petB and petD gene products with the goal of determining whether the missing transcripts are more efficiently translated than their precursors. To address the biochemistry of the defect in RNA metabolism, the crp1 gene is being cloned via the transposon tag. crp2 (formerly called hcf142) lacks the predominant mRNA encoding petA, but appears to be otherwise unimpaired in chloroplast RNA metabolism. The precise role of crp2 in synthesizing or stabilizing the petA mRNA is being investigated through biochemical studies.

  20. Genetic differentiation and admixture among Eurasian and North American Leymus (Triticeae) wildryes detected using chloroplast DNA sequences and AFLP profiles

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Leymus is a genomically defined group of allopolyploid Triticeae taxa that show homologous chromosome pairing when hybridized and contain the Ns and Xm subgenomes. Recent investigations showed that Leymus chloroplast DNA sequences are polyphyletic with most Eurasian taxa similar to the Psathyrostac...

  1. Genetic Differentiation and Hybridization Among Festuca Idahoensis, F. Roemeri, and F. Ovina Detected From AFLP, ITS, and Chloroplast DNA

    Technology Transfer Automated Retrieval System (TEKTRAN)

    North American forms of the F. ovina complex, Festuca idahoensis and F. roemert are distributed broadly east and narrowly west of the Cascade Mountains, respectively. The psbA-trnH and rps16-trnK chloroplast DNA intergenic sequences, 18S-5.8S-26S nuclear ribosomal DNA internal transcribed spacer (I...

  2. Abscisic acid represses the transcription of chloroplast genes*

    PubMed Central

    Yamburenko, Maria V.; Zubo, Yan O.; Börner, Thomas

    2013-01-01

    Numerous studies have shown effects of abscisic acid (ABA) on nuclear genes encoding chloroplast-localized proteins. ABA effects on the transcription of chloroplast genes, however, have not been investigated yet thoroughly. This work, therefore, studied the effects of ABA (75 μM) on transcription and steady-state levels of transcripts in chloroplasts of basal and apical segments of primary leaves of barley (Hordeum vulgare L.). Basal segments consist of young cells with developing chloroplasts, while apical segments contain the oldest cells with mature chloroplasts. Exogenous ABA reduced the chlorophyll content and caused changes of the endogenous concentrations not only of ABA but also of cytokinins to different extents in the basal and apical segments. It repressed transcription by the chloroplast phage-type and bacteria-type RNA polymerases and lowered transcript levels of most investigated chloroplast genes drastically. ABA did not repress the transcription of psbD and a few other genes and even increased psbD mRNA levels under certain conditions. The ABA effects on chloroplast transcription were more pronounced in basal vs. apical leaf segments and enhanced by light. Simultaneous application of cytokinin (22 μM 6-benzyladenine) minimized the ABA effects on chloroplast gene expression. These data demonstrate that ABA affects the expression of chloroplast genes differentially and points to a role of ABA in the regulation and coordination of the activities of nuclear and chloroplast genes coding for proteins with functions in photosynthesis. PMID:24078671

  3. Classification and differentiation of the genus Peganum indigenous to China based on chloroplast trnL-F and psbA-trnH sequences and seed coat morphology.

    PubMed

    Zhao, T; Wang, Z-T; Branford-White, C J; Xu, H; Wang, C-H

    2011-11-01

    The genus Peganum (Zygophyllaceae) consists of six species and one subspecies; three of which are distributed in China, P. harmala Linn, P. nigellastrum Bunge and P. multisectum (Maxim.) Bobr. A probable new or intermediate species, Peganum sp., has been suggested in the wild in northwest China. Traditional classification in genus Peganum has focused on hairs on the plant surface. In this study, seed coat characteristics of Peganum species were investigated using light and scanning electron microscopy, demonstrating clear differences in morphology between species. In addition, DNA sequence data from two sequences (chloroplast: trnL-F, psbA-trnH) were used to differentiate Peganum sp. and study polygenetic relationships. Diversity in DNA sequences among Peganum species was found, with inter-specific sequence divergence ranging from 0.6% to 5.6% in psbA-trnH, and 0.0% to 1.8% in trnL-F. The variations within species were low: from 0.0% to 0.4% in psbA-trnH and 0.0% to 0.4% in trnL-F. Therefore, Peganum species can now be easily identified as separate entities based on variations in DNA sequences. Phylogenetic trees were constructed from the combined data set for the two gene fragments, and the results indicate that Peganum sp. is monophyletic and sister to P. harmala and P. nigellastrum, while P. multisectum is also monophyletic. DNA data further confirmed that P. multisectum is an independent species and that a new species, Peganum sp., exists within the genus Peganum growing wild in China. PMID:21972911

  4. Chloroplast and Cytoplasmic Enzymes

    PubMed Central

    Anderson, Louise E.; Pacold, Ivan

    1972-01-01

    Several peaks of aldolase activity are found in the isoelectric focusing pattern of pea (Pisum sativum) leaf chloroplast extracts. One peak, separated by 0.5 pH unit from the major chloroplast aldolase peak, is found when cytoplasmic extracts are focused. The chloroplast and cytoplasmic enzymes have a pH 7.4 optimum with fructose 1,6-diphosphate. The Michaelis constant for fructose-1,6-diphosphate is 19 μM for the chloroplast, 21 μM for the cytoplasmic enzyme, and for sedoheptulose 1,7-diphosphate, 8 μM for the chloroplast enzyme, 18 μM for the cytoplasmic enzyme. Both enzymes are inhibited by d-glyceraldehyde 3-phosphate and by ribulose 1,5-diphosphate. The similarity in the catalytic properties of the isoenzymes suggests that both enzymes have an amphibolic role in carbon metabolism in the green leaf. PMID:16657968

  5. Auxin and chloroplast movements.

    PubMed

    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. PMID:26467664

  6. Activation of Polyphenol Oxidase of Chloroplasts 1

    PubMed Central

    Tolbert, N. E.

    1973-01-01

    Polyphenol oxidase of leaves is located mainly in chloroplasts isolated by differential or sucrose density gradient centrifugation. This activity is part of the lamellar structure that is not lost on repeated washing of the plastids. The oxidase activity was stable during prolonged storage of the particles at 4 C or —18 C. The Km (dihydroxyphenylalanine) for spinach leaf polyphenol oxidase was 7 mm by a spectrophotometric assay and 2 mm by the manometric assay. Polyphenol oxidase activity in the leaf peroxisomal fraction, after isopycnic centrifugation on a linear sucrose gradient, did not coincide with the peroxisomal enzymes but was attributed to proplastids at nearly the same specific density. Plants were grouped by the latency properties for polyphenol oxidase in their isolated chloroplasts. In a group including spinach, Swiss chard, and beet leaves the plastids immediately after preparation from fresh leaves required a small amount of light for maximal rates of oxidation of dihydroxyphenylalanine. Polyphenol oxidase activity in the dark or light increased many fold during aging of these chloroplasts for 1 to 5 days. Soluble polyphenol oxidase of the cytoplasm was not so stimulated. Chloroplasts prepared from stored leaves were also much more active than from fresh leaves. Maximum rates of dihydroxyphenylalanine oxidation were 2 to 6 mmoles × mg−1 chlorophyll × hr−1. Equal stimulation of latent polyphenol oxidase in fresh or aged chloroplasts in this group was obtained by either light, an aged trypsin digest, 3-(4-chlorophenyl)-1, 1-dimethylurea, or antimycin A. A variety of other treatments did not activate or had little effect on the oxidase, including various peptides, salts, detergents, and other proteolytic enzymes. Activation of latent polyphenol oxidase in spinach chloroplasts by trypsin amounted to as much as 30-fold. The trypsin activation occurred even after the trypsin had been treated with 10% trichloroacetic acid, 1.0 n HCl or boiled for 30

  7. Betaine synthesis in chenopods: localization in chloroplasts

    SciTech Connect

    Hanson, A.D.; May A.M.; Grumet, R.; Bode, J.; Jamieson, G.C.; Rhodes, D.

    1985-06-01

    Plants from several families (Chenopodiaceae, Gramineae, Compositae) accumulate betaine (glycine betaine) in response to salt or water stress via the pathway: choline betainal (betaine aldehyde) betaine. Betaine accumulation is probably a metabolic adaptation to stress. Intact protoplasts from leaves of spinach (Spinacia oleracea) oxidized ( UC)choline to betainal and betaine, as did protoplast lysates. Upon differential centrifugation, the ( UC)choline-oxidizing activity of lysates sedimented with chloroplasts. Chloroplasts purified from protoplast lysates by a Percoll cushion procedure retained strong ( UC)choline-oxidizing activity, although the proportion of the intermediate, ( UC)betainal, in the reaction products was usually higher than for protoplasts. Isolated chloroplasts also readily oxidized ( UC)betainal to betaine. Light increased the oxidation of both ( UC)choline and ( UC)betainal by isolated chloroplasts. Similar results were obtained with another chenopod (Beta vulgaris) but not with pea (Pisum sativum), a species that accumulates no betaine. The chloroplast site for betaine synthesis in chenopods contrasts with the mitochondrial site in mammals.

  8. Differential Contribution of Endoplasmic Reticulum and Chloroplast ω-3 Fatty Acid Desaturase Genes to the Linolenic Acid Content of Olive (Olea europaea) Fruit.

    PubMed

    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. PMID:26514651

  9. Longevity of guard cell chloroplasts in falling leaves: implication for stomatal function and cellular aging

    SciTech Connect

    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.

  10. Multiplexed Fragaria Chloroplast Genome Sequencing

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A method to sequence multiple chloroplast genomes that uses the sequencing depth of ultra high throughput sequencing technologies was recently described. Sequencing complete chloroplast genomes can resolve phylogenetic relationships at low taxonomic levels and identify point mutations and indels tha...

  11. Differential Annual Movement Patterns in a Migratory Species: Effects of Experience and Sexual Maturation

    PubMed Central

    Jorge, Paulo E.; Sowter, David; Marques, Paulo A. M.

    2011-01-01

    Some animals migrate long distances to exploit important seasonal food resources in the northern regions of the northern hemisphere, whilst avoiding winter starvation. Changes in the individual's age and navigational skills are likely to affect migration, which in turn influences the geographic distribution of individuals. Processes such as sexual maturation and navigational abilities are affected by age, and age is thus a key factor in understanding migration patterns and differences in distribution ranges. In the present study, we investigated the effects of age on the geographic distribution of a population of Lesser Black-backed Gulls Larus fuscus throughout its annual cycle, by analyzing a dataset of 19,096 records from 10,000 color-ringed gulls. In contrast to previous assumptions, the results showed that gulls were geographically segregated by age throughout the entire annual cycle, rather than showing a geographic age-related cline only in the wintering areas. This asymmetric distribution results from a reduction in the annual range of sexually mature gulls, and the differential distribution of mature and immature individuals (mature birds remained in more northern areas, compared to immature birds, throughout the annual cycle). Furthermore, although immature gulls travelled longer distances than adults, they initiated their fall migration with short movements, in contrast to adults that migrated using longer movements. The effects identified in this study explain the non-homogenous distribution of populations throughout the annual cycle, with wide implications for the development of effective human health policies and/or wildlife management strategies. PMID:21799853

  12. Studying the structure and processing of chloroplast transcripts.

    PubMed

    Barkan, Alice

    2011-01-01

    Most chloroplast genes in land plants are represented by multiple transcript isoforms that arise via differential splicing, endo- and exo-nucleolytic processing, and/or RNA editing. Exploration of the functional significance and mechanisms of these processing events is an active area of current research. This chapter focuses on methods that can be used to define the termini of chloroplast RNAs, quantify the relative levels of alternative processed RNA isoforms, and identify the binding sites of proteins that mediate chloroplast RNA processing. Various approaches for defining the sequence specificity of chloroplast RNA binding proteins are discussed, as are the parameters to consider in designing in vitro assays for RNA binding activities. A protocol is provided for a poisoned-primer extension assay for quantifying different splice isoforms. PMID:21822840

  13. Viability, ultrastructure and cytokinin metabolism of free and immobilized tobacco chloroplasts.

    PubMed

    Polanská, Lenka; Vicánková, Anna; Dobrev, Petre I; Cková, Ivana Macháv; Vanková, Radomíra

    2004-10-01

    Cytokinins play a decisive role in regulation of plastid development and differentiation, but their metabolism in plastids is not known. Metabolic studies using intact chloroplasts are prevented by their instability once they are isolated from leaf cells. Chloroplasts of Nicotiana tabacum L. cv. Petit Havana SR1 were therefore immobilized into low-viscosity alginate. Their intactness was assessed by a glyceraldehyde-3-phosphate dehydrogenase assay which indicated that free chloroplasts totally disintegrated within 7 h, while more than 50% of immobilized chloroplasts remained intact after 24 h. The immobilization had no marked impact on ultrastructure and postponed final destruction. The metabolite profile was similar in free and immobilized chloroplasts after 4 h incubation with tritiated zeatin. Nevertheless, the yield of conversion products decreased twice in immobilized chloroplasts, which was probably the outcome of mass transfer limitations and/or the sorption to polysaccharide matrix. PMID:15604795

  14. Post-transcriptional control of cell type-specific gene expression in bundle sheath and mesophyll chloroplasts of Amaranthus hypochondriacus.

    PubMed

    Boinski, J J; Wang, J L; Xu, P; Hotchkiss, T; Berry, J O

    1993-06-01

    Plants that utilize the highly efficient C4 photosynthetic pathway possess two types of specialized leaf cells, the mesophyll and bundle sheath. In mature leaves of amaranth, a dicotyledonous C4 plant, ribulose 1,5-bisphosphate carboxylase (Rubisco) is localized specifically to the chloroplasts of bundle sheath cells, and is not present in the chloroplasts of mesophyll cells. The cell type-specific expression of the chloroplast-encoded Rubisco large subunit (rbcL) gene, and other representative chloroplastic genes, was investigated by using separated bundle sheath and mesophyll chloroplasts prepared from mature amaranth leaves. One-dimensional SDS-polyacrylamide gel electrophoresis revealed several differences in the polypeptide compositions of the two chloroplast types. Western analysis demonstrated that, as in the intact leaves, the Rubisco LSU polypeptide was present only in chloroplast preparations from bundle sheath cells. Pyruvate orthophosphate dikinase (PPdK), a nuclear-encoded chloroplastic enzyme, was found only in the mesophyll chloroplast preparations. rbcL mRNA was present only in the bundle sheath chloroplast preparations, whereas transcripts for the chloroplast-encoded psbA, psaA-B, and rpl2 genes were present in both chloroplast types. Although the rbcL message accumulated only in bundle sheath chloroplasts, run-on transcription analysis indicated that the rbcL gene was transcribed in both bundle sheath and mesophyll chloroplast preparations. Therefore, differential rbcL gene expression in the isolated C4 chloroplasts is regulated, at least in part, at the post-transcriptional level. Possibly this control is mediated by differential processing or stabilization of the rbcL transcript. PMID:8329680

  15. Adaptive differentiation of traits related to resource use in a desert annual along a resource gradient.

    PubMed

    Brouillette, Larry C; Mason, Chase M; Shirk, Rebecca Y; Donovan, Lisa A

    2014-03-01

    • Plant resource-use traits are generally hypothesized to be adaptively differentiated for populations distributed along resource gradients. Although nutrient limitations are expected to select for resource-conservative strategies, water limitations may select for either resource-conservative or -acquisitive strategies. We test whether population differentiation reflects local adaptation for traits associated with resource-use strategies in a desert annual (Helianthus anomalus) distributed along a gradient of positively covarying water and nutrient availability. • We compared quantitative trait variation (Q(ST)) with neutral genetic differentiation (F(ST)), in a common garden glasshouse study, for leaf economics spectrum (LES) and related traits: photosynthesis (A(mass), A(area)), leaf nitrogen (N(mass), N(area)), leaf lifetime (LL), leaf mass per area (LMA), leaf water content (LWC), water-use efficiency (WUE, estimated as δ(13)C) and days to first flower (DFF). • Q(ST)-F(ST) differences support adaptive differentiation for Amass , N(mass), N(area), LWC and DFF. The trait combinations associated with drier and lower fertility sites represent correlated trait evolution consistent with the more resource-acquisitive end of the LES. There was no evidence for adaptive differentiation for A(area), LMA and WUE. • These results demonstrate that hot dry environments can selectively favor correlated evolution of traits contributing to a resource-acquisitive and earlier reproduction 'escape' strategy, despite lower fertility. PMID:24325125

  16. Chloroplast ribosomes and protein synthesis.

    PubMed Central

    Harris, E H; Boynton, J E; Gillham, N W

    1994-01-01

    Consistent with their postulated origin from endosymbiotic cyanobacteria, chloroplasts of plants and algae have ribosomes whose component RNAs and proteins are strikingly similar to those of eubacteria. Comparison of the secondary structures of 16S rRNAs of chloroplasts and bacteria has been particularly useful in identifying highly conserved regions likely to have essential functions. Comparative analysis of ribosomal protein sequences may likewise prove valuable in determining their roles in protein synthesis. This review is concerned primarily with the RNAs and proteins that constitute the chloroplast ribosome, the genes that encode these components, and their expression. It begins with an overview of chloroplast genome structure in land plants and algae and then presents a brief comparison of chloroplast and prokaryotic protein-synthesizing systems and a more detailed analysis of chloroplast rRNAs and ribosomal proteins. A description of the synthesis and assembly of chloroplast ribosomes follows. The review concludes with discussion of whether chloroplast protein synthesis is essential for cell survival. PMID:7854253

  17. Genetic Analysis of Chloroplast Translation

    SciTech Connect

    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.

  18. Heme content in developing chloroplasts

    SciTech Connect

    Thomas, J.; Weinstein, J.D. )

    1991-05-01

    Heme regulates tetrapyrrole biosynthesis by inhibition of {delta}-aminolevulinic acid synthesis, product inhibition of heme synthesis, and possibly other mechanisms. Determination of the physiological relevance of this inhibition requires a sensitive measurement which can distinguish regulatory free heme from heme which is an integral part of functional hemoproteins. A preliminary estimate was provided by reconstituting peroxidase activity from apo-peroxidase and the heme contained in broken plastids. However, subsequent experiments have suggested that this initial estimate was too large due to reconstitution of apo-peroxidase with heme from functional hemoproteins (i.e. heme stealing). The authors have now refined the measurement techniques to greatly reduce the extent of this heme stealing. Incubation of broken plastids with apo-peroxidase at 10C resolves the kinetics of reconstitution into two components. A fast component levels off after 100 min, and a slow component increases linearly for up to 6 hours. They believe that the heme which reconstitutes during the fast phase represents free heme, and the linear slow component represents heme stealing. In support of this theory, incubation at 15C increases the rate of both components. However, extrapolation to zero time of the slow components of the 10C and 15C time courses results in equivalent amounts of heme. Based on this kinetic differentiation between free heme and hemoprotein heme, chloroplasts isolated from cucumber cotyledons after 30 h of greening contain substantially greater amounts of free heme than etioplasts.

  19. Species versus guild level differentiation revealed across the annual cycle by isotopic niche examination.

    PubMed

    Bodey, Thomas W; Ward, Eric J; Phillips, Richard A; McGill, Rona A R; Bearhop, Stuart

    2014-03-01

    Interspecific competitive interactions typically result in niche differentiation to alleviate competition through mechanisms including character displacement. However, competition is not the sole constraint on resource partitioning, and its effects are mediated by factors including the environmental context in which species coexist. Colonial seabirds provide an excellent opportunity to investigate the importance of competition in shaping realized niche widths because their life histories lead to variation in intra- and interspecific competition across the annual cycle. Dense breeding aggregations result in intense competition for prey in surrounding waters, whereas non-breeding dispersal to larger geographical areas produces lower densities of competitors. Bayesian hierarchical models of the isotopic niche, closely aligned to the trophic niche, reveal the degree of segregation between species and functional groups during both time periods. Surprisingly, species explained far more of the variance in the isotopic niche during the non-breeding than the breeding period. Our results underline the key role of non-breeding dynamics in alleviating competition and promoting distinctions between species through the facilitation of resource partitioning. Such situations may be common in a diverse range of communities sustained by ephemeral but abundant food items. This highlights how consideration of the hierarchical grouping of competitive interactions alongside consideration of abiotic constraints across the complete annual cycle allows a full understanding of the role of competition in driving patterns of character displacement. PMID:24215391

  20. Mutation of the rice ASL2 gene encoding plastid ribosomal protein L21 causes chloroplast developmental defects and seedling death.

    PubMed

    Lin, D; Jiang, Q; Zheng, K; Chen, S; Zhou, H; Gong, X; Xu, J; Teng, S; Dong, Y

    2015-05-01

    The plastid ribosome proteins (PRPs) play important roles in plastid protein biosynthesis, chloroplast differentiation and early chloroplast development. However, the specialised functions of individual protein components of the chloroplast ribosome in rice (Oryza sativa) remain unresolved. In this paper, we identified a novel rice PRP mutant named asl2 (Albino seedling lethality 2) exhibiting an albino, seedling death phenotype. In asl2 mutants, the alteration of leaf colour was associated with chlorophyll (Chl) content and abnormal chloroplast development. Through map-based cloning and complementation, the mutated ASL2 gene was isolated and found to encode the chloroplast 50S ribosome protein L21 (RPL21c), a component of the chloroplast ribosome large subunit, which was localised in chloroplasts. ASL2 was expressed at a higher level in the plumule and leaves, implying its tissue-specific expression. Additionally, the expression of ASL2 was regulated by light. The transcript levels of the majority of genes for Chl biosynthesis, photosynthesis and chloroplast development were strongly affected in asl2 mutants. Collectively, the absence of functional ASL2 caused chloroplast developmental defects and seedling death. This report establishes the important role of RPL21c in chloroplast development in rice. PMID:25280352

  1. Evolution of Chloroplast J Proteins

    PubMed Central

    Chiu, Chi-Chou; Chen, Lih-Jen; Su, Pai-Hsiang; Li, Hsou-min

    2013-01-01

    Hsp70 chaperones are involved in multiple biological processes and are recruited to specific processes by designated J domain-containing cochaperones, or J proteins. To understand the evolution and functions of chloroplast Hsp70s and J proteins, we identified the Arabidopsis chloroplast J protein constituency using a combination of genomic and proteomic database searches and individual protein import assays. We show that Arabidopsis chloroplasts have at least 19 J proteins, the highest number of confirmed J proteins for any organelle. These 19 J proteins are classified into 11 clades, for which cyanobacteria and glaucophytes only have homologs for one clade, green algae have an additional three clades, and all the other 7 clades are specific to land plants. Each clade also possesses a clade-specific novel motif that is likely used to interact with different client proteins. Gene expression analyses indicate that most land plant-specific J proteins show highly variable expression in different tissues and are down regulated by low temperatures. These results show that duplication of chloroplast Hsp70 in land plants is accompanied by more than doubling of the number of its J protein cochaperones through adding new J proteins with novel motifs, not through duplications within existing families. These new J proteins likely recruit chloroplast Hsp70 to perform tissue specific functions related to biosynthesis rather than to stress resistance. PMID:23894646

  2. Chloroplast evolution, structure and functions

    PubMed Central

    Jensen, Poul Erik

    2014-01-01

    In this review, we consider a selection of recent advances in chloroplast biology. These include new findings concerning chloroplast evolution, such as the identification of Chlamydiae as a third partner in primary endosymbiosis, a second instance of primary endosymbiosis represented by the chromatophores found in amoebae of the genus Paulinella, and a new explanation for the longevity of captured chloroplasts (kleptoplasts) in sacoglossan sea slugs. The controversy surrounding the three-dimensional structure of grana, its recent resolution by tomographic analyses, and the role of the CURVATURE THYLAKOID1 (CURT1) proteins in supporting grana formation are also discussed. We also present an updated inventory of photosynthetic proteins and the factors involved in the assembly of thylakoid multiprotein complexes, and evaluate findings that reveal that cyclic electron flow involves NADPH dehydrogenase (NDH)- and PGRL1/PGR5-dependent pathways, both of which receive electrons from ferredoxin. Other topics covered in this review include new protein components of nucleoids, an updated inventory of the chloroplast proteome, new enzymes in chlorophyll biosynthesis and new candidate messengers in retrograde signaling. Finally, we discuss the first successful synthetic biology approaches that resulted in chloroplasts in which electrons from the photosynthetic light reactions are fed to enzymes derived from secondary metabolism. PMID:24991417

  3. Biosynthesis of starch in chloroplasts.

    PubMed

    Nomura, T; Nakayama, N; Murata, T; Akazawa, T

    1967-03-01

    The enzymic synthesis of ADP-glucose and UDP-glucose by chloroplastic pyrophosphorylase of bean and rice leaves has been demonstrated by paper chromatographic techniques. In both tissues, the activity of UDP-glucose-pyrophosphorylase was much higher than ADP-glucose-pyrophosphorylase. Glycerate-3-phosphate, phosphoenolpyruvate and fructose-1,6-diphosphate did not stimulate ADP-glucose formation by a pyrophosphorylation reaction. The major metabolic pathway for UDP-glucose utilization appears to be the synthesis of either sucrose or sucrose-P. On the other hand, a specific precursor role of ADP-glucose for synthesizing chloroplast starch by the ADP-glucose-starch transglucosylase reaction is supported by the coupled enzyme system of ADP-glucose-pyrophosphorylase and transglucosylase, isolated from chloroplasts. None of the glycolytic intermediates stimulated the glucose transfer in the enzyme sequence of reaction system employed. PMID:4292567

  4. Isolation of chloroplastic phosphoglycerate kinase

    SciTech Connect

    Macioszek, J.; Anderson, L.E. ); Anderson, J.B. )

    1990-09-01

    We report here a method for the isolation of high specific activity phosphoglycerate kinase (EC 2.7.2.3) from chloroplasts. The enzyme has been purified over 200-fold from pea (Pisum sativum L.) stromal extracts to apparent homogeneity with 23% recovery. Negative cooperativity is observed with the two enzyme phosphoglycerate kinase/glyceraldehyde-3-P dehydrogenase (EC 1.2.1.13) couple restored from the purified enzymes when NADPH is the reducing pyridine nucleotide, consistent with earlier results obtained with crude chloroplastic extracts. Michaelis Menten kinetics are observed when 3-phosphoglycerate is held constant and phosphoglycerate kinase is varied, which suggests that phosphoglycerate kinase-bound 1,3-bisphosphoglycerate may be the preferred substrate for glyceraldehyde-3-P dehydrogenase in the chloroplast.

  5. Purity of Chloroplasts Prepared from the Siphonous Green Alga, Caulerpa simpliciuscula, as Determined by Their Ultrastructure and Their Enzymic Content 1

    PubMed Central

    Grant, Bruce R.; Wright, Simon W.

    1980-01-01

    The ultrastructure and enzyme distribution in chloroplasts and other subcellular fractions isolated from the siphonous green alga, Caulerpa simpliciuscula, are described. The isolated chloroplasts were similar in appearance to those in the tissue from which they were derived, and in typical preparations 70% or more were intact. Chloroplasts which had lost their outer envelopes could be separated from intact plastids by centrifugation at low speeds through gradients of colloidal silica. Intact chloroplasts separated in this way retained their photosynthetic capacity and were impermeable to ferricyanide ions. The chloroplast preparations separated by differential centrifugation and refractionated using either discontinuous or continuous Percoll gradients contained non-chloroplast material. It was estimated that this amounted to a maximum of 10% of the mitochondrial population and 6% of cytoplasm extracted from the plant. The contaminating material surrounded the chloroplasts in a thin layer and was surrounded by a membrane. Images PMID:16661374

  6. Thermal Damage to Chloroplast Envelope Membranes 1

    PubMed Central

    McCain, Douglas C.; Croxdale, Judith; Markley, John L.

    1989-01-01

    Nuclear magnetic resonance was used to detect thermal injury to chloroplasts in vivo. A lesion occurs in the chloroplast envelope membrane at temperatures between 53°C and 57°C, depending on species, leaf condition, and heating rate. The injury is associated with a sudden loss of water from the chloroplast. PMID:16666815

  7. Photomorphogenic Regulation of Chloroplast Replication in Euglena

    PubMed Central

    Srinivas, Usha; Lyman, Harvard

    1980-01-01

    Chloroplast replication in Euglena gracilis is specifically inhibited by ultraviolet light and the effect is photoreactivable. The ability of irradiated cells to be photoreactivated is lost more rapidly if cells are incubated in red light than in darkness. A mutant, Y9ZNa1L, which lacks the red-blue photomorphogenic system regulating chloroplast synthesis does not show the red-light-enhanced loss of photoreactivability. Another mutant, Y11P27ZD which has the red-blue system, but lacks the blue-light system also regulating chloroplast synthesis, shows the red-light effect. The red-light effect is seen in a mutant of photosynthetic electron transport, P4ZUL, which rules out a product of photosynthesis as a mediator of the effect. Inhibitors of protein synthesis on chloroplast ribosomes do not prevent the red-light-enhanced loss of chloroplast DNA. Chloroplast DNA is lost rapidly when UV-irradiated cells are incubated in red light, showing that the loss of photoreactivability is due to the loss of the substrate for photoreactivation, chloroplast DNA. Therefore, the red-blue photomorphogenic system is activating a chloroplast DNA-specific nuclease(s). A model is proposed for a light-mediated mechanism regulating the amount of chloroplast DNA: blue light would promote chloroplast DNA synthesis; red light would promote its degradation. The photomorphogenic systems regulating chloroplast synthesis might work by activating a chloroplast-specific modification-restriction mechanism. PMID:16661425

  8. Protein import into chloroplasts requires a chloroplast ATPase

    SciTech Connect

    Pain, D.; Blobel, G.

    1987-05-01

    The authors have transcribed mRNA from a cDNA clone coding for pea ribulose-1,5-bisphosphate carboxylase, translated the mRNA in a wheat germ cell-free system, and studied the energy requirement for posttranslational import of the (/sup 35/S)methionine-labeled protein into the stroma of pea chloroplasts. They found that import depends on ATP hydrolysis within the stroma. Import is not inhibited when H/sup +/, K/sup +/, Na/sup +/, or divalent cation gradients across the chloroplast membranes are dissipated by ionophores, as long as exogenously added ATP is also present during the import reaction. The data suggest that protein import into the chloroplast stroma requires a chloroplast ATPase that does not function to generate a membrane potential for driving the import reaction but that exerts its effect in another, yet-to-be-determined, mode. They have carried out a preliminary characterization of this ATPase regarding its nucleotide specificity and the effects of various ATPase inhibitors.

  9. Mutational dynamics of aroid chloroplast genomes.

    PubMed

    Ahmed, Ibrar; Biggs, Patrick J; Matthews, Peter J; Collins, Lesley J; Hendy, Michael D; Lockhart, Peter J

    2012-01-01

    A characteristic feature of eukaryote and prokaryote genomes is the co-occurrence of nucleotide substitution and insertion/deletion (indel) mutations. Although similar observations have also been made for chloroplast DNA, genome-wide associations have not been reported. We determined the chloroplast genome sequences for two morphotypes of taro (Colocasia esculenta; family Araceae) and compared these with four publicly available aroid chloroplast genomes. Here, we report the extent of genome-wide association between direct and inverted repeats, indels, and substitutions in these aroid chloroplast genomes. We suggest that alternative but not mutually exclusive hypotheses explain the mutational dynamics of chloroplast genome evolution. PMID:23204304

  10. Chloroplast division in higher plants requires members of two functionally divergent gene families with homology to bacterial ftsZ.

    PubMed Central

    Osteryoung, K W; Stokes, K D; Rutherford, S M; Percival, A L; Lee, W Y

    1998-01-01

    The division of plastids is critical for viability in photosynthetic eukaryotes, but the mechanisms associated with this process are still poorly understood. We previously identified a nuclear gene from Arabidopsis encoding a chloroplast-localized homolog of the bacterial cell division protein FtsZ, an essential cytoskeletal component of the prokaryotic cell division apparatus. Here, we report the identification of a second nuclear-encoded FtsZ-type protein from Arabidopsis that does not contain a chloroplast targeting sequence or other obvious sorting signals and is not imported into isolated chloroplasts, which strongly suggests that it is localized in the cytosol. We further demonstrate using antisense technology that inhibiting expression of either Arabidopsis FtsZ gene (AtFtsZ1-1 or AtFtsZ2-1) in transgenic plants reduces the number of chloroplasts in mature leaf cells from 100 to one, indicating that both genes are essential for division of higher plant chloroplasts but that each plays a distinct role in the process. Analysis of currently available plant FtsZ sequences further suggests that two functionally divergent FtsZ gene families encoding differentially localized products participate in chloroplast division. Our results provide evidence that both chloroplastic and cytosolic forms of FtsZ are involved in chloroplast division in higher plants and imply that important differences exist between chloroplasts and prokaryotes with regard to the roles played by FtsZ proteins in the division process. PMID:9836740

  11. Comparative chromatography of chloroplast pigment

    NASA Technical Reports Server (NTRS)

    Grandolfo, M.; Sherma, J.; Strain, H. H.

    1969-01-01

    Methods for isolation of low concentration pigments of the cocklebur species are described. The methods entail two step chromatography so that the different sorption properties of the various pigments in varying column parameters can be utilized. Columnar and thin layer methods are compared. Many conditions influence separability of the chloroplasts.

  12. Two distinct redox cascades cooperatively regulate chloroplast functions and sustain plant viability.

    PubMed

    Yoshida, Keisuke; Hisabori, Toru

    2016-07-01

    The thiol-based redox regulation system is believed to adjust chloroplast functions in response to changes in light environments. A redox cascade via the ferredoxin-thioredoxin reductase (FTR)/thioredoxin (Trx) pathway has been traditionally considered to serve as a transmitter of light signals to target enzymes. However, emerging data indicate that chloroplasts have a complex redox network composed of diverse redox-mediator proteins and target enzymes. Despite extensive research addressing this system, two fundamental questions are still unresolved: How are redox pathways orchestrated within chloroplasts, and why are chloroplasts endowed with a complicated redox network? In this report, we show that NADPH-Trx reductase C (NTRC) is a key redox-mediator protein responsible for regulatory functions distinct from those of the classically known FTR/Trx system. Target screening and subsequent biochemical assays indicated that NTRC and the Trx family differentially recognize their target proteins. In addition, we found that NTRC is an electron donor to Trx-z, which is a key regulator of gene expression in chloroplasts. We further demonstrate that cooperative control of chloroplast functions via the FTR/Trx and NTRC pathways is essential for plant viability. Arabidopsis double mutants impaired in FTR and NTRC expression displayed lethal phenotypes under autotrophic growth conditions. This severe growth phenotype was related to a drastic loss of photosynthetic performance. These combined results provide an expanded map of the chloroplast redox network and its biological functions. PMID:27335455

  13. Covariations in the nuclear chloroplast transcriptome reveal a regulatory master-switch

    PubMed Central

    Richly, Erik; Dietzmann, Angela; Biehl, Alexander; Kurth, Joachim; Laloi, Christophe; Apel, Klaus; Salamini, Francesco; Leister, Dario

    2003-01-01

    The evolution of the endosymbiotic progenitor into the chloroplast organelle was associated with the transfer of numerous chloroplast genes into the nucleus. Hence, inter-organellar signalling, and the co-ordinated expression of sets of nuclear genes, was set up to control the metabolic and developmental status of the chloroplast. Here, we show by the differential-expression analysis of 3,292 genes, that most of the 35 environmental and genetic conditions tested, including plastid signalling mutations, elicit only three main classes of response from the nuclear chloroplast transcriptome. Two classes, probably involving GUN (genomes uncoupled)-type plastid signalling, are characterized by alterations, in opposite directions, in the expression of largely overlapping sets of genes. PMID:12776738

  14. Stable Expression of Basic Fibroblast Growth Factor in Chloroplasts of Tobacco.

    PubMed

    Wang, Yun-Peng; Wei, Zheng-Yi; Zhong, Xiao-Fang; Lin, Chun-Jing; Cai, Yu-Hong; Ma, Jian; Zhang, Yu-Ying; Liu, Yan-Zhi; Xing, Shao-Chen

    2016-01-01

    Basic fibroblast growth factor (bFGF) is a multifunctional factor in acceleration of cell proliferation, differentiation and transference, and therefore widely used in clinical applications. In this study, expression vector pWX-Nt03 harboring a codon-optimized bFGF gene was constructed and introduced into the tobacco chloroplasts by particle bombardment. After four rounds of selection, bFGF was proved to integrate into the chloroplast genome of regenerated plants and two of four transgenic plants were confirmed to be homoplastomic by PCR and Southern hybridization. ELISA assay indicated that bFGF represented approximately 0.1% of total soluble protein in the leaves of transplastomic tobacco plants. This is the first report of bFGF expression via chloroplast transformation in model plant, providing an additional option for the production of chloroplast-produced therapeutic proteins. PMID:26703590

  15. Stable Expression of Basic Fibroblast Growth Factor in Chloroplasts of Tobacco

    PubMed Central

    Wang, Yun-Peng; Wei, Zheng-Yi; Zhong, Xiao-Fang; Lin, Chun-Jing; Cai, Yu-Hong; Ma, Jian; Zhang, Yu-Ying; Liu, Yan-Zhi; Xing, Shao-Chen

    2015-01-01

    Basic fibroblast growth factor (bFGF) is a multifunctional factor in acceleration of cell proliferation, differentiation and transference, and therefore widely used in clinical applications. In this study, expression vector pWX-Nt03 harboring a codon-optimized bFGF gene was constructed and introduced into the tobacco chloroplasts by particle bombardment. After four rounds of selection, bFGF was proved to integrate into the chloroplast genome of regenerated plants and two of four transgenic plants were confirmed to be homoplastomic by PCR and Southern hybridization. ELISA assay indicated that bFGF represented approximately 0.1% of total soluble protein in the leaves of transplastomic tobacco plants. This is the first report of bFGF expression via chloroplast transformation in model plant, providing an additional option for the production of chloroplast-produced therapeutic proteins. PMID:26703590

  16. Formation of putative chloroplast cytochromes in isolated developing pea chloroplasts

    SciTech Connect

    Thaver, S.S.; Bhava, D.; Castelfranco, P.A.

    1986-04-01

    In addition to chlorophyll-protein complexes, other proteins were labeled when isolated developing pea chloroplasts were incubated with (/sup 14/C)-5-aminolevulinic acid (/sup 14/C)-ALA. The major labeled band (M/sub r/ = 43 kDa by LDS-PAGE) was labeled even in the presence of chloramphenicol. Heme-dependent peroxidase activity (as detected by the tetramethyl benzidine-H/sub 2/O/sub 2/ stain) was not visibly associated with this band. The radioactive band was stable to heat, 5% HCl in acetone, and was absent if the incubation with (/sup 14/C)-5-aminolevulinic acid was carried out in the presence of N-methyl protoporphyrin IX dimethyl ester (a specific inhibitor of ferrochelatase). Organic solvent extraction procedures for the enrichment of cytochrome f from chloroplast membranes also extracted this unknown labeled product. It was concluded that this labeled product was probably a c-type cytochrome. The effect of exogenous iron, iron chelators, gabaculine (an inhibitor of ALA synthesis) and other incubation conditions upon the in vitro formation of putative chloroplast cytochromes will be discussed.

  17. GROWTH REGULATING FACTOR5 Stimulates Arabidopsis Chloroplast Division, Photosynthesis, and Leaf Longevity1[OPEN

    PubMed Central

    Vercruyssen, Liesbeth; Tognetti, Vanesa B.; Gonzalez, Nathalie; Van Dingenen, Judith; De Milde, Liesbeth; Bielach, Agnieszka; De Rycke, Riet; Van Breusegem, Frank; Inzé, Dirk

    2015-01-01

    Arabidopsis (Arabidopsis thaliana) leaf development relies on subsequent phases of cell proliferation and cell expansion. During the proliferation phase, chloroplasts need to divide extensively, and during the transition from cell proliferation to expansion, they differentiate into photosynthetically active chloroplasts, providing the plant with energy. The transcription factor GROWTH REGULATING FACTOR5 (GRF5) promotes the duration of the cell proliferation period during leaf development. Here, it is shown that GRF5 also stimulates chloroplast division, resulting in a higher chloroplast number per cell with a concomitant increase in chlorophyll levels in 35S:GRF5 leaves, which can sustain higher rates of photosynthesis. Moreover, 35S:GRF5 plants show delayed leaf senescence and are more tolerant for growth on nitrogen-depleted medium. Cytokinins also stimulate leaf growth in part by extending the cell proliferation phase, simultaneously delaying the onset of the cell expansion phase. In addition, cytokinins are known to be involved in chloroplast development, nitrogen signaling, and senescence. Evidence is provided that GRF5 and cytokinins synergistically enhance cell division and chlorophyll retention after dark-induced senescence, which suggests that they also cooperate to stimulate chloroplast division and nitrogen assimilation. Taken together with the increased leaf size, ectopic expression of GRF5 has great potential to improve plant productivity. PMID:25604530

  18. In vivo effects of NbSiR silencing on chloroplast development in Nicotiana benthamiana.

    PubMed

    Kang, Yong-Won; Lee, Jae-Yong; Jeon, Young; Cheong, Gang-Won; Kim, Moonil; Pai, Hyun-Sook

    2010-04-01

    Sulfite reductase (SiR) performs dual functions, acting as a sulfur assimilation enzyme and as a chloroplast (cp-) nucleoid binding protein. In this study, we examined the in vivo effects of SiR deficiency on chloroplast development in Nicotiana benthamiana. Virus-induced gene silencing of NbSiR resulted in leaf yellowing and growth retardation phenotypes, which were not rescued by cysteine supplementation. NbSiR:GFP fusion protein was targeted to chloroplasts and colocalized with cp-nucleoids. Recombinant full-length NbSiR protein and the C-terminal half of NbSiR possessed cp-DNA compaction activities in vitro, and expression of full-length NbSiR in E. coli caused condensation of genomic DNA. NbSiR silencing differentially affected expression of plastid-encoded genes, inhibiting expression of several genes more severely than others. In the later stages, depletion of NbSiR resulted in chloroplast ablation. In NbSiR-silenced plants, enlarged cp-nucleoids containing an increased amount of cp-DNA were observed in the middle of the abnormal chloroplasts, and the cp-DNAs were predominantly of subgenomic sizes based on pulse field gel electrophoresis. The abnormal chloroplasts developed prolamellar body-like cubic lipid structures in the light without accumulating NADPH:protochlorophyllide oxidoreductase proteins. Our results suggest that NbSiR plays a role in cp-nucleoid metabolism, plastid gene expression, and thylakoid membrane development. PMID:20047069

  19. Fluorescence Properties of Guard Cell Chloroplasts

    PubMed Central

    Zeiger, Eduardo; Armond, Paul; Melis, Anastasios

    1981-01-01

    The presence of chloroplasts in guard cells from leaf epidermis, coleoptile, flowers, and albino portions of variegated leaves was established by incident fluorescence microscopy, thus confirming the notion that guard cell chloroplasts are remarkably conserved. Room temperature emission spectra from a few chloroplasts in a single guard cell of Vicia faba showed one major peak at around 683 nanometers. Low-temperature (77 K) emission spectra from peels of albino portions of Chlorophytum comosum leaves and from mesophyll chloroplasts of green parts of the same leaves showed major peaks at around 687 and 733 nanometers, peaks usually attributed to photosystem II and photosystem I pigment systems, respectively. Spectra of peels of V. faba leaves showed similar peaks. However, fluorescence microscopy revealed that the Vicia peels, as well as those from Allium cepa and Tulipa sp., were contaminated with non-guard cell chloroplasts which were practically undetectable under bright field illumination. These observations pose restrictions on the use of epidermal peels as a source of isolated guard cell chloroplasts. Studies on the 3-(3,4-dichlorophenyl)-1,1-dimethylurea-sensitive variable fluorescence kinetics of uncontaminated epidermal peels of C. comosum indicated that guard cell chloroplasts operate a normal, photosystem II-dependent, linear electron transport. The above properties in combination with their reported inability to fix CO2 photosynthetically may render the guard cell chloroplasts optimally suited to supply the reducing and high-energy phosphate equivalents needed to sustain active ion transport during stomatal opening in daylight. PMID:16661620

  20. Insights into chloroplast biogenesis and development.

    PubMed

    Pogson, Barry J; Ganguly, Diep; Albrecht-Borth, Verónica

    2015-09-01

    In recent years many advances have been made to obtain insight into chloroplast biogenesis and development. In plants several plastids types exist such as the proplastid (which is the progenitor of all plastids), leucoplasts (group of colourless plastids important for storage including elaioplasts (lipids), amyloplasts (starch) or proteinoplasts (proteins)), chromoplasts (yellow to orange-coloured due to carotenoids, in flowers or in old leaves as gerontoplasts), and the green chloroplasts. Chloroplasts are indispensable for plant development; not only by performing photosynthesis and thus rendering the plant photoautotrophic, but also for biochemical processes (which in some instances can also take place in other plastids types), such as the synthesis of pigments, lipids, and plant hormones and sensing environmental stimuli. Although we understand many aspects of these processes there are gaps in our understanding of the establishment of functional chloroplasts and their regulation. Why is that so? Even though chloroplast function is comparable in all plants and most of the algae, ferns and moss, detailed analyses have revealed many differences, specifically with respect to its biogenesis. As an update to our prior review on the genetic analysis of chloroplast biogenesis and development [1] herein we will focus on recent advances in Angiosperms (monocotyledonous and dicotyledonous plants) that provide novel insights and highlight the challenges and prospects for unravelling the regulation of chloroplast biogenesis specifically during the establishment of the young plants. This article is part of a Special Issue entitled: Chloroplast Biogenesis. PMID:25667967

  1. Chloroplasts as functional organelles in animal tissues.

    PubMed

    Trench, R K; Greene, R W; Bystrom, B G

    1969-08-01

    The marine gastropod molluscs Tridachia crispata, Tridachiella diomedea, and Placobranchus ianthobapsus (Sacoglossa, Opisthobranchia) possess free functional chloroplasts within the cells of the digestive diverticula, as determined by observations on ultrastructure, pigment analyses, and experiments on photosynthetic capacity. In the light, the chloroplasts incorporate H(14)CO(3) (-)in situ. Reduced radiocarbon is translocated to various chloroplast-free tissues in the animals. The slugs feed on siphonaceous algae from which the chloroplasts are derived. Pigments from the slugs and from known siphonaceous algae, when separated chromatographically and compared, showed similar components. Absorption spectra of extracts of slugs and algae were very similar. The larvae of the slugs are pigment-free up to the post-veliger stage, suggesting that chloroplasts are acquired de novo. with each new generation. PMID:5792329

  2. Interaction of Chloroplasts with Inhibitors

    PubMed Central

    Ridley, Stuart M.

    1983-01-01

    Several effects on pea (Pisum sativum L. var Onwards) chloroplasts of a new diphenylether herbicide, fomesafen (5-[2-chloro-4-trifluoromethyl-phenoxy]-N-methanesulfonyl-2 -nitrobenzamide) have been compared with those of a herbicide of related structure, nitrofluorfen (2-chloro-1-[4-nitrophenoxy]-4-[trifluoromethyl]benzene). Although both compounds produce the same light-dependent symptoms of desiccation and chlorosis indicative of a common primary mechanism of action, this study is concerned with a more broadly based investigation of different effects on the electron transport system. Comparisons have also been made with other compounds interacting with the chloroplast. Unlike nitrofluorfen, fomesafen has little effect as an inhibitor of electron flow or energy transfer. Both compounds have the ability to stimulate superoxide production through a functional electron transport system, and this involves specifically the p-nitro substituent. The stimulation, which is not likely to be an essential part of the primary herbicidal effect, is diminished under conditions that remove the coupling factor. Evidence suggests that both diphenylethers may be able to bind to the coupling factor, and kinetic studies reveal this for dibromothymoquinone as well. Such a binding site might be an important feature in allowing the primary effect of the diphenylether herbicides to be expressed. PMID:16663025

  3. Plastidic Isoprenoid Synthesis during Chloroplast Development 1

    PubMed Central

    Heintze, Adolf; Görlach, Jörn; Leuschner, Carola; Hoppe, Petra; Hagelstein, Petra; Schulze-Siebert, Detlef; Schultz, Gernot

    1990-01-01

    The chloroplast isoprenoid synthesis of very young leaves is supplied by the plastidic CO2 → pyruvate → acetyl-coenzyme A (C3 → C2) metabolism (D Schulze-Siebert, G Schultz [1987] Plant Physiol 84: 1233-1237) and occurs via the plastidic mevalonate pathway. The plastidic C3 → C2 metabolism and/or plastidic mevalonate pathway of barley (Hordeum vulgare L.) seedlings changes from maximal activity at the leaf base (containing developing chloroplasts with incomplete thylakoid stacking but a considerable rate of photosynthetic CO2-fixation) almost to ineffectivity at the leaf tip (containing mature chloroplasts with maximal photosynthetic activity). The ability to import isopentenyl diphosphate from the extraplastidic space gradually increases to substitute for the loss of endogenous intermediate supply for chloroplast isoprenoid synthesis (change from autonomic to division-of-labor stage). Fatty acid synthesis from NaH14CO3 decreases in the same manner as shown for leaf sections and chloroplasts isolated from these. Evidence has been obtained for a drastic decrease of pyruvate decarboxylase-dehydrogenase activity during chloroplast development compared with other anabolic chloroplast pathways (synthesis of aromatic amino acid and branched chain amino acids). The noncompetition of pyruvate and acetate in isotopic dilution studies indicates that both a pyruvate-derived and an acetate-derived compound are simultaneously needed to form introductory intermediates of the mevalonate pathway, presumably acetoacetyl-coenzyme A. PMID:16667567

  4. 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

    PubMed Central

    Manandhar-Shrestha, K.; Tamot, B.; Pratt, E. P. S.; Saitie, S.; Bräutigam, A.; Weber, A. P. M.; Hoffmann-Benning, Susanne

    2013-01-01

    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 particularly 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. Seventy-four percent have a known or predicted membrane association. Twenty-one membrane proteins were 2–15 times more abundant in BS cells, while 36 of the proteins were more abundant in M chloroplast 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 13 candidate genes. Chloroplast association for seven proteins was confirmed using YFP/GFP labeling. Gene expression of four putative transporters was examined throughout the leaf and during the greening of leaves. 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

  5. The complexity of chloroplast chaperonins.

    PubMed

    Vitlin Gruber, Anna; Nisemblat, Shahar; Azem, Abdussalam; Weiss, Celeste

    2013-12-01

    Type I chaperonins are large oligomeric protein ensembles that are involved in the folding and assembly of other proteins. Chloroplast chaperonins and co-chaperonins exist in multiple copies of two distinct isoforms that can combine to form a range of labile oligomeric structures. This complex system increases the potential number of chaperonin substrates and possibilities for regulation. The incorporation of unique subunits into the oligomer can modify substrate specificity. Some subunits are upregulated in response to heat shock and some show organ-specific expression, whereas others possess additional functions that are unrelated to their role in protein folding. Accumulating evidence suggests that specific subunits have distinct roles in biogenesis of ribulose-1,5-bisphosphate carboxylase oxygenase (Rubisco). PMID:24035661

  6. Analysis of the differential response of five annuals to elevated CO sub 2 during growth

    SciTech Connect

    Garbutt, K. ); Williams, W.E. ); Bazzaz, F.A. )

    1990-06-01

    In order to investigate the effects, without competition, of CO{sub 2} on germination, growth, physiological response, and reproduction, the authors focussed on co-occurring species that are prominent members of an annual community in Illinois. Five species of old field annual plants - Abutilon theophrasti (C{sub 3}), Amaranthus retroflexus (C{sub 4}), Ambrosia artemisiifolia (C{sub 3}), Chenopodium album (C{sub 3}), and Setaria faberii (C{sub 4}) - were grown for their entire life cycle as individuals at CO{sub 2} concentration of 350 {mu}L/O, 500 {mu}L/L, and 700 {mu}L/L. Emergence time, growth rate, shoot water status, photosynthesis, conductance, flowering time, nitrogen content, and biomass and reproductive biomass were measured. There was no detectable effect of enhanced CO{sub 2} on timing of emergency in any of the species. The three levels of carbon dioxide concentration were shown to produce varying effects on remaining quantities measured in the five different plants. Some of these differences were not statistically significant. The response of most characters had a significant species {times} CO{sub 2} interaction. However, this was not simply caused by the C{sub 3}/C{sub 4} dichotomy. Reproductive biomass (seed, fruits, and flowers) increased with increasing CO{sub 2} in Amaranthus (C{sub 4}) and in Chenopodium and Ambrosia (both C{sub 3}), but there was no change in Setaria (C{sub 4}), and Abutilon (C{sub 3}) showed a peak at 500 {mu}L/L. Species of the same community differed in their response to CO{sub 2}, and these differences may help explain the outcome of competitive interactions among these species above ambient CO{sub 2} levels.

  7. An Arabidopsis soluble chloroplast proteomic analysis reveals the participation of the Executer pathway in response to increased light conditions

    PubMed Central

    Uberegui, Estefanía; Hall, Michael; Lorenzo, Óscar; Schröder, Wolfgang P.; Balsera, Mónica

    2015-01-01

    The Executer1 and Executer2 proteins have a fundamental role in the signalling pathway mediated by singlet oxygen in chloroplast; nonetheless, not much is known yet about their specific activity and features. Herein, we have followed a differential-expression proteomics approach to analyse the impact of Executer on the soluble chloroplast protein abundance in Arabidopsis. Because singlet oxygen plays a significant role in signalling the oxidative response of plants to light, our analysis also included the soluble chloroplast proteome of plants exposed to a moderate light intensity in the time frame of hours. A number of light- and genotype-responsive proteins were detected, and mass-spectrometry identification showed changes in abundance of several photosynthesis- and carbon metabolism-related proteins as well as proteins involved in plastid mRNA processing. Our results support the participation of the Executer proteins in signalling and control of chloroplast metabolism, and in the regulation of plant response to environmental changes. PMID:25740923

  8. Chloroplasts at work during plant innate immunity.

    PubMed

    Serrano, Irene; Audran, Corinne; Rivas, Susana

    2016-06-01

    The major role played by chloroplasts during light harvesting, energy production, redox homeostasis, and retrograde signalling processes has been extensively characterized. Beyond the obvious link between chloroplast functions in primary metabolism and as providers of photosynthesis-derived carbon sources and energy, a growing body of evidence supports a central role for chloroplasts as integrators of environmental signals and, more particularly, as key defence organelles. Here, we review the importance of these organelles as primary sites for the biosynthesis and transmission of pro-defence signals during plant immune responses. In addition, we highlight interorganellar communication as a crucial process for amplification of the immune response. Finally, molecular strategies used by microbes to manipulate, directly or indirectly, the production/function of defence-related signalling molecules and subvert chloroplast-based defences are also discussed. PMID:26994477

  9. Solar energy conversion by chloroplast photoelectrochemical cells

    NASA Astrophysics Data System (ADS)

    Bhardwaj, R.; Pan, R. L.; Gross, E. L.

    1981-01-01

    A photoelectrochemical cell based on chloroplasts which generates large photovoltages and photocurrents from solar energy is presented. The cell contains broken Type C chloroplasts placed on a filter separating compartments containing an electron acceptor and electron donor with platinum electrodes in each. Photovoltages were observed across a load resistance of 3000 ohms with either flavin mononucleotide or anthroquinone 2-sulphonate as the electron acceptor and dichlorophenol indophenol as the donor, and persisted for 1-2 hr after the light was turned off. The powers and short circuit currents obtained in the chloroplast cells are nearly equal to those obtained in cells based on isolated photosystem I particles. Finally, an efficiency of 2.3% has been measured for the chloroplast contribution to the total power in flavin mononucleotide cells.

  10. Differentiation of the seed coat and composition of the mucilage of Lepidium perfoliatum L.: a desert annual with typical myxospermy.

    PubMed

    Huang, Daihong; Wang, Cui; Yuan, Junwen; Cao, Jing; Lan, Haiyan

    2015-10-01

    Myxospermy is an important feature in seeds of many plant species grown in desert region. Fertilization can initiate differentiation of the seed coat epidermis into a specialized cell type with mucilage production. In the present study, comprehensive analyses were performed on the seed coat differentiation, mucilage production and composition, and seed germination in Lepidium perfoliatum (Brassicaceae), a desert annual with typical myxospermy in China. First, results indicated that mucilage was secreted uniformly at the outer tangential wall, resulting in compression of the cytoplasm to the bottom of the epidermal cells. Secondly, the inner tangential wall and two radial walls of the subepidermal cells were apparently thickened by production of a secondary cell wall material, which resulted in a 'typical' palisade appearance. Thirdly, immunohistochemical staining combined with the enzymatic digestion and infrared spectrum analysis of the mucilage indicated that, while one important component of the seed coat mucilage in L. perfoliatum was pectin, it also contained β-1,3-d-glucan and xyloglucan. Finally, seed germination showed that seeds with mucilage displayed significantly higher germination percentage than that of demucilaged seeds in abundant or excess water conditions. These results suggest that the possible ecological role of mucilage in L. perfoliatum is in the adaptation to habitats with well-watered and water-logged conditions, rather than water stress. PMID:26341978

  11. Proteomic comparison reveals the contribution of chloroplast to salt tolerance of a wheat introgression line.

    PubMed

    Xu, Wenjing; Lv, Hongjun; Zhao, Mingming; Li, Yongchao; Qi, Yueying; Peng, Zhenying; Xia, Guangmin; Wang, Mengcheng

    2016-01-01

    We previously bred a salt tolerant wheat cv. SR3 with bread wheat cv. JN177 as the parent via asymmetric somatic hybridization, and found that the tolerance is partially attributed to the superior photosynthesis capacity. Here, we compared the proteomes of two cultivars to unravel the basis of superior photosynthesis capacity. In the maps of two dimensional difference gel electrophoresis (2D-DIGE), there were 26 differentially expressed proteins (DEPs), including 18 cultivar-based and 8 stress-responsive ones. 21 of 26 DEPs were identified and classified into four categories, including photosynthesis, photosynthesis system stability, linolenic acid metabolism, and protein synthesis in chloroplast. The chloroplast localization of some DEPs confirmed that the identified DEPs function in the chloroplast. The overexpression of a DEP enhanced salt tolerance in Arabidopsis thaliana. In line with these data, it is concluded that the contribution of chloroplast to high salinity tolerance of wheat cv. SR3 appears to include higher photosynthesis efficiency by promoting system protection and ROS clearance, stronger production of phytohormone JA by enhancing metabolism activity, and modulating the in chloroplast synthesis of proteins. PMID:27562633

  12. Proteomic comparison reveals the contribution of chloroplast to salt tolerance of a wheat introgression line

    PubMed Central

    Xu, Wenjing; Lv, Hongjun; Zhao, Mingming; Li, Yongchao; Qi, Yueying; Peng, Zhenying; Xia, Guangmin; Wang, Mengcheng

    2016-01-01

    We previously bred a salt tolerant wheat cv. SR3 with bread wheat cv. JN177 as the parent via asymmetric somatic hybridization, and found that the tolerance is partially attributed to the superior photosynthesis capacity. Here, we compared the proteomes of two cultivars to unravel the basis of superior photosynthesis capacity. In the maps of two dimensional difference gel electrophoresis (2D-DIGE), there were 26 differentially expressed proteins (DEPs), including 18 cultivar-based and 8 stress-responsive ones. 21 of 26 DEPs were identified and classified into four categories, including photosynthesis, photosynthesis system stability, linolenic acid metabolism, and protein synthesis in chloroplast. The chloroplast localization of some DEPs confirmed that the identified DEPs function in the chloroplast. The overexpression of a DEP enhanced salt tolerance in Arabidopsis thaliana. In line with these data, it is concluded that the contribution of chloroplast to high salinity tolerance of wheat cv. SR3 appears to include higher photosynthesis efficiency by promoting system protection and ROS clearance, stronger production of phytohormone JA by enhancing metabolism activity, and modulating the in chloroplast synthesis of proteins. PMID:27562633

  13. Transient foreign gene expression in chloroplasts of cultured tobacco cells after biolistic delivery of chloroplast vectors.

    PubMed Central

    Daniell, H; Vivekananda, J; Nielsen, B L; Ye, G N; Tewari, K K; Sanford, J C

    1990-01-01

    Expression of chloramphenicol acetyltransferase (cat) by suitable vectors in chloroplasts of cultured tobacco cells, delivered by high-velocity microprojectiles, is reported here. Several chloroplast expression vectors containing bacterial cat genes, placed under the control of either psbA promoter region from pea (pHD series) or rbcL promoter region from maize (pAC series) have been used in this study. In addition, chloroplast expression vectors containing replicon fragments from pea, tobacco, or maize chloroplast DNA have also been tested for efficiency and duration of cat expression in chloroplasts of tobacco cells. Cultured NT1 tobacco cells collected on filter papers were bombarded with tungsten particles coated with pUC118 (negative control), 35S-CAT (nuclear expression vector), pHD312 (repliconless chloroplast expression vector), and pHD407, pACp18, and pACp19 (chloroplast expression vectors with replicon). Sonic extracts of cells bombarded with pUC118 showed no detectable cat activity in the autoradiograms. Nuclear expression of cat reached two-thirds of the maximal 48 hr after bombardment and the maximal at 72 hr. Cells bombarded with chloroplast expression vectors showed a low level of expression until 48 hr of incubation. A dramatic increase in the expression of cat was observed 24 hr after the addition of fresh medium to cultured cells in samples bombarded with pHD407; the repliconless vector pHD312 showed about 50% of this maximal activity. The expression of nuclear cat and the repliconless chloroplast vector decreased after 72 hr, but a high level of chloroplast cat expression was maintained in cells bombarded with pHD407. Organelle-specific expression of cat in appropriate compartments was checked by introducing various plasmid constructions into tobacco protoplasts by electroporation. Although the nuclear expression vector 35S-CAT showed expression of cat, no activity was observed with any chloroplast vectors. Images PMID:2404285

  14. REDUCED CHLOROPLAST COVERAGE genes from Arabidopsis thaliana help to establish the size of the chloroplast compartment

    PubMed Central

    Larkin, Robert M.; Stefano, Giovanni; Ruckle, Michael E.; Stavoe, Andrea K.; Sinkler, Christopher A.; Brandizzi, Federica; Malmstrom, Carolyn M.; Osteryoung, Katherine W.

    2016-01-01

    Eukaryotic cells require mechanisms to establish the proportion of cellular volume devoted to particular organelles. These mechanisms are poorly understood. From a screen for plastid-to-nucleus signaling mutants in Arabidopsis thaliana, we cloned a mutant allele of a gene that encodes a protein of unknown function that is homologous to two other Arabidopsis genes of unknown function and to FRIENDLY, which was previously shown to promote the normal distribution of mitochondria in Arabidopsis. In contrast to FRIENDLY, these three homologs of FRIENDLY are found only in photosynthetic organisms. Based on these data, we proposed that FRIENDLY expanded into a small gene family to help regulate the energy metabolism of cells that contain both mitochondria and chloroplasts. Indeed, we found that knocking out these genes caused a number of chloroplast phenotypes, including a reduction in the proportion of cellular volume devoted to chloroplasts to 50% of wild type. Thus, we refer to these genes as REDUCED CHLOROPLAST COVERAGE (REC). The size of the chloroplast compartment was reduced most in rec1 mutants. The REC1 protein accumulated in the cytosol and the nucleus. REC1 was excluded from the nucleus when plants were treated with amitrole, which inhibits cell expansion and chloroplast function. We conclude that REC1 is an extraplastidic protein that helps to establish the size of the chloroplast compartment, and that signals derived from cell expansion or chloroplasts may regulate REC1. PMID:26862170

  15. Three-Dimensional Visualization of the Tubular-Lamellar Transformation of the Internal Plastid Membrane Network during Runner Bean Chloroplast Biogenesis.

    PubMed

    Kowalewska, Łucja; Mazur, Radosław; Suski, Szymon; Garstka, Maciej; Mostowska, Agnieszka

    2016-04-01

    Chloroplast biogenesis is a complex process that is integrated with plant development, leading to fully differentiated and functionally mature plastids. In this work, we used electron tomography and confocal microscopy to reconstruct the process of structural membrane transformation during the etioplast-to-chloroplast transition in runner bean (Phaseolus coccineus). During chloroplast development, the regular tubular network of paracrystalline prolamellar bodies (PLBs) and the flattened porous membranes of prothylakoids develop into the chloroplast thylakoids. Three-dimensional reconstruction is required to provide us with a more complete understanding of this transformation. We provide spatial models of the bean chloroplast biogenesis that allow such reconstruction of the internal membranes of the developing chloroplast and visualize the transformation from the tubular arrangement to the linear system of parallel lamellae. We prove that the tubular structure of the PLB transforms directly to flat slats, without dispersion to vesicles. We demonstrate that the grana/stroma thylakoid connections have a helical character starting from the early stages of appressed membrane formation. Moreover, we point out the importance of particular chlorophyll-protein complex components in the membrane stacking during the biogenesis. The main stages of chloroplast internal membrane biogenesis are presented in a movie that shows the time development of the chloroplast biogenesis as a dynamic model of this process. PMID:27002023

  16. Genome Sequences of Populus tremula Chloroplast and Mitochondrion: Implications for Holistic Poplar Breeding

    PubMed Central

    Mader, Malte; Le Paslier, Marie-Christine; Bounon, Rémi; Berard, Aurélie; Vettori, Cristina; Schroeder, Hilke; Leplé, Jean-Charles; Fladung, Matthias

    2016-01-01

    Complete Populus genome sequences are available for the nucleus (P. trichocarpa; section Tacamahaca) and for chloroplasts (seven species), but not for mitochondria. Here, we provide the complete genome sequences of the chloroplast and the mitochondrion for the clones P. tremula W52 and P. tremula x P. alba 717-1B4 (section Populus). The organization of the chloroplast genomes of both Populus clones is described. A phylogenetic tree constructed from all available complete chloroplast DNA sequences of Populus was not congruent with the assignment of the related species to different Populus sections. In total, 3,024 variable nucleotide positions were identified among all compared Populus chloroplast DNA sequences. The 5-prime part of the LSC from trnH to atpA showed the highest frequency of variations. The variable positions included 163 positions with SNPs allowing for differentiating the two clones with P. tremula chloroplast genomes (W52, 717-1B4) from the other seven Populus individuals. These potential P. tremula-specific SNPs were displayed as a whole-plastome barcode on the P. tremula W52 chloroplast DNA sequence. Three of these SNPs and one InDel in the trnH-psbA linker were successfully validated by Sanger sequencing in an extended set of Populus individuals. The complete mitochondrial genome sequence of P. tremula is the first in the family of Salicaceae. The mitochondrial genomes of the two clones are 783,442 bp (W52) and 783,513 bp (717-1B4) in size, structurally very similar and organized as single circles. DNA sequence regions with high similarity to the W52 chloroplast sequence account for about 2% of the W52 mitochondrial genome. The mean SNP frequency was found to be nearly six fold higher in the chloroplast than in the mitochondrial genome when comparing 717-1B4 with W52. The availability of the genomic information of all three DNA-containing cell organelles will allow a holistic approach in poplar molecular breeding in the future. PMID:26800039

  17. FtsHi4 Is Essential for Embryogenesis Due to Its Influence on Chloroplast Development in Arabidopsis

    PubMed Central

    Li, Shipeng; Su, Yanping; Liang, Qiuju; Meng, Hongyan; Shen, Songdong; Fan, Yunliu; Liu, Chunming; Zhang, Chunyi

    2014-01-01

    Chloroplast formation is associated with embryo development and seedling growth. However, the relationship between chloroplast differentiation and embryo development remains unclear. Five FtsHi genes that encode proteins with high similarity to FtsH proteins, but lack Zn2+-binding motifs, are present in the Arabidopsis genome. In this study, we showed that T-DNA insertion mutations in the Arabidopsis FtsHi4 gene resulted in embryo arrest at the globular-to-heart–shaped transition stage. Transmission electron microscopic analyses revealed abnormal plastid differentiation with a severe defect in thylakoid formation in the mutant embryos. Immunocytological studies demonstrated that FtsHi4 localized in chloroplasts as a thylakoid membrane-associated protein, supporting its essential role in thylakoid membrane formation. We further showed that FtsHi4 forms protein complexes, and that there was a significant reduction in the accumulation of D2 and PsbO (two photosystem II proteins) in mutant ovules. The role of FtsHi4 in chloroplast development was confirmed using an RNA-interfering approach. Additionally, mutations in other FtsHi genes including FtsHi1, FtsHi2, and FtsHi5 caused phenotypic abnormalities similar to ftshi4 with respect to plastid differentiation during embryogenesis. Taken together, our data suggest that FtsHi4, together with FtsHi1, FtsHi2, and FtsHi5 are essential for chloroplast development in Arabidopsis. PMID:24964212

  18. Defining the Core Proteome of the Chloroplast Envelope Membranes

    PubMed Central

    Simm, Stefan; Papasotiriou, Dimitrios G.; Ibrahim, Mohamed; Leisegang, Matthias S.; Müller, Bernd; Schorge, Tobias; Karas, Michael; Mirus, Oliver; Sommer, Maik S.; Schleiff, Enrico

    2013-01-01

    High-throughput protein localization studies require multiple strategies. Mass spectrometric analysis of defined cellular fractions is one of the complementary approaches to a diverse array of cell biological methods. In recent years, the protein content of different cellular (sub-)compartments was approached. Despite of all the efforts made, the analysis of membrane fractions remains difficult, in that the dissection of the proteomes of the envelope membranes of chloroplasts or mitochondria is often not reliable because sample purity is not always warranted. Moreover, proteomic studies are often restricted to single (model) species, and therefore limited in respect to differential individual evolution. In this study we analyzed the chloroplast envelope proteomes of different plant species, namely, the individual proteomes of inner and outer envelope (OE) membrane of Pisum sativum and the mixed envelope proteomes of Arabidopsis thaliana and Medicago sativa. The analysis of all three species yielded 341 identified proteins in total, 247 of them being unique. 39 proteins were genuine envelope proteins found in at least two species. Based on this and previous envelope studies we defined the core envelope proteome of chloroplasts. Comparing the general overlap of the available six independent studies (including ours) revealed only a number of 27 envelope proteins. Depending on the stringency of applied selection criteria we found 231 envelope proteins, while less stringent criteria increases this number to 649 putative envelope proteins. Based on the latter we provide a map of the outer and inner envelope core proteome, which includes many yet uncharacterized proteins predicted to be involved in transport, signaling, and response. Furthermore, a foundation for the functional characterization of yet unidentified functions of the inner and OE for further analyses is provided. PMID:23390424

  19. Chloroplasts Are Central Players in Sugar-Induced Leaf Growth.

    PubMed

    Van Dingenen, Judith; De Milde, Liesbeth; Vermeersch, Mattias; Maleux, Katrien; De Rycke, Riet; De Bruyne, Michiel; Storme, Véronique; Gonzalez, Nathalie; Dhondt, Stijn; Inzé, Dirk

    2016-05-01

    Leaves are the plant's powerhouses, providing energy for all organs through sugar production during photosynthesis. However, sugars serve not only as a metabolic energy source for sink tissues but also as signaling molecules, affecting gene expression through conserved signaling pathways to regulate plant growth and development. Here, we describe an in vitro experimental assay, allowing one to alter the sucrose (Suc) availability during early Arabidopsis (Arabidopsis thaliana) leaf development, with the aim to identify the affected cellular and molecular processes. The transfer of seedlings to Suc-containing medium showed a profound effect on leaf growth by stimulating cell proliferation and postponing the transition to cell expansion. Furthermore, rapidly after transfer to Suc, mesophyll cells contained fewer and smaller plastids, which are irregular in shape and contain fewer starch granules compared with control mesophyll cells. Short-term transcriptional responses after transfer to Suc revealed the repression of well-known sugar-responsive genes and multiple genes encoded by the plastid, on the one hand, and up-regulation of a GLUCOSE-6-PHOSPHATE TRANSPORTER (GPT2), on the other hand. Mutant gpt2 seedlings showed no stimulation of cell proliferation and no repression of chloroplast-encoded transcripts when transferred to Suc, suggesting that GPT2 plays a critical role in the Suc-mediated effects on early leaf growth. Our findings, therefore, suggest that induction of GPT2 expression by Suc increases the import of glucose-6-phosphate into the plastids that would repress chloroplast-encoded transcripts, restricting chloroplast differentiation. Retrograde signaling from the plastids would then delay the transition to cell expansion and stimulate cell proliferation. PMID:26932234

  20. Quercetin interaction with the chloroplast ATPase complex.

    PubMed

    Shoshan, V; Shahak, Y; Shavit, N

    1980-07-01

    1. Quercetin, a flavonoid which acts as an energy transfer inhibitor in photophosphorylation is shown to inhibit the P-ATP exchange activity of membrane-bound CF1 and the ATPase activity of isolated CF1. Quercetin, affects also the proton uptake in chloroplasts in a manner similar to that of dicyclohexylcarbodiimide. 2. The light-dependent proton uptake in EDTA-treated chloroplasts is stimulated by quercetin. In untreated chloroplasts quercetin has a dual effect: it enhances at pH above 7.5 while at lower pH values it decreases the extent of H+ uptake. Similar effects were obtained with dicyclohexylcarbodiimide. 3. Like quercetin, dicyclohexylcarbodiimide was also found to inhibit the ATPase activity of isolated CF1. 4. Quercetin inhibits uncoupled electron transport induced by either EDTA-treatment of chloroplasts or by addition of uncouplers. Quercetin restores H+ uptake in both types of uncoupled chloroplasts. 5. The mode of action of quercetin and dicyclohexylcarbodiimide in photophosphorylation is discussed, and interaction with both CF1 and F0 is suggested. PMID:6446936

  1. Glycolate transporter of the pea chloroplast envelope

    SciTech Connect

    Howitz, K.T.

    1985-01-01

    The discovery of a glycolate transporter in the pea (Pisum sativum) chloroplast envelope is described. Several novel silicone oil centrifugation methods were developed to resolve the initial rate kinetics of (/sup 14/C)glycolate transport by isolated, intact pea chloroplasts. Chloroplast glycolate transport was found to be carrier mediated. Transport rates saturated with increasing glycolate concentration. N-Ethylmaleimide (NEM) pretreatment of chloroplasts inhibited transport, an inhibition prevented by glycolate. Glycolate distributed across the envelope in a way which equalized stromal and medium glycolic acid concentrations, limiting possible transport mechanisms to facilitated glycolic acid diffusion, proton symport or hydroxyl antiport. The effects of stomal and medium pH's on the K/sub m/ and V/sub max/ fit the predictions of mobile carrier kinetic models of hydroxyl antiport or proton symport (H/sup +/ binds first). The carrier mediated transport was fast enough to be consistent with in vivo rates of photorespiration. The 2-hydroxymonocarboxylates, glycerate, lactate and glyoxylate are competitive inhibitors of chloroplast glycolate uptake. Glyoxylate, D-lactate and D-glycerate cause glycolate counterflow, indicating that they are also substrates of the glycolate carrier. This finding was confirmed for D-glycerate by studies on glycolate effects on (1-/sup 14/C)D-glycerate transport.

  2. Heme content and breakdown in developing chloroplasts

    SciTech Connect

    Thomas, J.; Weinstein, J.D. )

    1990-05-01

    Heme regulates tetrapyrrole biosynthesis in plants by inhibition of {delta}-aminolevulinic acid (ALA) synthesis, product inhibition of heme synthesis, and possibly other mechanisms. Plastid heme levels may be modulated by heme synthesis, breakdown and/or efflux. Heme breakdown may be catalyzed by a chloroplast localized heme oxygenase. Chloroplasts isolated from greening cucumber cotyledons were incubated in the presence or absence of various components thought to modulate heme breakdown. Following the incubations, the chloroplasts were broken (freeze-thaw) and then supplemented with horseradish peroxidase apoenzyme. The reconstituted peroxidase activity was used to determine the amount of free heme remaining (Thomas Weinstein (1989) Plant Physiol. 89S: 74). Chloroplasts, freshly isolated from seedlings greened for 16 hours, contained approximately 37 pmol heme/mg protein. When chloroplasts were incubated with 5 mM NADPH for 30 min, the endogenous heme dropped to unmeasurable levels. Exogenous heme was also broken down when NADPH was included in the incubation. Heme levels could be increased by the inclusion of 50 {mu}M ALA and/or p-hydroxymercuribenzoate. The increase due to exogenous ALA was blocked by levulinic acid, an inhibitor of ALA utilization. NADPH-dependent heme breakdown acid was inhibited by p-hydroxymercuribenzoate.

  3. Vectorial photocurrents and photoconductivity in metalized chloroplasts

    SciTech Connect

    Greenbaum, E. )

    1990-08-09

    A novel photobiophysical phenomenon was observed in isolated spinach chloroplasts that were metalized by precipitating colloidal platinum onto the surface of the thylakoid membranes. A two-point irradiation and detection system was constructed in which a continuous-beam helium-neon laser ({lambda} = 632.8 nm) was used to irradiate the platinized chloroplasts at varying perpendicular distances (Figure 1) from a single linear platinum electrode in pressure contact with the platinized chloroplasts. No external voltage bias was applied to the system. The key objective of the experiments reported in this report was to measure the relative photoconductivity of the chloroplast-metal composite matrix. Unlike conventional photosynthetic electrochemical cells, in which irradiated chloroplasts are in close proximity to an electrode or linked to the electrode by an electrode-active mediator, the flow of photocurrent was through the biocomposite material. A sustained steady-state vectorial flow of current in the plane of the entrapped composite from the point of laser irradiation to the wire electrode was measured.

  4. Isolation, quantification, and analysis of chloroplast DNA.

    PubMed

    Rowan, Beth A; Bendich, Arnold J

    2011-01-01

    Many areas of chloroplast research require methods that can assess the quality and quantity of chloroplast DNA (cpDNA). The study of chloroplast functions that depend on the proper maintenance and expression of the chloroplast genome, understanding cpDNA replication and repair, and the development of technologies for chloroplast transformation are just some of the disciplines that require the isolation of high-quality cpDNA. Arabidopsis thaliana offers several advantages for studying these processes because of the sizeable collection of mutants and natural varieties (accessions) available from stock centers and a broad community of researchers that has developed many other genetic resources. Several approaches for the isolation and quantification of cpDNA have been developed, but little consideration has been given to the strengths and weaknesses and the type of information obtained by each method, especially with respect to A. thaliana. Here, we provide protocols for obtaining high-quality cpDNA for PCR and other applications, and we evaluate several different isolation and analytical methods in order to build a robust framework for the study of cpDNA with this model organism. PMID:21822838

  5. Proteomic Insight into the Response of Arabidopsis Chloroplasts to Darkness

    PubMed Central

    Wang, Jing; Yu, Qingbo; Xiong, Haibo; Wang, Jun; Chen, Sixue; Yang, Zhongnan; Dai, Shaojun

    2016-01-01

    Chloroplast function in photosynthesis is essential for plant growth and development. It is well-known that chloroplasts respond to various light conditions. However, it remains poorly understood about how chloroplasts respond to darkness. In this study, we found 81 darkness-responsive proteins in Arabidopsis chloroplasts under 8 h darkness treatment. Most of the proteins are nucleus-encoded, indicating that chloroplast darkness response is closely regulated by the nucleus. Among them, 17 ribosome proteins were obviously reduced after darkness treatment. The protein expressional patterns and physiological changes revealed the mechanisms in chloroplasts in response to darkness, e.g., (1) inhibition of photosystem II resulted in preferential cyclic electron flow around PSI; (2) promotion of starch degradation; (3) inhibition of chloroplastic translation; and (4) regulation by redox and jasmonate signaling. The results have improved our understanding of molecular regulatory mechanisms in chloroplasts under darkness. PMID:27137770

  6. A Putative Chloroplast Thylakoid Metalloprotease VIRESCENT3 Regulates Chloroplast Development in Arabidopsis thaliana.

    PubMed

    Qi, Yafei; Liu, Xiayan; Liang, Shuang; Wang, Rui; Li, Yuanfeng; Zhao, Jun; Shao, Jingxia; An, Lijun; Yu, Fei

    2016-02-12

    The chloroplast is the site of photosynthesis and many other essential plant metabolic processes, and chloroplast development is an integral part of plant growth and development. Mutants defective in chloroplast development can display various color phenotypes including the intriguing virescence phenotype, which shows yellow/white coloration at the leaf base and greening toward the leaf tip. Through large scale genetic screens, we identified a series of new virescent mutants including virescent3-1 (vir3-1), vir4-1, and vir5-1 in Arabidopsis thaliana. We showed that VIR3 encodes a putative chloroplast metalloprotease by map-based cloning. Through site-directed mutagenesis, we showed that the conserved histidine 235 residue in the zinc binding motif HEAGH of VIR3 is indispensable for VIR3 accumulation in the chloroplast. The chloroplast localization of VIR3 was confirmed by the transient expression of VIR3-GFP in leaf protoplasts. Furthermore, taking advantage of transgenic lines expressing VIR3-FLAG, we demonstrated that VIR3 is an intrinsic thylakoid membrane protein that mainly resides in the stromal lamellae. Moreover, topology analysis using transgenic lines expressing a dual epitope-tagged VIR3 indicated that both the N and C termini of VIR3 are located in the stroma, and the catalytic domain of VIR3 is probably facing the stroma. Blue native gel analysis indicated that VIR3 is likely present as a monomer or part of a small complex in the thylakoid membrane. This work not only implicates VIR3 as a new factor involved in early chloroplast development but also provides more insight into the roles of chloroplast proteases in chloroplast biogenesis. PMID:26702056

  7. Analyses of Charophyte Chloroplast Genomes Help Characterize the Ancestral Chloroplast Genome of Land Plants

    PubMed Central

    Civáň, Peter; Foster, Peter G.; Embley, Martin T.; Séneca, Ana; Cox, Cymon J.

    2014-01-01

    Despite the significance of the relationships between embryophytes and their charophyte algal ancestors in deciphering the origin and evolutionary success of land plants, few chloroplast genomes of the charophyte algae have been reconstructed to date. Here, we present new data for three chloroplast genomes of the freshwater charophytes Klebsormidium flaccidum (Klebsormidiophyceae), Mesotaenium endlicherianum (Zygnematophyceae), and Roya anglica (Zygnematophyceae). The chloroplast genome of Klebsormidium has a quadripartite organization with exceptionally large inverted repeat (IR) regions and, uniquely among streptophytes, has lost the rrn5 and rrn4.5 genes from the ribosomal RNA (rRNA) gene cluster operon. The chloroplast genome of Roya differs from other zygnematophycean chloroplasts, including the newly sequenced Mesotaenium, by having a quadripartite structure that is typical of other streptophytes. On the basis of the improbability of the novel gain of IR regions, we infer that the quadripartite structure has likely been lost independently in at least three zygnematophycean lineages, although the absence of the usual rRNA operonic synteny in the IR regions of Roya may indicate their de novo origin. Significantly, all zygnematophycean chloroplast genomes have undergone substantial genomic rearrangement, which may be the result of ancient retroelement activity evidenced by the presence of integrase-like and reverse transcriptase-like elements in the Roya chloroplast genome. Our results corroborate the close phylogenetic relationship between Zygnematophyceae and land plants and identify 89 protein-coding genes and 22 introns present in the chloroplast genome at the time of the evolutionary transition of plants to land, all of which can be found in the chloroplast genomes of extant charophytes. PMID:24682153

  8. Analyses of charophyte chloroplast genomes help characterize the ancestral chloroplast genome of land plants.

    PubMed

    Civaň, Peter; Foster, Peter G; Embley, Martin T; Séneca, Ana; Cox, Cymon J

    2014-04-01

    Despite the significance of the relationships between embryophytes and their charophyte algal ancestors in deciphering the origin and evolutionary success of land plants, few chloroplast genomes of the charophyte algae have been reconstructed to date. Here, we present new data for three chloroplast genomes of the freshwater charophytes Klebsormidium flaccidum (Klebsormidiophyceae), Mesotaenium endlicherianum (Zygnematophyceae), and Roya anglica (Zygnematophyceae). The chloroplast genome of Klebsormidium has a quadripartite organization with exceptionally large inverted repeat (IR) regions and, uniquely among streptophytes, has lost the rrn5 and rrn4.5 genes from the ribosomal RNA (rRNA) gene cluster operon. The chloroplast genome of Roya differs from other zygnematophycean chloroplasts, including the newly sequenced Mesotaenium, by having a quadripartite structure that is typical of other streptophytes. On the basis of the improbability of the novel gain of IR regions, we infer that the quadripartite structure has likely been lost independently in at least three zygnematophycean lineages, although the absence of the usual rRNA operonic synteny in the IR regions of Roya may indicate their de novo origin. Significantly, all zygnematophycean chloroplast genomes have undergone substantial genomic rearrangement, which may be the result of ancient retroelement activity evidenced by the presence of integrase-like and reverse transcriptase-like elements in the Roya chloroplast genome. Our results corroborate the close phylogenetic relationship between Zygnematophyceae and land plants and identify 89 protein-coding genes and 22 introns present in the chloroplast genome at the time of the evolutionary transition of plants to land, all of which can be found in the chloroplast genomes of extant charophytes. PMID:24682153

  9. Protein methylation reactions in intact pea chloroplasts

    SciTech Connect

    Niemi, K.J. )

    1989-04-01

    Post-translational protein methylation was investigated in Pisum sativum chloroplasts. Intact pea chloroplasts were incubated with ({sup 3}H-methyl)-S-adenosylmethionine under various conditions. The chloroplasts were then separated into stromal and thylakoid fractions and analyzed for radioactivity transferred to protein. Light enhanced the magnitude of labeling in both fractions. One thylakoid polypeptide with an apparent molecular mass of 43 kDa was labeled only in the light. Several other thylakoid and stromal proteins were labeled in both light and dark-labeling conditions. Both base-labile methylation, carboxy-methylesters and base-stable groups, N-methylations were found. Further characterization of the methyl-transfer reactions will be presented.

  10. Chloroplast-Diphenyl Ether Interactions II 1

    PubMed Central

    Wettlaufer, S. H.; Alscher, Ruth; Strick, Christine

    1985-01-01

    Acifluorfen, a p-nitrodiphenyl ether herbicide, is inhibitory to those photosynthetic functions that require a functioning chloroplast envelope. Functions involving the stroma are also affected. Acifluorfen does not lyse intact spinach chloroplasts, yet does increase the sensitivity of CO2-dependent O2 evolution to exogenous inorganic phosphate without directly affecting the function of the phosphate translocator. Acifluorfen penetrates into the chloroplast stroma in a light-independent fashion. Once inside, it causes the inactivation of light and dithiothreitol-activated fructose 1,6-bisphosphatase. Light-activated glyceraldehyde-3-phosphate dehydrogenase (NADP) is also inactivated by acifluorfen. These data suggest that acifluorfen stimulates a pathway for inactivation of fructose 1,6-bisphosphatase and glyceraldehyde 3-phosphate dehydrogenase (NADP) which uses oxygen as a terminal oxidant and which involves thioredoxin and ferredoxin-thioredoxin reductase. PMID:16664219

  11. The chloroplast genome exists in multimeric forms

    SciTech Connect

    Deng, Xingwang; Wing, R.A.; Gruissem, W. )

    1989-06-01

    Chloroplast DNA conformation was analyzed by pulse-field gel electrophoresis. The authors found that spinach leaf chloroplast DNA molecules exist in at least four distinct forms with the apparent molecular weights of monomer, dimer, trimer, and tetramer. Two-dimensional gel analysis of DNA after UV nicking and in the presence of ethidium bromide indicates that they are not isomers that differ in superhelical density. DNA gyrase decatenation analysis demonstrates that the majority of the DNA molecules are oligomers rather than catenanes. The relative amounts of monomer, dimer, trimer, and tetramer forms, quantitated by molecular hybridization, are 1, 1/3, 1/9, and 1/27, respectively, and do not change during leaf maturation. The possible mechanisms of chloroplast DNA oligomer formation are discussed.

  12. Chloroplast Response to Low Leaf Water Potentials

    PubMed Central

    Keck, R. W.; Boyer, J. S.

    1974-01-01

    Cyclic and noncyclic photophosphorylation and electron transport by photosystem 1, photosystem 2, and from water to methyl viologen (“whole chain”) were studied in chloroplasts isolated from sunflower (Helianthus annus L. var Russian Mammoth) leaves that had been desiccated to varying degrees. Electron transport showed considerable inhibition at leaf water potentials of −9 bars when the chloroplasts were exposed to an uncoupler in vitro, and it continued to decline in activity as leaf water potentials decreased. Electron transport by photosystem 2 and coupled electron transport by photosystem 1 and the whole chain were unaffected at leaf water potentials of −10 to −11 bars but became progressively inhibited between leaf water potentials of −11 and −17 bars. A low, stable activity remained at leaf water potentials below −17 bars. In contrast, both types of photophosphorylation were unaffected by leaf water potentials of −10 to −11 bars, but then ultimately became zero at leaf water potentials of −17 bars. Although the chloroplasts isolated from the desiccated leaves were coupled at leaf water potentials of −11 to −12 bars, they became progressively uncoupled as leaf water potentials decreased to −17 bars. Abscisic acid and ribonuclease had no effect on chloroplast photophosphorylation. The results are generally consistent with the idea that chloroplast activity begins to decrease at the same leaf water potentials that cause stomatal closure in sunflower leaves and that chloroplast electron transport begins to limit photosynthesis at leaf water potentials below about −11 bars. However, it suggests that, during severe desiccation, the limitation may shift from electron transport to photophosphorylation. PMID:16658727

  13. Chloroplast signaling within, between and beyond cells

    PubMed Central

    Bobik, Krzysztof; Burch-Smith, Tessa M.

    2015-01-01

    The most conspicuous function of plastids is the 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 plastids possess their own genomes 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 nuclei, 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 less widely known 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 to avoid

  14. Tic32, an essential component in chloroplast biogenesis.

    PubMed

    Hörmann, Friederike; Küchler, Michael; Sveshnikov, Dmitry; Oppermann, Udo; Li, Yong; Soll, Jürgen

    2004-08-13

    Chloroplast protein import across the inner envelope is facilitated by the translocon of the inner envelope of chloroplasts (Tic). Here we have identified Tic32 as a novel subunit of the Tic complex. Tic32 can be purified from solubilized inner envelope membranes by chromatography on Tic110 containing affinity matrix. Co-immunoprecipitation experiments using either Tic32 or Tic110 antisera indicated a tight association between these polypeptides as well as with other Tic subunits, e.g. Tic40, Tic22, or Tic62, whereas the outer envelope protein Toc75 was not found in this complex. Chemical cross-linking suggests that Tic32 is involved late in the overall translocation process, because both the precursor form as well as the mature form of Rubisco small subunit can be detected. We were unable to isolate Arabidopsis null mutants of the attic32 gene, indicating that Tic32 is essential for viability. Deletion of the attic32 gene resulted in early seed abortion because the embryo was unable to differentiate from the heart stage to the torpedo stage. The homology of Tic32 to short-chain dehydrogenases suggests a dual role of Tic32 in import, one as a regulatory component and one as an important subunit in the assembly of the entire complex. PMID:15180984

  15. Chloroplast-derived enzyme cocktails hydrolyse lignocellulosic biomass and release fermentable sugars

    PubMed Central

    Verma, Dheeraj; Kanagaraj, Anderson; Jin, Shuangxia; Singh, Nameirakpam D.; Kolattukudy, Pappachan E; Daniell, Henry

    2009-01-01

    Summary It is widely recognized that biofuel production from lignocellulosic materials is limited by inadequate technology to efficiently and economically release fermentable sugars from the complex multi-polymeric raw materials. Therefore, endoglucanases, exoglucanase, pectate lyases, cutinase, swollenin, xylanase, acetyl xylan esterase, beta glucosidase and lipase genes from bacteria or fungi were expressed in E. coli or tobacco chloroplasts. A PCR based method was used to clone genes without introns from Trichoderma reesei genomic DNA. Homoplasmic transplastomic lines showed normal phenotype and were fertile. Based on observed expression levels, up to 49, 64 and 10,751 million units of pectate lyases or endoglucanase can be produced annually, per acre of tobacco. Plant production cost of endoglucanase is 3,100-fold and pectate lyase is 1,057 or 1,480 fold lower than the same recombinant enzymes sold commercially, produced via fermentation. Chloroplast-derived enzymes had higher temperature stability and wider pH optima than enzymes expressed in E. coli. Plant crude-extracts showed higher enzyme activity than E. coli with increasing protein concentration, demonstrating their direct utility without purification. Addition of E. coli extracts to the chloroplast-derived enzymes significantly decreased their activity. Chloroplast-derived crude-extract enzyme cocktails yielded more (up to 3,625%) glucose from filter paper, pine wood or citrus peel than commercial cocktails. Furthermore, pectate lyase transplastomic plants showed enhanced resistance to Erwina soft rot. This is the first report of using plant-derived enzyme cocktails for production of fermentable sugars from lignocellulosic biomass. Limitations of higher cost and lower production capacity of fermentation systems are addressed by chloroplast-derived enzyme cocktails. PMID:20070870

  16. The evolution of chloroplast RNA editing.

    PubMed

    Tillich, Michael; Lehwark, Pascal; Morton, Brian R; Maier, Uwe G

    2006-10-01

    RNA editing alters the nucleotide sequence of an RNA molecule so that it deviates from the sequence of its DNA template. Different RNA-editing systems are found in the major eukaryotic lineages, and these systems are thought to have evolved independently. In this study, we provide a detailed analysis of data on C-to-U editing sites in land plant chloroplasts and propose a model for the evolution of RNA editing in land plants. First, our data suggest that the limited RNA-editing system of seed plants and the much more extensive systems found in hornworts and ferns are of monophyletic origin. Further, although some eukaryotic editing systems appear to have evolved to regulate gene expression, or at least are now involved in gene regulation, there is no evidence that RNA editing plays a role in gene regulation in land plant chloroplasts. Instead, our results suggest that land plant chloroplast C-to-U RNA editing originated as a mechanism to generate variation at the RNA level, which could complement variation at the DNA level. Under this model, many of the original sites, particularly in seed plants, have been subsequently lost due to mutation at the DNA level, and the function of extant sites is merely to conserve certain codons. This is the first comprehensive model for the evolution of the chloroplast RNA-editing system of land plants and may also be applicable to the evolution of RNA editing in plant mitochondria. PMID:16835291

  17. From extracellular to intracellular: the establishment of mitochondria and chloroplasts.

    PubMed

    Whatley, J M; John, P; Whatley, F R

    1979-04-11

    Paracoccus and Rhodopseudomonas are unusual among bacteria in having a majority of the biochemical features of mitochondria; blue-green algae have many of the features of chloroplasts. The theory of serial endosymbiosis proposes that a primitive eukaryote successively took up bacteria and blue-green algae to yield mitochondria and chloroplasts respectively. Possible characteristics of transitional forms are indicated both by the primitive amoeba, Pelomyxa, which lacks mitochondria but contains a permanent population of endosymbiotic bacteria, and by several anomalous eukaryotic algae, e.g. Cyanophora, which contain cyanelles instead of chloroplasts. Blue-green algae appear to be obvious precursors of red algal chloroplasts but the ancestry of other chloroplasts is less certain, though the epizoic symbiont, Prochloron, may resemble the ancestral green algal chloroplast. We speculate that the chloroplasts of the remaining algae may have been a eukaryotic origin. The evolution or organelles from endosymbiotic precursors would involve their integration with the host cell biochemically, structurally and numerically. PMID:36620

  18. Motif analysis unveils the possible co-regulation of chloroplast genes and nuclear genes encoding chloroplast proteins.

    PubMed

    Wang, Ying; Ding, Jun; Daniell, Henry; Hu, Haiyan; Li, Xiaoman

    2012-09-01

    Chloroplasts play critical roles in land plant cells. Despite their importance and the availability of at least 200 sequenced chloroplast genomes, the number of known DNA regulatory sequences in chloroplast genomes are limited. In this paper, we designed computational methods to systematically study putative DNA regulatory sequences in intergenic regions near chloroplast genes in seven plant species and in promoter sequences of nuclear genes in Arabidopsis and rice. We found that -35/-10 elements alone cannot explain the transcriptional regulation of chloroplast genes. We also concluded that there are unlikely motifs shared by intergenic sequences of most of chloroplast genes, indicating that these genes are regulated differently. Finally and surprisingly, we found five conserved motifs, each of which occurs in no more than six chloroplast intergenic sequences, are significantly shared by promoters of nuclear-genes encoding chloroplast proteins. By integrating information from gene function annotation, protein subcellular localization analyses, protein-protein interaction data, and gene expression data, we further showed support of the functionality of these conserved motifs. Our study implies the existence of unknown nuclear-encoded transcription factors that regulate both chloroplast genes and nuclear genes encoding chloroplast protein, which sheds light on the understanding of the transcriptional regulation of chloroplast genes. PMID:22733202

  19. Changes in Chloroplast mRNA Stability during Leaf Development.

    PubMed Central

    Klaff, P; Gruissem, W

    1991-01-01

    During spinach leaf development, chloroplast-encoded mRNAs accumulate to different steady-state levels. Their relative transcription rates alone, however, cannot account for the changes in mRNA amount. In this study, we examined the importance of mRNA stability for the regulation of plastid mRNA accumulation using an in vivo system to measure mRNA decay in intact leaves by inhibiting transcription with actinomycin D. Decay of psbA and rbcL mRNAs was assayed in young and mature leaves. The psbA mRNA half-life was increased more than twofold in mature leaves compared with young leaves, whereas rbcL mRNA decayed with a similar relative half-life at both leaf developmental stages. The direct in vivo measurements demonstrated that differential mRNA stability in higher plant plastids can account for differences in mRNA accumulation during leaf development. The role of polysome association in mRNA decay was also investigated. Using organelle-specific translation inhibitors that force mRNAs into a polysome-bound state or deplete mRNAs of ribosomes, we measured mRNA decay in vivo in either state. The results showed that rbcL and psbA mRNAs are less stable when bound to polysomes relative to the polysome-depleted mRNAs and that their stabilities are differentially affected by binding to polysomes. The results suggested that ribosome binding and/or translation of the psbA and rbcL mRNAs may function to modulate the rate of their decay in chloroplasts. PMID:12324602

  20. Polymorphic simple sequence repeat regions in chloroplast genomes: applications to the population genetics of pines.

    PubMed Central

    Powell, W; Morgante, M; McDevitt, R; Vendramin, G G; Rafalski, J A

    1995-01-01

    Simple sequence repeats (SSRs), consisting of tandemly repeated multiple copies of mono-, di-, tri-, or tetranucleotide motifs, are ubiquitous in eukaryotic genomes and are frequently used as genetic markers, taking advantage of their length polymorphism. We have examined the polymorphism of such sequences in the chloroplast genomes of plants, by using a PCR-based assay. GenBank searches identified the presence of several (dA)n.(dT)n mononucleotide stretches in chloroplast genomes. A chloroplast (cp) SSR was identified in three pine species (Pinus contorta, Pinus sylvestris, and Pinus thunbergii) 312 bp upstream of the psbA gene. DNA amplification of this repeated region from 11 pine species identified nine length variants. The polymorphic amplified fragments were isolated and the DNA sequence was determined, confirming that the length polymorphism was caused by variation in the length of the repeated region. In the pines, the chloroplast genome is transmitted through pollen and this PCR assay may be used to monitor gene flow in this genus. Analysis of 305 individuals from seven populations of Pinus leucodermis Ant. revealed the presence of four variants with intrapopulational diversities ranging from 0.000 to 0.629 and an average of 0.320. Restriction fragment length polymorphism analysis of cpDNA on the same populations previously failed to detect any variation. Population subdivision based on cpSSR was higher (Gst = 0.22, where Gst is coefficient of gene differentiation) than that revealed in a previous isozyme study (Gst = 0.05). We anticipate that SSR loci within the chloroplast genome should provide a highly informative assay for the analysis of the genetic structure of plant populations. Images Fig. 2 PMID:7644491

  1. Effect of light intensity on pigments and main acyl lipids during 'natural' chloroplast development in wheat seedlings.

    PubMed

    Lechowicz, W; Maternicka, K; Faltynowicz, M; Poskuta, J

    1986-01-01

    The content and composition of pigments and acyl lipids (monogalactosyl diacylglycerol, digalactosyl diacylglycerol and phosphatidyl glycerol) have been investigated in developing chloroplasts isolated from successive 2-cm sections along the leaves of wheat seedlings grown either under 100, 30 or 3 W·m(-2). In all examined stages of plastid development chlorophyll a/b and chlorophyll/carotenoid ratios were higher with increasing irradiance, whereas chlorophyll content expressed on fresh weight basis gradually decreased.Concentrations of monogalactosyl diacylglycerol, digalactosyl diacylglycerol and phosphatidyl glycerol decreased per chlorophyll unit with increasing plastid maturity. The higher was the light intensity applied during plant growth, the higher were galactolipid and phosphatidyl glycerol contents in developing chloroplasts. During plastid development the percentage of α-linolenic acid markedly increased in total and individual acyl lipids. Under high light conditions, the accumulation of this fatty acid proceeded more rapidly. Significantly higher proportion of α-linolenic acid was found in acyl lipid fraction of chloroplasts differentiating in high light grown plants, than in those from plants exposed to lower light intensities. The differences in the double bond index may indicate higher fluidity of thylakoid membranes in sun-type chloroplasts.Trans-3Δ-hexadecenoic acid, virtually absent in the youngest plastids, was found in much higher concentration (per chlorophyll unit and as mol % of phosphatidyl glycerol fatty acids) in chloroplasts developing at high light conditions. PMID:24443210

  2. Direct Chloroplast Sequencing: Comparison of Sequencing Platforms and Analysis Tools for Whole Chloroplast Barcoding

    PubMed Central

    Brozynska, Marta; Furtado, Agnelo; Henry, Robert James

    2014-01-01

    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. PMID:25329378

  3. The complete chloroplast genome of Cynara humilis.

    PubMed

    Curci, Pasquale Luca; Sonnante, Gabriella

    2016-07-01

    The complete chloroplast genome of the wild thistle Cynara humilis L. (Asteraceae) is presented in this study. The genome is 152,585 bp in length and has a quadripartite structure composed by a large single-copy (LSC) of 83,622 bp, a small single-copy (SSC) of 18,651 bp and two inverted repeats (IRb/a) of 25,156 bp each. The GC content corresponds to 37.7%. The amount of unique genes is 114, in which 17 are duplicated in the IRs, for a total of 131 genes. A maximum parsimony phylogenetic analysis revealed that C. humilis chloroplast genome is closely related to that of the globe artichoke within the Carduoideae subfamily. PMID:25812057

  4. Isolation and identification of chloroplast lipids.

    PubMed

    Sato, Norihiro; Tsuzuki, Mikio

    2011-01-01

    Glycerolipids of photosynthetic organisms are accounted for largely by thylakoid membrane lipids consisting of chloroplast-specific glycolipids such as monogalactosyl diacylglycerol, digalactosyl diacylglycerol, and sulfoquinovosyl diacylglycerol, and a sole phospholipid, phosphatidylglycerol. In this chapter, methods for characterization of lipids from plant cells are described. The methods include extraction of total lipids from the cells, separation of these lipids into individual lipid classes by thin-layer chromatography, and identification of respective lipid classes by their mobility. We also present methods for the determination of compositions of constituent fatty acids, distribution of fatty acids between sn-1 and sn-2 positions, and determination of contents of individual lipid classes by gas-liquid chromatography. These methods are applicable to isolated chloroplasts or some membrane fractions such as thylakoid membranes. PMID:20960124

  5. Posttranslational Modifications of Chloroplast Proteins: An Emerging Field1

    PubMed Central

    2015-01-01

    Posttranslational modifications of proteins are key effectors of enzyme activity, protein interactions, targeting, and turnover rate, but despite their importance, they are still poorly understood in plants. Although numerous reports have revealed the regulatory role of protein phosphorylation in photosynthesis, various other protein modifications have been identified in chloroplasts only recently. It is known that posttranslational Nα-acetylation occurs in both nuclear- and plastid-encoded chloroplast proteins, but the physiological significance of this acetylation is not yet understood. Lysine acetylation affects the localization and activity of key metabolic enzymes, and it may work antagonistically or cooperatively with lysine methylation, which also occurs in chloroplasts. In addition, tyrosine nitration may help regulate the repair cycle of photosystem II, while N-glycosylation determines enzyme activity of chloroplastic carbonic anhydrase. This review summarizes the progress in the research field of posttranslational modifications of chloroplast proteins and points out the importance of these modifications in the regulation of chloroplast metabolism. PMID:25911530

  6. Action of Nalidixic Acid on Chloroplast Replication in Euglena gracilis.

    PubMed

    Lyman, H; Jupp, A S; Larrinua, I

    1975-02-01

    The role of light in nalidixic acid bleaching of Euglena gracilis var. bacillaris was investigated. The kinetics of loss of the chloroplast-associated DNA and the sensitivity of chloroplast replication to ultraviolet light was followed during treatment with nalidixic acid. By using the mutant P(4)ZUL, and 3-(3,4-dichlorophenyl)-, 1-dimethylurea, it was demonstrated that the requirement for light was a functioning photosynthetic electron transport system. Ultracentifugal analysis showed a substantial decrease in chloroplast-associated DNA after 6 hours of treatment with nalidixic acid. Ultraviolet target analysis revealed that the number of chloroplast genomes per cell had been reduced. The possible role of light and implications of the reduction in chloroplast genomes for chloroplast replication are discussed. PMID:16659089

  7. Posttranslational Modifications of Chloroplast Proteins: An Emerging Field.

    PubMed

    Lehtimäki, Nina; Koskela, Minna M; Mulo, Paula

    2015-07-01

    Posttranslational modifications of proteins are key effectors of enzyme activity, protein interactions, targeting, and turnover rate, but despite their importance, they are still poorly understood in plants. Although numerous reports have revealed the regulatory role of protein phosphorylation in photosynthesis, various other protein modifications have been identified in chloroplasts only recently. It is known that posttranslational N(α)-acetylation occurs in both nuclear- and plastid-encoded chloroplast proteins, but the physiological significance of this acetylation is not yet understood. Lysine acetylation affects the localization and activity of key metabolic enzymes, and it may work antagonistically or cooperatively with lysine methylation, which also occurs in chloroplasts. In addition, tyrosine nitration may help regulate the repair cycle of photosystem II, while N-glycosylation determines enzyme activity of chloroplastic carbonic anhydrase. This review summarizes the progress in the research field of posttranslational modifications of chloroplast proteins and points out the importance of these modifications in the regulation of chloroplast metabolism. PMID:25911530

  8. Protein methylation in pea chloroplasts. [Pisum sativum

    SciTech Connect

    Niemi, K.J.; Adler, J.; Selman, B.R. )

    1990-07-01

    The methylation of chloroplast proteins has been investigated by incubating intact pea (Pisum sativum) chloroplasts with ({sup 3}H-methyl)-S-adenosylmethionine. Incubation in the light increases the amount of methylation in both the thylakoid and stromal fractions. Numerous thylakoid proteins serve as substrates for the methyltransfer reactions. Three of these thylakoid proteins are methylated to a significantly greater extent in the light than in the dark. The primary stromal polypeptide methylated is the large subunit of ribulose bisphosphate carboxylase/oxygenase. One other stromal polypeptide is also methylated much more in the light than in the dark. Two distinct types of protein methylation occur. One methylinkage is stable to basic conditions whereas a second type is base labile. The base-stable linkage is indicative of N-methylation of amino acid residues while base-lability is suggestive of carboxymethylation of amino acid residues. Labeling in the light increases the percentage of methylation that is base labile in the thylakoid fraction while no difference is observed in the amount of base-labile methylations in light-labeled and dark-labeled stromal proteins. Also suggestive of carboxymethylation is the detection of volatile ({sup 3}H)methyl radioactivity which increases during the labeling period and is greater in chloroplasts labeled in the light as opposed to being labeled in the dark; this implies in vivo turnover of the ({sup 3}H)methyl group.

  9. Chloroplast Genome Evolution in the Euglenaceae.

    PubMed

    Bennett, Matthew S; Triemer, Richard E

    2015-01-01

    Over the last few years multiple studies have been published outlining chloroplast genomes that represent many of the photosynthetic euglenid genera. However, these genomes were scattered throughout the euglenophyceaean phylogenetic tree, and focused on comparisons with Euglena gracilis. Here, we present a study exclusively on taxa within the Euglenaceae. Six new chloroplast genomes were characterized, those of Cryptoglena skujai, E. gracilis var. bacillaris, Euglena viridis, Euglenaria anabaena, Monomorphina parapyrum, and Trachelomonas volvocina, and added to six previously published chloroplast genomes to determine if trends existed within the family. With this study: at least one genome has now been characterized for each genus, the genomes of different strains from two taxa were characterized to explore intraspecific variability, and a second taxon has been characterized for the genus Monomorphina to examine intrageneric variability. Overall results showed a large amount of variability among the genomes, though a few trends could be identified both within Euglenaceae and within Euglenophyta. In addition, the intraspecific analysis indicated that the similarity of a genome sequence between strains was taxon dependent, and the intrageneric analysis indicated that the majority of the evolutionary changes within the Euglenaceae occurred intergenerically. PMID:25976746

  10. A Large Population of Small Chloroplasts in Tobacco Leaf Cells Allows More Effective Chloroplast Movement Than a Few Enlarged Chloroplasts1

    PubMed Central

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

    2002-01-01

    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. PMID:12011343

  11. Cold-induced responses in annual bluegrass genotypes with differential resistance to pink snow mold (Microdochium nivale).

    PubMed

    Bertrand, Annick; Castonguay, Yves; Azaiez, Aïda; Hsiang, Tom; Dionne, Julie

    2011-01-01

    Greens-type annual bluegrass (Poa annua L.) is susceptible to winter stresses including subfreezing temperatures and pink snow mold (SM). To better understand the mechanisms of SM resistance in annual bluegrass, four SM-resistant and four SM-sensitive genotypes were incubated at low temperature with Microdochium nivale (Fries) Samuels & Hallett, the causal agent of pink snow mold. We assessed the impact of a 6-week incubation period with SM at 2 °C under high humidity (≥ 98%) on the accumulation of cold-induced metabolites and on freezing tolerance. Incubation of annual bluegrass inoculated with SM lead to a major decrease in concentration of cryoprotective sugars such as sucrose and HDP (high degree of polymerization) fructans. Conversely, major amino acids linked to stress resistance such as glutamine and arginine increased in crowns of annual bluegrass in response to SM inoculation. One of the major differences between resistant and sensitive genotypes was found in the concentration of HDP fructans, which remained higher in SM-resistant genotypes throughout the incubation period. HDP fructans were also more abundant in freeze-tolerant genotypes, reinforcing their positive impact on winter survival of annual bluegrass. The identification of genotypes that are resistant to both SM and freezing shows the possibility of being able to improve both traits concomitantly. PMID:21421353

  12. Preparation of stroma, thylakoid membrane, and lumen fractions from Arabidopsis thaliana chloroplasts for proteomic analysis.

    PubMed

    Hall, Michael; Mishra, Yogesh; Schröder, Wolfgang P

    2011-01-01

    For many studies regarding important chloroplast processes such as oxygenic photosynthesis, fractionation of the total chloroplast proteome is a necessary first step. Here, we describe a method for isolating the stromal, the thylakoid membrane, and the thylakoid lumen subchloroplast fractions from Arabidopsis thaliana leaf material. All three fractions can be isolated sequentially from the same plant material in a single day preparation. The isolated fractions are suitable for various proteomic analyses such as simple mapping studies or for more complex experiments such as differential expression analysis using two-dimensional difference gel electrophoresis (2D-DIGE) or mass spectrometry (MS)-based techniques. Besides this, the obtained fractions can also be used for many other purposes such as immunological assays, enzymatic activity assays, and studies of protein complexes by native-polyacrylamide gel electrophoresis (native-PAGE). PMID:21863445

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

    PubMed Central

    Maier, Uwe G; Bozarth, Andrew; Funk, Helena T; Zauner, Stefan; Rensing, Stefan A; Schmitz-Linneweber, Christian; Börner, Thomas; Tillich, Michael

    2008-01-01

    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. PMID:18755031

  14. The complete chloroplast genome of Phalaenopsis "Tiny Star".

    PubMed

    Kim, Goon-Bo; Kwon, Youngeun; Yu, Hee-Ju; Lim, Ki-Byung; Seo, Jae-Hwan; Mun, Jeong-Hwan

    2016-01-01

    We determined the complete chloroplast DNA sequence of Phalaenopsis "Tiny Star" based on Illumina sequencing. The total length of the chloroplast genome is 148,918 bp long with GC content of 36.7%. It contains 70 protein-coding genes, 30 tRNA genes, and 4 rRNA genes. Comparative analysis with the reported orchid chloroplast sequences identified unique InDel variations in the "Tiny Star" chloroplast genome that have potential as genetic markers to investigate the maternal lineage of Phalaenopsis and Doritaenopsis cultivars. PMID:25093401

  15. Fine tuning chloroplast movements through physical interactions between phototropins

    PubMed Central

    Sztatelman, Olga; Łabuz, Justyna; Hermanowicz, Paweł; Banaś, Agnieszka Katarzyna; Bażant, Aneta; Zgłobicki, Piotr; Aggarwal, Chhavi; Nadzieja, Marcin; Krzeszowiec, Weronika; Strzałka, Wojciech; Gabryś, Halina

    2016-01-01

    Phototropins are plant photoreceptors which regulate numerous responses to blue light, including chloroplast relocation. Weak blue light induces chloroplast accumulation, whereas strong light leads to an avoidance response. Two Arabidopsis phototropins are characterized by different light sensitivities. Under continuous light, both can elicit chloroplast accumulation, but the avoidance response is controlled solely by phot2. As well as continuous light, brief light pulses also induce chloroplast displacements. Pulses of 0.1s and 0.2s of fluence rate saturating the avoidance response lead to transient chloroplast accumulation. Longer pulses (up to 20s) trigger a biphasic response, namely transient avoidance followed by transient accumulation. This work presents a detailed study of transient chloroplast responses in Arabidopsis. Phototropin mutants display altered chloroplast movements as compared with the wild type: phot1 is characterized by weaker responses, while phot2 exhibits enhanced chloroplast accumulation, especially after 0.1s and 0.2s pulses. To determine the cause of these differences, the abundance and phosphorylation levels of both phototropins, as well as the interactions between phototropin molecules are examined. The formation of phototropin homo- and heterocomplexes is the most plausible explanation of the observed phenomena. The physiological consequences of this interplay are discussed, suggesting the universal character of this mechanism that fine-tunes plant reactions to blue light. Additionally, responses in mutants of different protein phosphatase 2A subunits are examined to assess the role of protein phosphorylation in signaling of chloroplast movements. PMID:27406783

  16. Copper Delivery to Chloroplast Proteins and its Regulation

    PubMed Central

    Aguirre, Guadalupe; Pilon, Marinus

    2016-01-01

    Copper is required for photosynthesis in chloroplasts of plants because it is a cofactor of plastocyanin, an essential electron carrier in the thylakoid lumen. Other chloroplast copper proteins are copper/zinc superoxide dismutase and polyphenol oxidase, but these proteins seem to be dispensable under conditions of low copper supply when transcripts for these proteins undergo microRNA-mediated down regulation. Two ATP-driven copper transporters function in tandem to deliver copper to chloroplast compartments. This review seeks to summarize the mechanisms of copper delivery to chloroplast proteins and its regulation. We also delineate some of the unanswered questions that still remain in this field. PMID:26793223

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

    NASA Astrophysics Data System (ADS)

    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.

  18. Pyruvate Dehydrogenase Complex from Chloroplasts of Pisum sativum L 1

    PubMed Central

    Williams, Michael; Randall, Douglas D.

    1979-01-01

    Pyruvate dehydrogenase complex is associated with intact chloroplasts and mitochondria of 9-day-old Pisum sativum L. seedlings. The ratio of the mitochondrial complex to the chloroplast complex activities is about 3 to 1. Maximal rates observed for chloroplast pyruvate dehydrogenase complex activity ranged from 6 to 9 micromoles of NADH produced per milligram of chlorophyll per hour. Osmotic rupture of pea chloroplasts released 88% of the complex activity, indicating that chloroplast pyruvate dehydrogenase complex is a stromal complex. The pH optimum for chloroplast pyruvate dehydrogenase complex was between 7.8 and 8.2, whereas the mitochondrial pyruvate dehydrogenase complex had a pH optimum between 7.3 and 7.7. Chloroplast pyruvate dehydrogenase complex activity was specific for pyruvate, dependent upon coenzyme A and NAD and partially dependent upon Mg2+ and thiamine pyrophosphate. Chloroplast-associated pyruvate dehydrogenase complex provides a direct link between pyruvate metabolism and chloroplast fatty acid biosynthesis by providing the substrate, acetyl-CoA, necessary for membrane development in young plants. Images PMID:16661100

  19. Phylogeny of the genus Pistacia as determined from analysis of the chloroplast genome.

    PubMed

    Parfitt, D E; Badenes, M L

    1997-07-22

    Classification within the genus Pistacia has been based on leaf morphology and geographical distribution. Molecular genetic tools (PCR amplification followed by restriction analysis of a 3.2-kb region of variable chloroplast DNA, and restriction fragment length polymorphism analysis of the Pistacia cpDNA with tobacco chloroplast DNA probes) provided a new set of variables to study the phylogenetic relationships of 10 Pistacia species. Both parsimony and cluster analyses were used to divide the genus into two major groups. P. vera was determined to be the least derived species. P. weinmannifolia, an Asian species, is most closely related to P. texana and P. mexicana, New World species. These three species share a common origin, suggesting that a common ancestor of P. texana and P. mexicana originated in Asia. P. integerrima and P. chinensis were shown to be distinct whereas the pairs of species were monophyletic within each of two tertiary groups, P. vera:P. khinjuk and P. mexicana:P. texana. An evolutionary trend from large to small nuts and leaves with few, large leaflets to many, small leaflets was supported. The genus Pistacia was shown to have a low chloroplast DNA mutation rate: 0.05-0.16 times that expected of annual plants. PMID:9223300

  20. Analyses of the Complete Genome and Gene Expression of Chloroplast of Sweet Potato [Ipomoea batata

    PubMed Central

    Yan, Lang; Lai, Xianjun; Li, Xuedan; Wei, Changhe; Tan, Xuemei; Zhang, Yizheng

    2015-01-01

    Sweet potato [Ipomoea batatas (L.) Lam] ranks among the top seven most important food crops cultivated worldwide and is hexaploid plant (2n=6x=90) in the Convolvulaceae family with a genome size between 2,200 to 3,000 Mb. The genomic resources for this crop are deficient due to its complicated genetic structure. Here, we report the complete nucleotide sequence of the chloroplast (cp) genome of sweet potato, which is a circular molecule of 161,303 bp in the typical quadripartite structure with large (LSC) and small (SSC) single-copy regions separated by a pair of inverted repeats (IRs). The chloroplast DNA contains a total of 145 genes, including 94 protein-encoding genes of which there are 72 single-copy and 11 double-copy genes. The organization and structure of the chloroplast genome (gene content and order, IR expansion/contraction, random repeating sequences, structural rearrangement) of sweet potato were compared with those of Ipomoea (L.) species and some basal important angiosperms, respectively. Some boundary gene-flow and gene gain-and-loss events were identified at intra- and inter-species levels. In addition, by comparing with the transcriptome sequences of sweet potato, the RNA editing events and differential expressions of the chloroplast functional-genes were detected. Moreover, phylogenetic analysis was conducted based on 77 protein-coding genes from 33 taxa and the result may contribute to a better understanding of the evolution progress of the genus Ipomoea (L.), including phylogenetic relationships, intraspecific differentiation and interspecific introgression. PMID:25874767

  1. Analyses of the complete genome and gene expression of chloroplast of sweet potato [Ipomoea batata].

    PubMed

    Yan, Lang; Lai, Xianjun; Li, Xuedan; Wei, Changhe; Tan, Xuemei; Zhang, Yizheng

    2015-01-01

    Sweet potato [Ipomoea batatas (L.) Lam] ranks among the top seven most important food crops cultivated worldwide and is hexaploid plant (2n=6x=90) in the Convolvulaceae family with a genome size between 2,200 to 3,000 Mb. The genomic resources for this crop are deficient due to its complicated genetic structure. Here, we report the complete nucleotide sequence of the chloroplast (cp) genome of sweet potato, which is a circular molecule of 161,303 bp in the typical quadripartite structure with large (LSC) and small (SSC) single-copy regions separated by a pair of inverted repeats (IRs). The chloroplast DNA contains a total of 145 genes, including 94 protein-encoding genes of which there are 72 single-copy and 11 double-copy genes. The organization and structure of the chloroplast genome (gene content and order, IR expansion/contraction, random repeating sequences, structural rearrangement) of sweet potato were compared with those of Ipomoea (L.) species and some basal important angiosperms, respectively. Some boundary gene-flow and gene gain-and-loss events were identified at intra- and inter-species levels. In addition, by comparing with the transcriptome sequences of sweet potato, the RNA editing events and differential expressions of the chloroplast functional-genes were detected. Moreover, phylogenetic analysis was conducted based on 77 protein-coding genes from 33 taxa and the result may contribute to a better understanding of the evolution progress of the genus Ipomoea (L.), including phylogenetic relationships, intraspecific differentiation and interspecific introgression. PMID:25874767

  2. Field production and functional evaluation of chloroplast-derived interferon-α2b

    PubMed Central

    Arlen, Philip A.; Falconer, Regina; Cherukumilli, Sri; Cole, Amy; Cole, Alexander M.; Oishi, Karen K.; Daniell, Henry

    2008-01-01

    Summary Type I interferons (IFNs) inhibit viral replication and cell growth and enhance the immune response, and therefore have many clinical applications. IFN-α2b ranks third in world market use for a biopharmaceutical, behind only insulin and erythropoietin. The average annual cost of IFN-α2b for the treatment of hepatitis C infection is $26 000, and is therefore unavailable to the majority of patients in developing countries. Therefore, we expressed IFN-α2b in tobacco chloroplasts, and transgenic lines were grown in the field after obtaining United States Department of Agriculture Animal and Plant Health Inspection Service (USDA-APHIS) approval. Stable, site-specific integration of transgenes into chloroplast genomes and homoplasmy through several generations were confirmed. IFN-α2b levels reached up to 20% of total soluble protein, or 3 mg per gram of leaf (fresh weight). Transgenic IFN-α2b had similar in vitro biological activity to commercially produced PEG-Intron™ when tested for its ability to protect cells against cytopathic viral replication in the vesicular stomatitis virus cytopathic effect (VSV CPE) assay and to inhibit early-stage human immunodeficiency virus (HIV) infection. The antitumour and immunomodulating properties of IFN-α2b were also seen in vivo . Chloroplast-derived IFN-α2b increased the expression of major histocompatibility complex class I (MHC I) on splenocytes and the total number of natural killer (NK) cells. Finally, IFN-α2b purified from chloroplast transgenic lines (cpIFN-α2b) protected mice from a highly metastatic tumour line. This demonstration of high levels of expression of IFN-α2b, transgene containment and biological activity akin to that of commercial preparations of IFN-α2b facilitated the first field production of a plant-derived human blood protein, a critical step towards human clinical trials and commercialization. PMID:17490449

  3. Computational methods and software for differential equations and their applications. Annual report, November 16, 1984-November 15, 1985

    SciTech Connect

    Gear, C.W.

    1985-01-01

    This report summarizes the progress on the project over the last twelve months. Work has continued in two areas: numerical techniques for differential equations and software tools for scientific problem solving. Within these areas we have been working on the following: (1) Syntax-directed editors for mathematical software, (2) Polylith: a system for module interconnection, (3) mathematical expression input and editing, (4) a tool for examining fill-in in Gauss elimination, (5) a theoretical examination of fill-in, (6) a technique for determining the structural index of a differential-algebraic system, (7) invariant-conserving methods for differential equations, (9) iterative methods for stiff equations, (10) singular two-point boundary value problems, (11) initial value problems in Banach spaces, (12) computational error estimates for stiff ODEs, (13); multigrid solution of collocation equations, and (14) parallel techniques for ordinary differential equations.

  4. Ectopic Overexpression of The Transcription Factor OsGLK1 Induces Chloroplast Development in Non-Green Rice Cells

    PubMed Central

    Nakamura, Hidemitsu; Muramatsu, Masayuki; Hakata, Makoto; Ueno, Osamu; Nagamura, Yoshiaki; Hirochika, Hirohiko; Takano, Makoto; Ichikawa, Hiroaki

    2009-01-01

    For systematic and genome-wide analyses of rice gene functions, we took advantage of the full-length cDNA overexpresser (FOX) gene-hunting system and generated >12 000 independent FOX-rice lines from >25 000 rice calli treated with the rice-FOX Agrobacterium library. We found two FOX-rice lines generating green calli on a callus-inducing medium containing 2,4-D, on which wild-type rice calli became ivory yellow. In both lines, OsGLK1 cDNA encoding a GARP transcription factor was ectopically overexpressed. Using rice expression-microarray and northern blot analyses, we found that a large number of nucleus-encoded genes involved in chloroplast functions were highly expressed and transcripts of plastid-encoded genes, psaA, psbA and rbcL, increased in the OsGLK1-FOX calli. Transmission electron microscopy showed the existence of differentiated chloroplasts with grana stacks in OsGLK1-FOX calli cells. However, in darkness, OsGLK1-FOX calli did not show a green color or develop grana stacks. Furthermore, we found developed chloroplasts in vascular bundle and bundle sheath cells of coleoptiles and leaves from OsGLK1-FOX seedlings. The OsGLK1-FOX calli exhibited high photosynthetic activity and were able to grow on sucrose-depleted media, indicating that developed chloroplasts in OsGLK1-FOX rice calli are functional and active. We also observed that the endogenous OsGLK1 mRNA level increased synchronously with the greening of wild-type calli after transfer to plantlet regeneration medium. These results strongly suggest that OsGLK1 regulates chloroplast development under the control of light and phytohormones, and that it is a key regulator of chloroplast development. PMID:19808806

  5. KNOX genes influence a gradient of fruit chloroplast development through regulation of GOLDEN2-LIKE expression in tomato.

    PubMed

    Nadakuduti, Satya Swathi; Holdsworth, William L; Klein, Chelsey L; Barry, Cornelius S

    2014-06-01

    The chlorophyll content of unripe fleshy fruits is positively correlated with the nutrient content and flavor of ripe fruit. In tomato (Solanum lycopersicum) fruit, the uniform ripening (u) locus, which encodes a GOLDEN 2-LIKE transcription factor (SlGLK2), influences a gradient of chloroplast development that extends from the stem end of the fruit surrounding the calyx to the base of the fruit. With the exception of the u locus, the factors that influence the formation of this developmental gradient are unknown. In this study, characterization and positional cloning of the uniform gray-green (ug) locus of tomato reveals a thus far unknown role for the Class I KNOTTED1-LIKE HOMEOBOX (KNOX) gene, TKN4, in specifying the formation of this chloroplast gradient. The involvement of KNOX in fruit chloroplast development was confirmed through characterization of the Curl (Cu) mutant, a dominant gain-of-function mutation of TKN2, which displays ectopic fruit chloroplast development that resembles SlGLK2 over-expression. TKN2 and TKN4 act upstream of SlGLK2 and the related gene ARABIDOPSIS PSEUDO RESPONSE REGULATOR 2-LIKE (SlAPRR2-LIKE) to establish their latitudinal gradient of expression across developing fruit that leads to a gradient of chloroplast development. Class I KNOX genes typically influence plant morphology through maintenance of meristem activity, but this study identifies a role for TKN2 and TKN4 in specifically influencing chloroplast development in fruit but not leaves, suggesting that this fundamental process is differentially regulated in these two organs. PMID:24689783

  6. Why have chloroplasts developed a unique motility system?

    PubMed Central

    Suetsugu, Noriyuki; Dolja, Valerian V

    2010-01-01

    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. PMID:20855973

  7. Chloroplast Proteases: Updates on Proteolysis within and across Suborganellar Compartments.

    PubMed

    Nishimura, Kenji; Kato, Yusuke; Sakamoto, Wataru

    2016-08-01

    Chloroplasts originated from the endosymbiosis of ancestral cyanobacteria and maintain transcription and translation machineries for around 100 proteins. Most endosymbiont genes, however, have been transferred to the host nucleus, and the majority of the chloroplast proteome is composed of nucleus-encoded proteins that are biosynthesized in the cytosol and then imported into chloroplasts. How chloroplasts and the nucleus communicate to control the plastid proteome remains an important question. Protein-degrading machineries play key roles in chloroplast proteome biogenesis, remodeling, and maintenance. Research in the past few decades has revealed more than 20 chloroplast proteases, which are localized to specific suborganellar locations. In particular, two energy-dependent processive proteases of bacterial origin, Clp and FtsH, are central to protein homeostasis. Processing endopeptidases such as stromal processing peptidase and thylakoidal processing peptidase are involved in the maturation of precursor proteins imported into chloroplasts by cleaving off the amino-terminal transit peptides. Presequence peptidases and organellar oligopeptidase subsequently degrade the cleaved targeting peptides. Recent findings have indicated that not only intraplastidic but also extraplastidic processive protein-degrading systems participate in the regulation and quality control of protein translocation across the envelopes. In this review, we summarize current knowledge of the major chloroplast proteases in terms of type, suborganellar localization, and diversification. We present details of these degradation processes as case studies according to suborganellar compartment (envelope, stroma, and thylakoids). Key questions and future directions in this field are discussed. PMID:27288365

  8. Tools for regulated gene expression in the chloroplast of Chlamydomonas.

    PubMed

    Rochaix, Jean-David; Surzycki, Raymond; Ramundo, Silvia

    2014-01-01

    The green unicellular alga Chlamydomonas reinhardtii has emerged as a very attractive model system for chloroplast genetic engineering. Algae can be transformed readily at the chloroplast level through bombardment of cells with a gene gun, and transformants can be selected using antibiotic resistance or phototrophic growth. An inducible chloroplast gene expression system could be very useful for several reasons. First, it could be used to elucidate the function of essential chloroplast genes required for cell growth and survival. Second, it could be very helpful for expressing proteins which are toxic to the algal cells. Third, it would allow for the reversible depletion of photosynthetic complexes thus making it possible to study their biogenesis in a controlled fashion. Fourth, it opens promising possibilities for hydrogen production in Chlamydomonas. Here we describe an inducible/repressible chloroplast gene expression system in Chlamydomonas in which the copper-regulated Cyc6 promoter drives the expression of the nuclear Nac2 gene encoding a protein which is targeted to the chloroplast where it acts specifically on the chloroplast psbD 5'-untranslated region and is required for the stable accumulation of the psbD mRNA and photosystem II. The system can be used for any chloroplast gene or transgene by placing it under the control of the psbD 5'-untranslated region. PMID:24599871

  9. Protein Methylation in Pea Chloroplasts 1

    PubMed Central

    Niemi, Kevin J.; Adler, Julius; Selman, Bruce R.

    1990-01-01

    The methylation of chloroplast proteins has been investigated by incubating intact pea (Pisum sativum) chloroplasts with [3H-methyl]-S-adenosylmethionine. Incubation in the light increases the amount of methylation in both the thylakoid and stromal fractions. Numerous thylakoid proteins serve as substrates for the methyltransfer reactions. Three of these thylakoid proteins are methylated to a significantly greater extent in the light than in the dark. One is a polypeptide with a molecular mass of 64 kD, a second has an Mr of 48 kD, and the third has a molecular mass of less than 10 kD. The primary stromal polypeptide methylated is the large subunit of ribulose bisphosphate carboxylase/oxygenase. One other stromal polypeptide, having a molecular mass of 24 kD, is also methylated much more in the light than in the dark. Two distinct types of protein methylation occur. One methyl-linkage is stable to basic conditions whereas a second type is base labile. The base-stable linkage is indicative of N-methylation of amino acid residues while base-lability is suggestive of carboxymethylation of amino acid residues. Labeling in the light increases the percentage of methylation that is base labile in the thylakoid fraction while no difference is observed in the amount of base-labile methylations in light-labeled and dark-labeled stromal proteins. Also suggestive of carboxymethylation is the detection of volatile [3H]methyl radioactivity which increases during the labeling period and is greater in chloroplasts labeled in the light as opposed to being labeled in the dark; this implies in vivo turnover of the [3H]methyl group. Images Figure 1 PMID:16667584

  10. Expression of eukaryotic polypeptides in chloroplasts

    DOEpatents

    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.

  11. The complete chloroplast genome of the Dendrobium strongylanthum (Orchidaceae: Epidendroideae).

    PubMed

    Li, Jing; Chen, Chen; Wang, Zhe-Zhi

    2016-07-01

    Complete chloroplast genome sequence is very useful for studying the phylogenetic and evolution of species. In this study, the complete chloroplast genome of Dendrobium strongylanthum was constructed from whole-genome Illumina sequencing data. The chloroplast genome is 153 058 bp in length with 37.6% GC content and consists of two inverted repeats (IRs) of 26 316 bp. The IR regions are separated by large single-copy region (LSC, 85 836 bp) and small single-copy (SSC, 14 590 bp) region. A total of 130 chloroplast genes were successfully annotated, including 84 protein coding genes, 38 tRNA genes, and eight rRNA genes. Phylogenetic analyses showed that the chloroplast genome of Dendrobium strongylanthum is related to that of the Dendrobium officinal. PMID:26153739

  12. Non-contact intracellular binding of chloroplasts in vivo

    NASA Astrophysics Data System (ADS)

    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.

  13. The complete chloroplast genome sequence of Panax quinquefolius (L.).

    PubMed

    Kim, Kyunghee; Lee, Sang-Choon; Lee, Junki; Kim, Nam-Hoon; Jang, Woojong; Yang, Tae-Jin

    2016-07-01

    The complete chloroplast genome sequence of Panax quinquefolius, an important medicinal herb, was generated by de novo assembly with low-coverage whole-genome sequence data and manual correction. A circular 156 088-bp chloroplast genome showed typical chloroplast genome structure comprising a large single copy region of 86 095 bp, a small single copy region of 17 993 bp, and a pair of inverted repeats of 26 000 bp. The chloroplast genome had 87 protein-coding genes, 37 tRNA genes, and eight rRNA genes. Phylogenetic analysis with the chloroplast genome revealed that P. quinquefolius is much closer to P. ginseng than P. notoginseng. PMID:26162051

  14. Comparative proteomic analysis of amaranth mesophyll and bundle sheath chloroplasts and their adaptation to salt stress.

    PubMed

    Joaquín-Ramos, Ahuitzolt; Huerta-Ocampo, José Á; Barrera-Pacheco, Alberto; De León-Rodríguez, Antonio; Baginsky, Sacha; Barba de la Rosa, Ana P

    2014-09-15

    The effect of salt stress was analyzed in chloroplasts of Amaranthus cruentus var. Amaranteca, a plant NAD-malic enzyme (NAD-ME) type. Morphology of chloroplasts from bundle sheath (BSC) and mesophyll (MC) was observed by transmission electron microscopy (TEM). BSC and MC from control plants showed similar morphology, however under stress, changes in BSC were observed. The presence of ribulose bisphosphate carboxylase/oxygenase (RuBisCO) was confirmed by immunohistochemical staining in both types of chloroplasts. Proteomic profiles of thylakoid protein complexes from BSC and MC, and their changes induced by salt stress were analyzed by blue-native polyacrylamide gel electrophoresis followed by SDS-PAGE (2-D BN/SDS-PAGE). Differentially accumulated protein spots were analyzed by LC-MS/MS. Although A. cruentus photosynthetic tissue showed the Kranz anatomy, the thylakoid proteins showed some differences at photosystem structure level. Our results suggest that A. cruentus var. Amaranteca could be better classified as a C3-C4 photosynthetic plant. PMID:25046763

  15. Phylogenetic affinities of the grasses to other monocots as revealed by molecular analysis of chloroplast DNA.

    PubMed

    Katayama, H; Ogihara, Y

    1996-05-01

    The distribution of structural alterations of the chloroplast genome found in grass chloroplast (cp) DNA in comparison with that of tobacco was systematically surveyed in the cpDNAs of monocots. Southern hybridization and/or PCR analyses for the detection of (1) three inversions in the large single-copy region, (2) loss of an intron in the rpoC1 gene, (3) an extra-sequence insertion in the rpoC2 gene, (4) the deletion of ORF2280, (5) rearrangements of the accD (ORF512) gene, and (6) non-reciprocal translocation of the rpl23 gene, were carried out on cpDNAs isolated from 58 species, 22 families, and 11 orders, which covered almost all families of monocots. These structural alterations of cpDNA mostly occurred at the family level. However, only part of the Restionaceae possessed the inversion that characterizes the lineage of grass differentiation. The order of mutational events made it possible to reconstruct grass phylogeny in monocots. Since no variations in structural alterations of the cpDNA were found among the Poaceae, grass plants were inferred to have originated from an ancestor harboring these structural alterations of the chloroplast genome. These phylogenetic relationships were supported by the sequence data of rbcL. PMID:8662197

  16. Chloroplast targeting of FtsHprotease is essential for chloroplast development and thylakoid stability at elevated temperatures in plants

    Technology Transfer Automated Retrieval System (TEKTRAN)

    AtFtsH11 is a chloroplast and mitochondria dual targeted metalloprotease, identified as essential for Arabidopsis plant to survive at moderate high temperatures at all developmental stages. Our study showed that FtsH11 plays critical roles in both the early stages of chloroplast biogenesis and main...

  17. The Chloroplast Genome of Pellia endiviifolia: Gene Content, RNA-Editing Pattern, and the Origin of Chloroplast Editing

    PubMed Central

    Grosche, Christopher; Funk, Helena T.; Maier, Uwe G.; Zauner, Stefan

    2012-01-01

    RNA editing is a post-transcriptional process that can act upon transcripts from mitochondrial, nuclear, and chloroplast genomes. In chloroplasts, single-nucleotide conversions in mRNAs via RNA editing occur at different frequencies across the plant kingdom. These range from several hundred edited sites in some mosses and ferns to lower frequencies in seed plants and the complete lack of RNA editing in the liverwort Marchantia polymorpha. Here, we report the sequence and edited sites of the chloroplast genome from the liverwort Pellia endiviifolia. The type and frequency of chloroplast RNA editing display a pattern highly similar to that in seed plants. Analyses of the C to U conversions and the genomic context in which the editing sites are embedded provide evidence in favor of the hypothesis that chloroplast RNA editing evolved to compensate mutations in the first land plants. PMID:23221608

  18. Ferredoxin-linked chloroplast enzymes. Progress report

    SciTech Connect

    1993-12-31

    This report summarizes research on ferredoxin:NADP{sup +} oxidoreductase and ferredoxin:thioredoxin reductase. One of the primary goals of the original proposal was to map the ferredoxin-binding sites on three soluble enzymes that are located in spinach chloroplasts and utilize ferredoxin as an electron donor:Ferredoxin:NADP{sup +} oxidoreductase (FNR); ferredoxin:thioredoxin reductase (FTR) and glutamate synthase. As the availability of amino acid sequences for the enzymes are important in such studies, it was proposed that the amino acid sequence of glutamate synthase be determined. The amino acid sequences of FNR, FTR and ferredoxin are already known. An aim related to elucidating the binding sites on these enzymes for ferredoxin was to determine whether there is a common site on ferredoxin involved in binding to all of these ferredoxin-dependent chloroplast enzymes and, if so, to map it. One additional aim was to characterize thioredoxin binding by FTR and determine whether the same site on FTR is involved in binding both ferredoxin and thioredoxin. Considerable progress has been made on most of these original projects, although work conducted on FTR is still in its preliminary stages.

  19. The complete chloroplast genome of Capsella rubella.

    PubMed

    Wu, Zhiqiang

    2016-07-01

    The whole nucleotide sequence of the chloroplast genome from Capsella rubella is determined in this study using short Illumina sequence data from public database. The circular double-stranded DNA, which consists of 154,601 base pairs (bp) in size, contains a pair of inverted repeats (IRa and IRb) of 26,462 bp each, which are separated by a small and large single-copies (SSC and LSC) of 17,855 and 83,822 bp, respectively. The overall GC content of the chloroplast genome is 36.54% and the GC contents of LSC, IRs and SSC are 34.33%, 42.38% and 29.61% separately. One hundred and twelve unique genes were annotated, including 78 protein-coding genes, 30 tRNA genes and 4 rRNA genes. Among these, 16 are duplicated in the inverted repeat regions, 15 genes contained 1 intron, and 3 genes (rps12, clpP and ycf3) comprised of 2 introns. Two protein genes (rps19 and ycf1) span in the boundaries of LSC-IR and IR-SSC to produce two partial pseudogenes. PMID:26024136

  20. Programmed cell death in plants: A chloroplastic connection

    PubMed Central

    Ambastha, Vivek; Tripathy, Baishnab C; Tiwari, Budhi Sagar

    2015-01-01

    Programmed cell death (PCD) is an integral cellular program by which targeted cells culminate to demise under certain developmental and pathological conditions. It is essential for controlling cell number, removing unwanted diseased or damaged cells and maintaining the cellular homeostasis. The details of PCD process has been very well elucidated and characterized in animals but similar understanding of the process in plants has not been achieved rather the field is still in its infancy that sees some sporadic reports every now and then. The plants have 2 energy generating sub-cellular organelles- mitochondria and chloroplasts unlike animals that just have mitochondria. The presence of chloroplast as an additional energy transducing and ROS generating compartment in a plant cell inclines to advocate the involvement of chloroplasts in PCD execution process. As chloroplasts are supposed to be progenies of unicellular photosynthetic organisms that evolved as a result of endosymbiosis, the possibility of retaining some of the components involved in bacterial PCD by chloroplasts cannot be ruled out. Despite several excellent reviews on PCD in plants, there is a void on an update of information at a place on the regulation of PCD by chloroplast. This review has been written to provide an update on the information supporting the involvement of chloroplast in PCD process and the possible future course of the field. PMID:25760871

  1. Mining the soluble chloroplast proteome by affinity chromatography.

    PubMed

    Bayer, Roman G; Stael, Simon; Csaszar, Edina; Teige, Markus

    2011-04-01

    Chloroplasts are fundamental organelles enabling plant photoautotrophy. Besides their outstanding physiological role in fixation of atmospheric CO(2), they harbor many important metabolic processes such as biosynthesis of amino acids, vitamins or hormones. Technical advances in MS allowed the recent identification of most chloroplast proteins. However, for a deeper understanding of chloroplast function it is important to obtain a complete list of constituents, which is so far limited by the detection of low-abundant proteins. Therefore, we developed a two-step strategy for the enrichment of low-abundant soluble chloroplast proteins from Pisum sativum and their subsequent identification by MS. First, chloroplast protein extracts were depleted from the most abundant protein ribulose-1,5-bisphosphate carboxylase/oxygenase by SEC or heating. Further purification was carried out by affinity chromatography, using ligands specific for ATP- or metal-binding proteins. By these means, we were able to identify a total of 448 proteins including 43 putative novel chloroplast proteins. Additionally, the chloroplast localization of 13 selected proteins was confirmed using yellow fluorescent protein fusion analyses. The selected proteins included a phosphoglycerate mutase, a cysteine protease, a putative protein kinase and an EF-hand containing substrate carrier protein, which are expected to exhibit important metabolic or regulatory functions. PMID:21365755

  2. Chloroplast-like transfer RNA genes expressed in wheat mitochondria.

    PubMed Central

    Joyce, P B; Gray, M W

    1989-01-01

    In the course of a systematic survey of wheat mitochondrial tRNA genes, we have sequenced chloroplast-like serine (trnS-GGA), phenylalanine (trnF-GAA) and cysteine (trnC-GCA) tRNA genes and their flanking regions. These genes are remnants of 'promiscuous' chloroplast DNA that has been incorporated into wheat mtDNA in the course of its evolution. Each gene differs by one or a few nucleotides from the authentic chloroplast homolog previously characterized in wheat or other plants, and each could potentially encode a functional tRNA whose secondary structure shows no deviations from the generalized model. To determine whether these chloroplast-like tRNA genes are actually expressed, wheat mitochondrial tRNAs were resolved by a series of polyacrylamide gel electrophoreses, after being specifically end-labeled in vitro by 3'-CCA addition mediated by wheat tRNA nucleotidyltransferase. Subsequent direct RNA sequence analysis identified prominent tRNA species corresponding to the mitochondrial and not the chloroplast trnS, trnF and trnC genes. This analysis also revealed chloroplast-like elongator methionine, asparagine and tryptophan tRNAs. Our results suggest that at least some chloroplast-like tRNA genes in wheat mtDNA are transcribed, with transcripts undergoing processing, post-transcriptional modification and 3'-CCA addition, to produce mature tRNAs that may participate in mitochondrial protein synthesis. Images PMID:2762145

  3. Transposon-induced nuclear mutations that alter chloroplast gene expression

    SciTech Connect

    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.

  4. Mining the soluble chloroplast proteome by affinity chromatography

    PubMed Central

    Bayer, Roman G; Stael, Simon; Csaszar, Edina; Teige, Markus

    2011-01-01

    Chloroplasts are fundamental organelles enabling plant photoautotrophy. Besides their outstanding physiological role in fixation of atmospheric CO2, they harbor many important metabolic processes such as biosynthesis of amino acids, vitamins or hormones. Technical advances in MS allowed the recent identification of most chloroplast proteins. However, for a deeper understanding of chloroplast function it is important to obtain a complete list of constituents, which is so far limited by the detection of low-abundant proteins. Therefore, we developed a two-step strategy for the enrichment of low-abundant soluble chloroplast proteins from Pisum sativum and their subsequent identification by MS. First, chloroplast protein extracts were depleted from the most abundant protein ribulose-1,5-bisphosphate carboxylase/oxygenase by SEC or heating. Further purification was carried out by affinity chromatography, using ligands specific for ATP- or metal-binding proteins. By these means, we were able to identify a total of 448 proteins including 43 putative novel chloroplast proteins. Additionally, the chloroplast localization of 13 selected proteins was confirmed using yellow fluorescent protein fusion analyses. The selected proteins included a phosphoglycerate mutase, a cysteine protease, a putative protein kinase and an EF-hand containing substrate carrier protein, which are expected to exhibit important metabolic or regulatory functions. PMID:21365755

  5. Chloroplast diversity in a putative hybrid swarm of Ponderosae (Pinaceae).

    PubMed

    Epperson, Bryan K; Telewski, Frank W; Willyard, Ann

    2009-03-01

    The Ponderosae subsection of the genus Pinus contains numerous taxa in disjunct mountain ranges of southern Arizona and New Mexico, differing for several leaf and cone traits, key among which is the number of leaf needles per fascicle. Trees with three needles are often found together with trees having five needles and mixed numbers. One taxonomic hypothesis is that there are swarms of hybrids between P. ponderosa and P. arizonica. A second hypothesis is that there are spatial mixtures of two separate taxa, five-needle P. arizonica and a "taxon X" containing three needle and mixed needle trees. We genotyped chloroplasts in one putative hybrid swarm on Mt. Lemmon using microsatellite markers and show that cpDNA is almost completely differentiated between two separate morphotypes corresponding to P. arizonica and "taxon X." Little if any introgression has occurred on Mt. Lemmon, and the simplest explanation is that little or no effective hybridization has occurred. Further results indicate that not only is taxon X not of hybrid origin, it is more closely related to nonregional Ponderosae other than P. ponderosa and P. arizonica. The results further suggest that other putative hybrid swarms in the region are also spatial mixtures of distinct taxa. PMID:21628225

  6. Production of therapeutic proteins in the chloroplast of Chlamydomonas reinhardtii

    PubMed Central

    2014-01-01

    Chloroplast transformation in the photosynthetic alga Chlamydomonas reinhardtii has been used to explore the potential to use it as an inexpensive and easily scalable system for the production of therapeutic recombinant proteins. Diverse proteins, such as bacterial and viral antigens, antibodies and, immunotoxins have been successfully expressed in the chloroplast using endogenous and chimeric promoter sequences. In some cases, proteins have accumulated to high level, demonstrating that this technology could compete with current production platforms. This review focuses on the works that have engineered the chloroplast of C. reinhardtii with the aim of producing recombinant proteins intended for therapeutical use in humans or animals. PMID:25136510

  7. Production of therapeutic proteins in the chloroplast of Chlamydomonas reinhardtii.

    PubMed

    Almaraz-Delgado, Alma Lorena; Flores-Uribe, José; Pérez-España, Víctor Hugo; Salgado-Manjarrez, Edgar; Badillo-Corona, Jesús Agustín

    2014-01-01

    Chloroplast transformation in the photosynthetic alga Chlamydomonas reinhardtii has been used to explore the potential to use it as an inexpensive and easily scalable system for the production of therapeutic recombinant proteins. Diverse proteins, such as bacterial and viral antigens, antibodies and, immunotoxins have been successfully expressed in the chloroplast using endogenous and chimeric promoter sequences. In some cases, proteins have accumulated to high level, demonstrating that this technology could compete with current production platforms. This review focuses on the works that have engineered the chloroplast of C. reinhardtii with the aim of producing recombinant proteins intended for therapeutical use in humans or animals. PMID:25136510

  8. Chloroplast Phylogenomic Inference of Green Algae Relationships

    PubMed Central

    Sun, Linhua; Fang, Ling; Zhang, Zhenhua; Chang, Xin; Penny, David; Zhong, Bojian

    2016-01-01

    The green algal phylum Chlorophyta has six diverse classes, but the phylogenetic relationship of the classes within Chlorophyta remains uncertain. In order to better understand the ancient Chlorophyta evolution, we have applied a site pattern sorting method to study compositional heterogeneity and the model fit in the green algal chloroplast genomic data. We show that the fastest-evolving sites are significantly correlated with among-site compositional heterogeneity, and these sites have a much poorer fit to the evolutionary model. Our phylogenomic analyses suggest that the class Chlorophyceae is a monophyletic group, and the classes Ulvophyceae, Trebouxiophyceae and Prasinophyceae are non-monophyletic groups. Our proposed phylogenetic tree of Chlorophyta will offer new insights to investigate ancient green algae evolution, and our analytical framework will provide a useful approach for evaluating and mitigating the potential errors of phylogenomic inferences. PMID:26846729

  9. Dynamics of chloroplast genomes in green plants.

    PubMed

    Xu, Jian-Hong; Liu, Qiuxiang; Hu, Wangxiong; Wang, Tingzhang; Xue, Qingzhong; Messing, Joachim

    2015-10-01

    Chloroplasts are essential organelles, in which genes have widely been used in the phylogenetic analysis of green plants. Here, we took advantage of the breadth of plastid genomes (cpDNAs) sequenced species to investigate their dynamic changes. Our study showed that gene rearrangements occurred more frequently in the cpDNAs of green algae than in land plants. Phylogenetic trees were generated using 55 conserved protein-coding genes including 33 genes for photosynthesis, 16 ribosomal protein genes and 6 other genes, which supported the monophyletic evolution of vascular plants, land plants, seed plants, and angiosperms. Moreover, we could show that seed plants were more closely related to bryophytes rather than pteridophytes. Furthermore, the substitution rate for cpDNA genes was calculated to be 3.3×10(-10), which was almost 10 times lower than genes of nuclear genomes, probably because of the plastid homologous recombination machinery. PMID:26206079

  10. Chloroplast phylogeny indicates that bryophytes are monophyletic.

    PubMed

    Nishiyama, Tomoaki; Wolf, Paul G; Kugita, Masanori; Sinclair, Robert B; Sugita, Mamoru; Sugiura, Chika; Wakasugi, Tatsuya; Yamada, Kyoji; Yoshinaga, Koichi; Yamaguchi, Kazuo; Ueda, Kunihiko; Hasebe, Mitsuyasu

    2004-10-01

    Opinions on the basal relationship of land plants vary considerably and no phylogenetic tree with significant statistical support has been obtained. Here, we report phylogenetic analyses using 51 genes from the entire chloroplast genome sequences of 20 representative green plant species. The analyses, using translated amino acid sequences, indicated that extant bryophytes (mosses, liverworts, and hornworts) form a monophyletic group with high statistical confidence and that extant bryophytes are likely sisters to extant vascular plants, although the support for monophyletic vascular plants was not strong. Analyses at the nucleotide level could not resolve the basal relationship with statistical confidence. Bryophyte monophyly inferred using amino acid sequences has a good statistical foundation and is not rejected statistically by other data sets. We propose bryophyte monophyly as the currently best hypothesis. PMID:15240838

  11. Origins of prokaryotes, eukaryotes, mitochondria, and chloroplasts

    NASA Technical Reports Server (NTRS)

    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.

  12. Light Quality Effects on Corn Chloroplast Development

    PubMed Central

    Eskins, Kenneth; Duysen, Murray; Dybas, Linda; McCarthy, Susan

    1985-01-01

    Corn was grown under greenhouse and controlled light quality conditions incluing full spectrum, red (R), and far-red (FR) sources. Young leaf samples were analyzed for pigments, pigment-proteins, membrane polypeptides, and ultrastructure. Chloroplast development in full spectrum white light was similar to that found in R but different from that found in FR plus low R. Compared to greenhouse and R, FR plus low R (670-760) repressed the formation of photosystem I reaction center protein (CP1 + CP1a) and enhanced those of photosystem II (CPa) in both bundle sheath and mesophyll cells. Photosystem II polypeptides were present in both cell types, with the 46 and 34 kilodalton proteins predominant in mesophyll cells. Bundle sheath cells contained relatively more of the 51 kilodalton and less of the 46 kilodalton proteins. However, they also contained measurable amounts of ribulose bisphosphate carboxylase which may interfere with estimates of the 51 kilodalton protein. Images Fig. 4 PMID:16664023

  13. Photosynthetic light reactions: integral to chloroplast retrograde signalling.

    PubMed

    Gollan, Peter J; Tikkanen, Mikko; Aro, Eva-Mari

    2015-10-01

    Chloroplast retrograde signalling is ultimately dependent on the function of the photosynthetic light reactions and not only guides the acclimation of the photosynthetic apparatus to changing environmental and metabolic cues, but has a much wider influence on the growth and development of plants. New information generated during the past few years about regulation of photosynthetic light reactions and identification of the underlying regulatory proteins has paved the way towards better understanding of the signalling molecules produced in chloroplasts upon changes in the environment. Likewise, the availability of various mutants lacking regulatory functions has made it possible to address the role of excitation energy distribution and electron flow in the thylakoid membrane in inducing the retrograde signals from chloroplasts to the nucleus. Such signalling molecules also induce and interact with hormonal signalling cascades to provide comprehensive information from chloroplasts to the nucleus. PMID:26318477

  14. The complete chloroplast genome sequence of Zanthoxylum piperitum.

    PubMed

    Lee, Jonghoon; Lee, Hyeon Ju; Kim, Kyunghee; Lee, Sang-Choon; Sung, Sang Hyun; Yang, Tae-Jin

    2016-09-01

    The complete chloroplast genome sequence of Zanthoxylum piperitum, a plant species with useful aromatic oils in family Rutaceae, was generated in this study by de novo assembly with whole-genome sequence data. The chloroplast genome was 158 154 bp in length with a typical quadripartite structure containing a pair of inverted repeats of 27 644 bp, separated by large single copy and small single copy of 85 340 bp and 17 526 bp, respectively. The chloroplast genome harbored 112 genes consisting of 78 protein-coding genes 30 tRNA genes and 4 rRNA genes. Phylogenetic analysis of the complete chloroplast genome sequences with those of known relatives revealed that Z. piperitum is most closely related to the Citrus species. PMID:26260183

  15. Full transcription of the chloroplast genome in photosynthetic eukaryotes

    PubMed Central

    Shi, Chao; Wang, Shuo; Xia, En-Hua; Jiang, Jian-Jun; Zeng, Fan-Chun; Gao, Li-Zhi

    2016-01-01

    Prokaryotes possess a simple genome transcription system that is different from that of eukaryotes. In chloroplasts (plastids), it is believed that the prokaryotic gene transcription features govern genome transcription. However, the polycistronic operon transcription model cannot account for all the chloroplast genome (plastome) transcription products at whole-genome level, especially regarding various RNA isoforms. By systematically analyzing transcriptomes of plastids of algae and higher plants, and cyanobacteria, we find that the entire plastome is transcribed in photosynthetic green plants, and that this pattern originated from prokaryotic cyanobacteria — ancestor of the chloroplast genomes that diverged about 1 billion years ago. We propose a multiple arrangement transcription model that multiple transcription initiations and terminations combine haphazardly to accomplish the genome transcription followed by subsequent RNA processing events, which explains the full chloroplast genome transcription phenomenon and numerous functional and/or aberrant pre-RNAs. Our findings indicate a complex prokaryotic genome regulation when processing primary transcripts. PMID:27456469

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

    SciTech Connect

    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.

  17. Prenylquinone profiling in whole leaves and chloroplast subfractions.

    PubMed

    Kessler, Felix; Glauser, Gaetan

    2014-01-01

    Prenylquinones are indispensable molecules in plants and animals. In plants, phylloquinone (vitamin K) and plastoquinone are electron carriers during photosynthesis in chloroplasts, whereas tocopherol (vitamin E) functions as a lipid antioxidant. The biosynthetic pathways of the prenylquinones have been largely characterized but the mechanisms regulating their production and distribution in various subcompartments of the chloroplast are only starting to emerge. Research on chloroplast lipid droplets (plastoglobules) has unraveled a complex network of intersecting prenylquinone metabolic pathways that are providing unprecedented insight into the regulatory processes. In this chapter, we describe how to isolate chloroplast membrane fractions, in particular the plastoglobule lipid droplets, and how to profile the prenylquinones that are contained in these fractions. PMID:24777800

  18. Full transcription of the chloroplast genome in photosynthetic eukaryotes.

    PubMed

    Shi, Chao; Wang, Shuo; Xia, En-Hua; Jiang, Jian-Jun; Zeng, Fan-Chun; Gao, Li-Zhi

    2016-01-01

    Prokaryotes possess a simple genome transcription system that is different from that of eukaryotes. In chloroplasts (plastids), it is believed that the prokaryotic gene transcription features govern genome transcription. However, the polycistronic operon transcription model cannot account for all the chloroplast genome (plastome) transcription products at whole-genome level, especially regarding various RNA isoforms. By systematically analyzing transcriptomes of plastids of algae and higher plants, and cyanobacteria, we find that the entire plastome is transcribed in photosynthetic green plants, and that this pattern originated from prokaryotic cyanobacteria - ancestor of the chloroplast genomes that diverged about 1 billion years ago. We propose a multiple arrangement transcription model that multiple transcription initiations and terminations combine haphazardly to accomplish the genome transcription followed by subsequent RNA processing events, which explains the full chloroplast genome transcription phenomenon and numerous functional and/or aberrant pre-RNAs. Our findings indicate a complex prokaryotic genome regulation when processing primary transcripts. PMID:27456469

  19. Chloroplast genome variation in upland and lowland switchgrass

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Switchgrass (Panicum virgatum L.) exists at multiple ploidies and two phenotypically distinct ecotypes. To facilitate interploidal comparisons and to understand the extent of sequence variation within existing breeding pools, two complete switchgrass chloroplast genomes were sequenced from individu...

  20. Separation of Chloroplast Pigments Using Reverse Phase Chromatography.

    ERIC Educational Resources Information Center

    Reese, R. Neil

    1997-01-01

    Presents a protocol that uses reverse phase chromatography for the separation of chloroplast pigments. Provides a simple and relatively safe procedure for use in teaching laboratories. Discusses pigment extraction, chromatography, results, and advantages of the process. (JRH)

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

    PubMed Central

    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

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

    PubMed

    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

  3. Complete Chloroplast Genome of Tanaecium tetragonolobum: The First Bignoniaceae Plastome

    PubMed Central

    Nazareno, Alison Gonçalves; Carlsen, Monica; Lohmann, Lúcia Garcez

    2015-01-01

    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 plants is complex

  4. Sequence evidence for the symbiotic origins of chloroplasts and mitochondria

    NASA Technical Reports Server (NTRS)

    George, D. G.; Hunt, L. T.; Dayhoff, M. O.

    1983-01-01

    The origin of mitochondria and chloroplasts is investigated on the basis of prokaryotic and early-eukaryotic evolutionary trees derived from protein and nucleic-acid sequences by the method of Dayhoff (1979). Trees for bacterial ferrodoxins, 5S ribosomal RNA, c-type cytochromes, the lipid-binding subunit of ATPase, and dihydrofolate reductase are presented and discussed. Good agreement among the trees is found, and it is argued that the mitochondria and chloroplasts evolved by multiple symbiotic events.

  5. Export of Carbon from Chloroplasts at Night1

    PubMed Central

    Schleucher, Jürgen; Vanderveer, Peter J.; Sharkey, Thomas D.

    1998-01-01

    Hexose export from chloroplasts at night has been inferred in previous studies of mutant and transgenic plants. We have tested whether hexose export is the normal route of carbon export from chloroplasts at night. We used nuclear magnetic resonance to distinguish glucose (Glc) made from hexose export and Glc made from triose export. Glc synthesized in vitro from fructose-6-phosphate in the presence of deuterium-labeled water had deuterium incorporated at C-2, whereas synthesis from triose phosphates caused C-2 through C-5 to become deuterated. In both tomato (Lycopersicon esculentum L.) and bean (Phaseolus vulgaris L.), Glc from sucrose made at night in the presence of deuterium-enriched water was deuterated only in the C-2 position, indicating that >75% of carbon is exported as hexoses at night. In darkness the phosphate in the cytosol was 28 mm, whereas that in the chloroplasts was 5 mm, but hexose phosphates were 10-fold higher in the cytosol than in the chloroplasts. Therefore, hexose phosphates would not move out of chloroplasts without the input of energy. We conclude that most carbon leaves chloroplasts at night as Glc, maltose, or higher maltodextrins under normal conditions. PMID:9847119

  6. Evidence that sigma factors are components of chloroplast RNA polymerase.

    PubMed Central

    Troxler, R F; Zhang, F; Hu, J; Bogorad, L

    1994-01-01

    Plastid genes are transcribed by DNA-dependent RNA polymerase(s), which have been incompletely characterized and have been examined in a limited number of species. Plastid genomes contain rpoA, rpoB, rpoC1, and rpoC2 coding for alpha, beta, beta', and beta" RNA polymerase subunits that are homologous to the alpha, beta, and beta' subunits that constitute the core moiety of RNA polymerase in bacteria. However, genes with homology to sigma subunits in bacteria have not been found in plastid genomes. An antibody directed against the principal sigma subunit of RNA polymerase from the cyanobacterium Anabaena sp. PCC 7120 was used to probe western blots of purified chloroplast RNA polymerase from maize, rice, Chlamydomonas reinhardtii, and Cyanidium caldarium. Chloroplast RNA polymerase from maize and rice contained an immunoreactive 64-kD protein. Chloroplast RNA polymerase from C. reinhardtii contained immunoreactive 100- and 82-kD proteins, and chloroplast RNA polymerase from C. caldarium contained an immunoreactive 32-kD protein. The elution profile of enzyme activity of both algal chloroplast RNA polymerases coeluted from DEAE with the respective immunoreactive proteins, indicating that they are components of the enzyme. These results provide immunological evidence for sigma-like factors in chloroplast RNA polymerase in higher plants and algae. PMID:8159791

  7. Amino acid incorporation by ribosomes and polyribosomes from wheat chloroplasts.

    PubMed

    Hadziyev, D; Zalik, S

    1970-01-01

    Sucrose-gradient and analytical ultracentrifugation showed that chloroplast polyribosomes from 4-day-old seedlings had mono-, di-, tri-, tetra- and traces of penta-ribosomes, in contrast with those from 7-day-old seedlings in which only the mono-, di- and traces of tri-ribosomes were present. Without Mg(2+) the polyribosomes dissociated into ribosomal subunits. The rate of l-[U-(14)C]phenylalanine incorporation was threefold greater for preparations from 4- than from 7-day-old seedlings. Incorporation by the latter was stimulated by polyuridylic acid. The rates of incorporation were similar whether the reaction mixture contained chloroplast or wheat-germ transfer RNA and amino acid synthetases purified on methylated albumin-on-kieselguhr and Sephadex G-75 columns respectively. The cofactor requirement was the same as for isolated intact chloroplasts. Osmotic rupture of chloroplasts with and without Triton X-100 revealed the presence of free and bound ribosomes. Free single ribosomes isolated by osmotic shrinkage or prepared by pancreatic ribonuclease digestion of chloroplast polyribosomes had negligible incorporation activity. This activity was increased by washing or by polyuridylic acid, but was still only a fraction of that given by polyribosomes. A comparison of incorporation activity of chloroplast polyribosomes with those from the surrounding cytoplasm showed the former to be 20 times more active. PMID:5411422

  8. Chloroplast unfolded protein response, a new plastid stress signaling pathway?

    PubMed

    Ramundo, Silvia; Rochaix, Jean-David

    2014-01-01

    A unique feature of the ATP-dependent ClpP protease of eukaryotic photosynthetic organisms is that its catalytic subunit ClpP1 is encoded by the chloroplast genome. Attempts to inactivate this subunit through chloroplast transformation have failed because it is essential for cell survival. To study the function of ClpP we have developed a repressible chloroplast gene expression system in Chlamydomonas reinhardtii. This system is based on the use of a chimeric nuclear gene in which the vitamin-repressible MetE promoter and Thi4 riboswitch have been fused to the coding sequence of Nac2. Upon entry into the chloroplast the Nac2 protein specifically interacts with the psbD 5'UTR and is required for the proper processing/translation of the psbD mRNA. This property can be conveyed to any chloroplast mRNA by replacing its 5'UTR with that of psbD. In this study we have chosen clpP1 as plastid target gene and examined the cellular events induced upon depletion of ClpP through transcriptomic, proteomic, biochemical and electron microscope analysis. Among the most striking features, a massive increase in protein abundance occurs for plastid chaperones, proteases and proteins involved in membrane assembly/disassembly strongly suggesting the existence of a chloroplast unfolded protein response. PMID:25482768

  9. Role of mitochondria in sulfolipid biosynthesis by Euglena chloroplasts

    SciTech Connect

    Saidha, T.; Schiff, J.A.

    1987-04-01

    Sulfate activation occurs in Euglena mitochondria the authors now find that the sulfate activating enzymes are absent from Euglena chloroplasts. Cells of mutant W/sub 10/BSmL lacking plastids also lack detectable sulfolipid (SL) when grown on /sup 35/SO/sub 4//sup 2 -/ indicating that SL is absent from the mitochondria and is exclusively in the plastids. Plastids alone will convert /sup 35/S-cysteine to /sup 35/SL in the presence of ATP and Mg/sup 2 +/; light is stimulatory. Under similar conditions, chloroplasts and mitochondria incubated together convert /sup 35/SO/sub 4//sup 2 -/ to plastid-localized /sup 35/SL but either organelle incubated alone fails to effect this conversion. Unlabeled cysteine blocks SL labeling from sulfate in the mixed incubation; since cysteine is formed from sulfate by Euglena mitochrondria, cysteine (and other compounds) may move from the mitochondrion to the chloroplast to provide the sulfo group for SL formation. Although mitochondria form labeled protein from /sup 35/SO/sub 4//sup 2 -/ via cysteine, chloroplasts alone do not form labeled protein from /sup 35/SO/sub 4//sup 2 -/, ATP and Mg/sup 2 +/ in light or darkness; incubation of chloroplasts plus mitochondria under these conditions labels chloroplast protein.

  10. Dimorphic chloroplasts in the epidermis of Podostemoideae, a subfamily of the unique aquatic angiosperm family Podostemaceae.

    PubMed

    Fujinami, Rieko; Yoshihama, Isao; Imaichi, Ryoko

    2011-09-01

    Plants of the Podostemoideae, a subfamily of the unique aquatic angiosperm family Podostemaceae, which are found in rapids and waterfalls of the tropics and subtropics, have two different sizes of chloroplasts in their epidermis. These small and large chloroplasts are located separately in each epidermal cell along its upper and inner tangential walls, respectively. This is the first case of the chloroplast dimorphism in a single epidermal cell of angiosperms. While the large chloroplasts have well developed starch grains, the small chloroplasts have a normal granal ultrastructure but very few starch grains. This suggests that the small chloroplasts mainly function in CO(2) uptake for photosynthesis from torrential water. PMID:21120679

  11. WHIRLY1 is a major organizer of chloroplast nucleoids

    PubMed Central

    Krupinska, Karin; Oetke, Svenja; Desel, Christine; Mulisch, Maria; Schäfer, Anke; Hollmann, Julien; Kumlehn, Jochen; Hensel, Götz

    2014-01-01

    WHIRLY1 is an abundant protein of chloroplast nucleoids, which has also been named pTAC-1 with regard to its detection in the proteome of transcriptionally active chromosomes (TAC). In barley primary foliage leaves, expression of the WHIRLY1 gene is highest at the base whereas protein accumulation is highest in the middle of the leaf where young developing chloroplasts are found. In order to elucidate the function of WHIRLY1 in chloroplast nucleoids, transgenic barley plants with an RNAi-mediated knock-down of the HvWHIRLY1 gene (RNAi-W1) were generated. The homozygous RNAi-W1-7 plants, barely containing traces of the WHIRLY1 protein, were chosen for detailed analyses of nucleoids. Nucleic acid specific-staining with YO-PRO®-1 revealed that in comparison to wild type chloroplasts, which have multiple small nucleoids attached to thylakoids, chloroplasts of the transgenic plants contain large irregularly formed patches of DNA besides nucleoids that are similar in size and shape to those of wild type chloroplasts. In large electron lucent areas, filamentous structures were detected by conventional transmission electron microscopy. Analyses of ptDNA levels by both DNA dot-blot hybridization and quantitative PCR showed that leaves of the transgenic plants have a two- to three-fold higher level of ptDNA than the wild type. The higher ptDNA level in RNAi-W1 plants coincided with an enhanced expression of the gene encoding a putative organelle targeted DNA polymerase in the mid part of primary foliage leaves. Furthermore, overexpression of the barley WHIRLY1 gene in E. coli cells revealed a higher compaction of bacterial nucleoids. These results suggest that WHIRLY1 belongs to the group of plastid nucleoid associated proteins (ptNAP) having a function in compacting a subpopulation of chloroplast nucleoids thereby affecting DNA replication. PMID:25237316

  12. Transplastomic integration of a cyanobacterial bicarbonate transporter into tobacco chloroplasts.

    PubMed

    Pengelly, J J L; Förster, B; von Caemmerer, S; Badger, M R; Price, G D; Whitney, S M

    2014-07-01

    Improving global yields of agricultural crops is a complex challenge with evidence indicating benefits in productivity are achieved by enhancing photosynthetic carbon assimilation. Towards improving rates of CO2 capture within leaf chloroplasts, this study shows the versatility of plastome transformation for expressing the Synechococcus PCC7002 BicA bicarbonate transporter within tobacco plastids. Fractionation of chloroplast membranes from transplastomic tob(BicA) lines showed that ~75% of the BicA localized to the thylakoid membranes and ~25% to the chloroplast envelope. BicA levels were highest in young emerging tob(BicA) leaves (0.12 μmol m(-2), ≈7mg m(-2)) accounting for ~0.1% (w/w) of the leaf protein. In these leaves, the molar amount of BicA was 16-fold lower than the abundant thylakoid photosystem II D1 protein (~1.9 μmol m(-2)) which was comparable to the 9:1 molar ratio of D1:BicA measured in air-grown Synechococcus PCC7002 cells. The BicA produced had no discernible effect on chloroplast ultrastructure, photosynthetic CO2-assimilation rates, carbon isotope discrimination, or growth of the tob(BicA) plants, implying that the bicarbonate transporter had little or no activity. These findings demonstrate the utility of plastome transformation for targeting bicarbonate transporter proteins into the chloroplast membranes without impeding growth or plastid ultrastructure. This study establishes the span of experimental measurements required to verify heterologous bicarbonate transporter function and location in chloroplasts and underscores the need for more detailed understanding of BicA structure and function to identify solutions for enabling its activation and operation in leaf chloroplasts. PMID:24965541

  13. Nitrite Uptake into Intact Pea Chloroplasts 1

    PubMed Central

    Brunswick, Pamela; Cresswell, Christopher F.

    1988-01-01

    The relationship between net nitrite uptake and its reduction in intact pea chloroplasts was investigated employing electron transport regulators, uncouplers, and photophosphorylation inhibitors. Observations confirmed the dependence of nitrite uptake on stromal pH and nitrite reduction but also suggested a partial dependance upon PSI phosphorylation. It was also suggested that ammonia stimulates nitrogen assimilation in the dark by association with stromal protons. Inhibition of nitrite uptake by N-ethylmaleimide and dinitrofluorobenzene could not be completely attributed to their inhibition of carbon dioxide fixation. Other protein binding reagents which inhibited photosynthesis showed no effect on nitrite uptake, except for p-chlormercuribenzoate which stimulated nitrite uptake. The results with N-ethylmaleimide and dinitrofluorobenzene tended to support the proposed presence of a protein permeation channel for nitrite uptake in addition to HNO2 penetration. On the basis of a lack of effect by known anion uptake inhibitors, it was concluded that the nitrite uptake mechanism was distinct from that of phosphate and chloride/sulfate transport. PMID:16665917

  14. Chloroplast genome structure in Ilex (Aquifoliaceae)

    PubMed Central

    Yao, Xin; Tan, Yun-Hong; Liu, Ying-Ying; Song, Yu; Yang, Jun-Bo; Corlett, Richard T.

    2016-01-01

    Aquifoliaceae is the largest family in the campanulid order Aquifoliales. It consists of a single genus, Ilex, the hollies, which is the largest woody dioecious genus in the angiosperms. Most species are in East Asia or South America. The taxonomy and evolutionary history remain unclear due to the lack of a robust species-level phylogeny. We produced the first complete chloroplast genomes in this family, including seven Ilex species, by Illumina sequencing of long-range PCR products and subsequent reference-guided de novo assembly. These genomes have a typical bicyclic structure with a conserved genome arrangement and moderate divergence. The total length is 157,741 bp and there is one large single-copy region (LSC) with 87,109 bp, one small single-copy with 18,436 bp, and a pair of inverted repeat regions (IR) with 52,196 bp. A total of 144 genes were identified, including 96 protein-coding genes, 40 tRNA and 8 rRNA. Thirty-four repetitive sequences were identified in Ilex pubescens, with lengths >14 bp and identity >90%, and 11 divergence hotspot regions that could be targeted for phylogenetic markers. This study will contribute to improved resolution of deep branches of the Ilex phylogeny and facilitate identification of Ilex species. PMID:27378489

  15. Limited variation across two chloroplast genomes with finishing chloroplast genome of Capsella grandiflora.

    PubMed

    Wu, Zhiqiang; Ma, Qiumao

    2016-09-01

    The complete chloroplast genome of Capsella grandiflora is finished in this study, which consists of 154 638 base pairs (bp) in size containing a pair of inverted repeats (IRa and IRb) of 26 462 bp each and a small and large single-copies (SSC and LSC) of 17 835 and 83 879 bp, respectively. The overall GC content is 36.54% and the GC contents of LSC, IRs, and SSC are 34.34%, 42.38%, and 29.61% separately. The gene contents and numbers are the same with other two published species in genus Capsella with 112 annotated unique genes including 78 protein-coding genes, 30 tRNA genes, and four rRNA genes. Among these, 16 are duplicated in the inverted repeat regions, 15 genes contained one intron, and three genes (rps12, clpP, and ycf3) comprising two introns. Based on the whole genome comparison, only 28 SNPs and 16 Indels (insertion and deletion) are detected between two closed relatives' chloroplast genomes. PMID:26186303

  16. Chloroplasts Are Central Players in Sugar-Induced Leaf Growth1[OPEN

    PubMed Central

    De Milde, Liesbeth; Maleux, Katrien

    2016-01-01

    Leaves are the plant’s powerhouses, providing energy for all organs through sugar production during photosynthesis. However, sugars serve not only as a metabolic energy source for sink tissues but also as signaling molecules, affecting gene expression through conserved signaling pathways to regulate plant growth and development. Here, we describe an in vitro experimental assay, allowing one to alter the sucrose (Suc) availability during early Arabidopsis (Arabidopsis thaliana) leaf development, with the aim to identify the affected cellular and molecular processes. The transfer of seedlings to Suc-containing medium showed a profound effect on leaf growth by stimulating cell proliferation and postponing the transition to cell expansion. Furthermore, rapidly after transfer to Suc, mesophyll cells contained fewer and smaller plastids, which are irregular in shape and contain fewer starch granules compared with control mesophyll cells. Short-term transcriptional responses after transfer to Suc revealed the repression of well-known sugar-responsive genes and multiple genes encoded by the plastid, on the one hand, and up-regulation of a GLUCOSE-6-PHOSPHATE TRANSPORTER (GPT2), on the other hand. Mutant gpt2 seedlings showed no stimulation of cell proliferation and no repression of chloroplast-encoded transcripts when transferred to Suc, suggesting that GPT2 plays a critical role in the Suc-mediated effects on early leaf growth. Our findings, therefore, suggest that induction of GPT2 expression by Suc increases the import of glucose-6-phosphate into the plastids that would repress chloroplast-encoded transcripts, restricting chloroplast differentiation. Retrograde signaling from the plastids would then delay the transition to cell expansion and stimulate cell proliferation. PMID:26932234

  17. The complete chloroplast genome of Capsicum frutescens (Solanaceae)1

    PubMed Central

    Shim, Donghwan; Raveendar, Sebastin; Lee, Jung-Ro; Lee, Gi-An; Ro, Na-Young; Jeon, Young-Ah; Cho, Gyu-Taek; Lee, Ho-Sun; Ma, Kyung-Ho; Chung, Jong-Wook

    2016-01-01

    Premise of the study: We report the complete sequence of the chloroplast genome of Capsicum frutescens (Solanaceae), a species of chili pepper. Methods and Results: Using an Illumina platform, we sequenced the chloroplast genome of C. frutescens. The total length of the genome is 156,817 bp, and the overall GC content is 37.7%. A pair of 25,792-bp inverted repeats is separated by small (17,853 bp) and large (87,380 bp) single-copy regions. The C. frutescens chloroplast genome encodes 132 unique genes, including 87 protein-coding genes, 37 transfer RNA (tRNA) genes, and eight ribosomal RNA (rRNA) genes. Of these, seven genes are duplicated in the inverted repeats and 12 genes contain one or two introns. Comparative analysis with the reference chloroplast genome revealed 125 simple sequence repeat motifs and 34 variants, mostly located in the noncoding regions. Conclusions: The complete chloroplast genome sequence of C. frutescens reported here is a valuable genetic resource for Capsicum species. PMID:27213127

  18. Comparison of intraspecific, interspecific and intergeneric chloroplast diversity in Cycads

    PubMed Central

    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

  19. Comparison of intraspecific, interspecific and intergeneric chloroplast diversity in Cycads.

    PubMed

    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

  20. The chloroplast view of the evolution of polyploid wheat.

    PubMed

    Gornicki, Piotr; Zhu, Huilan; Wang, Junwei; Challa, Ghana S; Zhang, Zhengzhi; Gill, Bikram S; Li, Wanlong

    2014-11-01

    Polyploid wheats comprise four species: Triticum turgidum (AABB genomes) and T. aestivum (AABBDD) in the Emmer lineage, and T. timopheevii (AAGG) and T. zhukovskyi (AAGGA(m) A(m) ) in the Timopheevi lineage. Genetic relationships between chloroplast genomes were studied to trace the evolutionary history of the species. Twenty-five chloroplast genomes were sequenced, and 1127 plant accessions were genotyped, representing 13 Triticum and Aegilops species. The A. speltoides (SS genome) diverged before the divergence of T. urartu (AA), A. tauschii (DD) and the Aegilops species of the Sitopsis section. Aegilops speltoides forms a monophyletic clade with the polyploid Emmer and Timopheevi wheats, which originated within the last 0.7 and 0.4 Myr, respectively. The geographic distribution of chloroplast haplotypes of the wild tetraploid wheats and A. speltoides illustrates the possible geographic origin of the Emmer lineage in the southern Levant and the Timopheevi lineage in northern Iraq. Aegilops speltoides is the closest relative of the diploid donor of the chloroplast (cytoplasm), as well as the B and G genomes to Timopheevi and Emmer lineages. Chloroplast haplotypes were often shared by species or subspecies within major lineages and between the lineages, indicating the contribution of introgression to the evolution and domestication of polyploid wheats. PMID:25059383

  1. Arabidopsis chloroplast chaperonin 10 is a calmodulin-binding protein

    NASA Technical Reports Server (NTRS)

    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.

  2. NUCLEIC ACIDS OF CHLOROPLASTS AND MITOCHONDRIA IN SWISS CHARD.

    PubMed

    KISLEV, N; SWIFT, H; BOGORAD, L

    1965-05-01

    Nucleic acids in young leaves of Swiss chard have been studied by light and electron microscope techniques. Leaf DNA has also been characterized by density gradient centrifugation and shown to contain a minor band of higher guanine plus cytosine (GC) content, presumably attributable to chloroplasts. The chloroplasts were faintly stained by the Feulgen reaction; radioautography demonstrated the incorporation of tritiated thymidine in the cytoplasm and in some nuclei. The Feulgen stainability and most of the radioactivity were removable with DNase. Under the electron microscope, both mitochondria and chloroplasts were found to contain filamentous and particulate components within the matrix areas. The morphology of the filamentous component was dependent on the fixation, being partially clumped after OSO(4) or formalin, but finely filamentous after Kellenberger fixation. The filaments were stainable with uranyl acetate, and were extractable with DNase following formalin fixation under conditions in which nuclear DNA was also extracted. The particulate component, after formalin fixation and uranyl staining, was prominent in chloroplasts from young leaves, but was only sparsely distributed in mitochondria. The stainability was removed with ribonuclease. We have concluded that chloroplasts and mitochondria of Swiss chard possess a filamentous component that contains DNA, probably responsible for both cytoplasmic thymidine incorporation and the minor band in CsCl centrifugation. A particulate ribosome-like component that contains RNA is also present. PMID:14287184

  3. Inhibition of chloroplastic respiration by osmotic dehydration. [Spinacia oleracea L

    SciTech Connect

    Willeford, K.O.; Ahluwalia, K.J.K.; Gibbs, M. )

    1989-04-01

    The respiratory capacity of isolated spinach (Spinacia oleracea L.) chloroplasts, measured as the rate of {sup 14}CO{sub 2} evolved from the oxidative pentose phosphate cycle in darkened chloroplasts exogenously supplied with ({sup 14}C)glucose, was progressively diminished by escalating osmotic dehydration with betaine or sorbitol. Comparing the inhibitions of CO{sub 2} evolution generated by osmotic dehydration in chloroplasts given C-1 and C-6 labeled glucose, 54% and 84%, respectively, indicates that osmotic dehydration effects to a greater extent the recycling of the oxidative pentose phosphate intermediates, fructose-6P and glyceraldehyde-3P. Respiratory inhibition in the darkened chloroplast could be alleviated by addition of NH{sub 4}Cl (a stromal alkylating agent), iodoacetamide (an inhibitor of glyceraldehyde-3P dehydrogenase), or glycolate-2P (an inhibitor of phosphofructokinase). It is concluded that the site which primarily mediates respiratory inhibition in the darkened chloroplast occurs at the fructose 1,6-bisphosphatase/phosphofructokinase junction.

  4. Ftsz Ring Formation at the Chloroplast Division Site in Plants

    PubMed Central

    Vitha, Stanislav; McAndrew, Rosemary S.; Osteryoung, Katherine W.

    2001-01-01

    Among the events that accompanied the evolution of chloroplasts from their endosymbiotic ancestors was the host cell recruitment of the prokaryotic cell division protein FtsZ to function in chloroplast division. FtsZ, a structural homologue of tubulin, mediates cell division in bacteria by assembling into a ring at the midcell division site. In higher plants, two nuclear-encoded forms of FtsZ, FtsZ1 and FtsZ2, play essential and functionally distinct roles in chloroplast division, but whether this involves ring formation at the division site has not been determined previously. Using immunofluorescence microscopy and expression of green fluorescent protein fusion proteins in Arabidopsis thaliana, we demonstrate here that FtsZ1 and FtsZ2 localize to coaligned rings at the chloroplast midpoint. Antibodies specific for recognition of FtsZ1 or FtsZ2 proteins in Arabidopsis also recognize related polypeptides and detect midplastid rings in pea and tobacco, suggesting that midplastid ring formation by FtsZ1 and FtsZ2 is universal among flowering plants. Perturbation in the level of either protein in transgenic plants is accompanied by plastid division defects and assembly of FtsZ1 and FtsZ2 into filaments and filament networks not observed in wild-type, suggesting that previously described FtsZ-containing cytoskeletal-like networks in chloroplasts may be artifacts of FtsZ overexpression. PMID:11285278

  5. Whole Chloroplast Genome Sequencing in Fragaria Using Deep Sequencing: A Comparison of Three Methods

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Chloroplast sequences previously investigated in Fragaria revealed low amounts of variation. Deep sequencing technologies enable economical sequencing of complete chloroplast genomes. These sequences can potentially provide robust phylogenetic resolution, even at low taxonomic levels within plant gr...

  6. Treatment with Antibiotics that Interfere with Peptidoglycan Biosynthesis Inhibits Chloroplast Division in the Desmid Closterium

    PubMed Central

    Matsumoto, Hiroko; Takechi, Katsuaki; Sato, Hiroshi; Takio, Susumu; Takano, Hiroyoshi

    2012-01-01

    Charophytes is a green algal group closely related to land plants. We investigated the effects of antibiotics that interfere with peptidoglycan biosynthesis on chloroplast division in the desmid Closterium peracerosum–strigosum–littorale complex. To detect cells just after division, we used colchicine, which inhibits Closterium cell elongation after division. Although normal Closterium cells had two chloroplasts before and after cell division, cells treated with ampicillin, D-cycloserine, or fosfomycin had only one chloroplast after cell division, suggesting that the cells divided without chloroplast division. The antibiotics bacitracin and vancomycin showed no obvious effect. Electron microscopic observation showed that irregular-shaped chloroplasts existed in ampicillin-treated Closterium cells. Because antibiotic treatments resulted in the appearance of long cells with irregular chloroplasts and cell death, we counted cell types in the culture. The results suggested that cells with one chloroplast appeared first and then a huge chloroplast was generated that inhibited cell division, causing elongation followed by cell death. PMID:22815801

  7. Chloroplast DNA diversity among wild and cultivated members of Cucurbita (Cucurbitaceae).

    PubMed

    Wilson, H D; Doebley, J; Duvall, M

    1992-09-01

    Cladistic analysis of 86 chloroplast DNA restriction-site mutations among 30 samples representing 15 species of Cucurbita indicates that annual species of the genus are derived from perennials. The Malabar Gourd, C. ficifolia, is placed as a basal, sister taxon relative to other domesticated species and allied wild-types. The pattern of variation supports three species groups as monophyletic: (1) C. fraterna, C. pepo, and C. texana, (2) C. lundelliana, C. martinezii, C. mixta, C. moschata and C. sororia, and (3) C. foetidissima and C. pedatifolia. Domesticated samples representing subspecies of C. pepo are divided into two concordant groups, one of which is allied to wild-types referable to C. texana and C. fraterna. The data failed to resolve relationships among cultivars of C. moschata and C. mixta and their association to the wild C. sororia. The South American domesticate, C. maxima, and its companion weed, C. andreana, show close affinity and alliance to C. equadorensis. PMID:24201487

  8. Primary structure of maize chloroplast adenylate kinase.

    PubMed

    Schiltz, E; Burger, S; Grafmüller, R; Deppert, W R; Haehnel, W; Wagner, E

    1994-06-15

    This paper describes the sequence of adenylate kinase (Mg-ATP+AMP<-->Mg-ADP+ADP) from maize chloroplasts. This light-inducible enzyme is important for efficient CO2 fixation in the C4 cycle, by removing and recycling AMP produced in the reversible pyruvate phosphate dikinase reaction. The complete sequence was determined by analyzing peptides from cleavages with trypsin, AspN protease and CNBr and subcleavage of a major CNBr peptide with chymotrypsin. N-terminal Edman degradation and carboxypeptidase digestion established the terminal residues. Electrospray mass spectrometry confirmed the final sequence of 222 residues (M(r) = 24867) including one cysteine and one tryptophan. The sequence shows this enzyme to be a long-variant-type adenylate kinase, the nearest relatives being adenylate kinases from Enterobacteriaceae. Alignment of the sequence with the adenylate kinase from Escherichia coli reveals 44% identical residues. Since the E. coli structure has been published recently at 0.19-nm resolution with the inhibitor adenosine(5')pentaphospho(5')adenosine (Ap5A) [Müller, C. W. & Schulz, G. E. (1992) J. Mol. Biol. 224, 159-177], catalytically essential residues could be compared and were found to be mostly conserved. Surprisingly, in the nucleotide-binding Gly-rich loop Gly-Xaa-Pro-Gly-Xaa-Gly-Lys the middle Gly is replaced by Ala. This is, however, compensated by an Ile-->Val exchange in the nearest spatial neighborhood. A Thr-->Ala exchange explains the unusual tolerance of the enzyme for pyrimidine nucleotides in the acceptor site. PMID:8026505

  9. Development of chloroplast genomic resources for Cynara.

    PubMed

    Curci, Pasquale L; De Paola, Domenico; Sonnante, Gabriella

    2016-03-01

    In this study, new chloroplast (cp) resources were developed for the genus Cynara, using whole cp genomes from 20 genotypes, by means of high-throughput sequencing technologies. Our target species included seven globe artichokes, two cultivated cardoons, eight wild artichokes, and three other wild Cynara species (C. baetica, C. cornigera and C. syriaca). One complete cp genome was isolated using short reads from a whole-genome sequencing project, while the others were obtained by means of long-range PCR, for which primer pairs are provided here. A de novo assembly strategy combined with a reference-based assembly allowed us to reconstruct each cp genome. Comparative analyses among the newly sequenced genotypes and two additional Cynara cp genomes ('Brindisino' artichoke and C. humilis) retrieved from public databases revealed 126 parsimony informative characters and 258 singletons in Cynara, for a total of 384 variable characters. Thirty-nine SSR loci and 34 other INDEL events were detected. After data analysis, 37 primer pairs for SSR amplification were designed, and these molecular markers were subsequently validated in our Cynara genotypes. Phylogenetic analysis based on all cp variable characters provided the best resolution when compared to what was observed using only parsimony informative characters, or only short 'variable' cp regions. The evaluation of the molecular resources obtained from this study led us to support the 'super-barcode' theory and consider the total cp sequence of Cynara as a reliable and valuable molecular marker for exploring species diversity and examining variation below the species level. PMID:26354522

  10. Chloroplast incorporation and long-term photosynthetic performance through the life cycle in laboratory cultures of Elysia timida (Sacoglossa, Heterobranchia)

    PubMed Central

    2014-01-01

    Introduction The Mediterranean sacoglossan Elysia timida is one of the few sea slug species with the ability to sequester chloroplasts from its food algae and to subsequently store them in a functional state in the digestive gland cells for more than a month, during which time the plastids retain high photosynthetic activity (= long-term retention). Adult E. timida have been described to feed on the unicellular alga Acetabularia acetabulum in their natural environment. The suitability of E. timida as a laboratory model culture system including its food source was studied. Results In contrast to the literature reporting that juvenile E. timida feed on Cladophora dalmatica first, and later on switch to the adult diet A. acetabulum, the juveniles in this study fed directly on A. acetabulum (young, non-calcified stalks); they did not feed on the various Cladophora spp. (collected from the sea or laboratory culture) offered. This could possibly hint to cryptic speciation with no clear morphological differences, but incipient ecological differentiation. Transmission electron microscopy of chloroplasts from A. acetabulum after initial intake by juvenile E. timida showed different states of degradation — in conglomerations or singularly — and fragments of phagosome membranes, but differed from kleptoplast images of C. dalmatica in juvenile E. timida from the literature. Based on the finding that the whole life cycle of E. timida can be completed with A. acetabulum as the sole food source, a laboratory culture system was established. An experiment with PAM-fluorometry showed that cultured E. timida are also able to store chloroplasts in long-term retention from Acetabularia peniculus, which stems from the Indo-Pacific and is not abundant in the natural environment of E. timida. Variations between three experiment groups indicated potential influences of temperature on photosynthetic capacities. Conclusions E. timida is a viable laboratory model system to study

  11. Oryza sativa Chloroplast Signal Recognition Particle 43 (OscpSRP43) Is Required for Chloroplast Development and Photosynthesis

    PubMed Central

    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. PMID:26600124

  12. Kinetic studies of interfacial photocurrents in platinized chloroplasts

    SciTech Connect

    Greenbaum, E.

    1992-12-01

    The present experiments focus on kinetic studies of phototocurrents generated in a photobioelectrochemical cell constructed from platinized chloroplast membranes. These chloroplast membranes although separated from the CO{sub 2}-reducing enzymes of the Calvin-Benson cycle, contain the full complement of photosystem I and II reaction centers along with the electron transport chain linking these two centers. The vectorial model of photosynthesis indicates that the orientation of the reaction centers in the photosynthetic membranes is such that electrons emerge from the membranes into the stroma region of the chloroplasts. Since the flattened saclike vesicles of the thylakoid membranes are topologically equivalent to spheres, it follows that, irrespective of the rotational orientation of the membranes, the photogenerated electrons emerge from the reaction centers in a radial direction away from the intra-thylakoid region.

  13. The complete chloroplast genome sequence of Anoectochilus roxburghii.

    PubMed

    Yu, Chao-Wei; Lian, Qin; Wu, Kang-Cheng; Yu, Shu-Han; Xie, Li-Yan; Wu, Zu-Jian

    2016-07-01

    The complete chloroplast sequence of the Anoectochilus roxburghii, a popular traditional Chinese medicine for the treatment of cancer, was determined in this study. The chloroplast genome (cpDNA)^ was 152,802 bp in length, containing a pair of inverted repeats of 52,728 bp separated by a large single-copy region and a small single-copy region of 82,641 bp and 17,433 bp, respectively. The chloroplast genome encodes 116 predicted functional genes, including 81 protein-coding genes, four ribosomal RNA genes, and 31 transfer RNA genes, 25 of which are duplicated in the inverted repeat regions. The cpDNA is GC-rich (36.9%). PMID:25865497

  14. The complete chloroplast genome sequence of Alocasia macrorrhizos.

    PubMed

    Wang, Bin; Han, Limin

    2016-09-01

    The complete chloroplast sequence of Alocasia macrorrhizos is 154 995 bp in length, containing a pair of inverted repeats of 25 944 bp separated by a large single-copy (LSC) region and a small single-copy (SSC) region of 87 366 bp and 15 741 bp, respectively. The chloroplast genome encodes 132 predicted functional genes, including 87 protein-coding genes, four ribosomal RNA genes, and 37 transfer RNA genes, 18 of which are duplicated in the inverted repeat regions. In these genes, 16 genes contained single intron and two genes comprising double introns. A maximum-likelihood phylogenetic analysis using complete chloroplast genome revealed that A. macrorrhizos does not belong to Araceae family, which infers that the A. macrorrhizos is distant from the species in Araceae family. PMID:26258514

  15. S-sulfocysteine synthase function in sensing chloroplast redox status

    PubMed Central

    Gotor, Cecilia; Romero, Luis C.

    2013-01-01

    The minor chloroplastic O-acetylserine(thiol)lyase isoform encoded by the CS26 gene in Arabidopsis thaliana has been described as an S-sulfocysteine synthase enzyme that plays an important role in chloroplast function. This enzyme is located in the thylakoid lumen, and its S-sulfocysteine activity is essential for the proper photosynthetic performance of the chloroplast under long-day growth conditions. Based on the present knowledge of this enzyme, we suggest that S-sulfocysteine synthase functions as a protein sensor to detect the accumulation of thiosulfate as a result of the inadequate detoxification of reactive oxygen species generated under conditions of excess light to produce the S-sulfocysteine molecule that triggers protection mechanisms of the photosynthetic apparatus. PMID:23333972

  16. The complete chloroplast genome sequence of Perilla frutescens (L.).

    PubMed

    Shen, Qi; Yang, Jun; Lu, Chaolong; Wang, Bo; Song, Chi

    2016-09-01

    Perilla frutescens (L.) is a traditionally medical herb of East Asia. The complete chloroplast genome of P. frutescens (L.) Britton var. frutescens was assembled in this study. Total chloroplast genome size of Perilla was 153,666 bp in length, containing a pair of inverted repeats (IRs) of 25,677 bp, separated by large single copy (LSC) and small single copy (SSC) of 84,288 bp and 17,925 bp, respectively. Overall GC contents of the genome were 37.6%. The chloroplast genome harbored 127 annotated genes, including 89 protein-coding genes, 29 tRNA genes and 8 rRNA genes. Eleven genes contained one or two introns. PMID:25714143

  17. The complete chloroplast genome sequence of Spathiphyllum kochii.

    PubMed

    Han, Limin; Wang, Bin; Wang, Zhe Zhi

    2016-07-01

    The complete chloroplast sequence of the Spathiphyllum kochii is 163 368 bp in length, containing a pair of inverted repeats of 25 270 bp separated by a large single-copy region and a small single-copy region of 90 482 bp and 22 346 bp, respectively. The chloroplast genome encodes 133 predicted functional genes, including 88 protein-coding genes, four ribosomal RNA genes and 37 transfer RNA genes, 18 of which are duplicated in the inverted repeat regions. The cpDNA is GC-rich (37.6%). The chloroplast genome of S. kochii reported here will lay basis for identification, utilization and protection of its germplasm resources. PMID:26134343

  18. Breakthrough in chloroplast genetic engineering of agronomically important crops

    PubMed Central

    Daniell, Henry; Kumar, Shashi; Dufourmantel, Nathalie

    2012-01-01

    Chloroplast genetic engineering offers several unique advantages, including high-level transgene expression, multi-gene engineering in a single transformation event and transgene containment by maternal inheritance, as well as a lack of gene silencing, position and pleiotropic effects and undesirable foreign DNA. More than 40 transgenes have been stably integrated and expressed using the tobacco chloroplast genome to confer desired agronomic traits or express high levels of vaccine antigens and biopharmaceuticals. Despite such significant progress, this technology has not been extended to major crops. However, highly efficient soybean, carrot and cotton plastid transformation has recently been accomplished through somatic embryogenesis using species-specific chloroplast vectors. This review focuses on recent exciting developments in this field and offers directions for further research and development. PMID:15866001

  19. The complete chloroplast genome sequence of Dieffenbachia seguine (Araceae).

    PubMed

    Wang, Bin; Han, Limin; Chen, Chen; Wang, Zhezhi

    2016-07-01

    The nucleotide sequence of the chloroplast genome from Dieffenbachia seguine is the first to have complete genome sequence from genus of Dieffenbachia family Araceae. The genome size is 163 699 bp in length, with 36.4% GC content. A pair of inverted repeats (IRs, 25 235 bp) is separated by a large single copy region (LSC, 90 780 bp) and a small single copy region (SSC, 22 449 bp). The chloroplast genome contains 113 unique genes, 88 protein-coding genes, 37 tRNA genes, and four rRNA genes. In these genes, 16 genes contained single intron and two genes composed of double introns. A maximum likelihood phylogenetic analysis using complete chloroplast genome revealed that Dieffenbachia seguine belongs to the Araceae family of the Arecidae group, which is conform to the traditional classification. PMID:26153749

  20. Post-Transcriptional Control of Chloroplast Gene Expression

    PubMed Central

    del Campo, Eva M.

    2009-01-01

    Chloroplasts contain their own genome, organized as operons, which are generally transcribed as polycistronic transcriptional units. These primary transcripts are processed into smaller RNAs, which are further modified to produce functional RNAs. The RNA processing mechanisms remain largely unknown and represent an important step in the control of chloroplast gene expression. Such mechanisms include RNA cleavage of pre-existing RNAs, RNA stabilization, intron splicing, and RNA editing. Recently, several nuclear-encoded proteins that participate in diverse plastid RNA processing events have been characterised. Many of them seem to belong to the pentatricopeptide repeat (PPR) protein family that is implicated in many crucial functions including organelle biogenesis and plant development. This review will provide an overview of current knowledge of the post-transcriptional processing in chloroplasts. PMID:19838333

  1. Phosphatidylinositol 4-Phosphate Negatively Regulates Chloroplast Division in Arabidopsis[OPEN

    PubMed Central

    Okazaki, Kumiko; Miyagishima, Shin-ya; Wada, Hajime

    2015-01-01

    Chloroplast division is performed by the constriction of envelope membranes at the division site. Although constriction of a ring-like protein complex has been shown to be involved in chloroplast division, it remains unknown how membrane lipids participate in the process. Here, we show that phosphoinositides with unknown function in envelope membranes are involved in the regulation of chloroplast division in Arabidopsis thaliana. PLASTID DIVISION1 (PDV1) and PDV2 proteins interacted specifically with phosphatidylinositol 4-phosphate (PI4P). Inhibition of phosphatidylinositol 4-kinase (PI4K) decreased the level of PI4P in chloroplasts and accelerated chloroplast division. Knockout of PI4Kβ2 expression or downregulation of PI4Kα1 expression resulted in decreased levels of PI4P in chloroplasts and increased chloroplast numbers. PI4Kα1 is the main contributor to PI4P synthesis in chloroplasts, and the effect of PI4K inhibition was largely abolished in the pdv1 mutant. Overexpression of DYNAMIN-RELATED PROTEIN5B (DRP5B), another component of the chloroplast division machinery, which is recruited to chloroplasts by PDV1 and PDV2, enhanced the effect of PI4K inhibition, whereas overexpression of PDV1 and PDV2 had additive effects. The amount of DRP5B that associated with chloroplasts increased upon PI4K inhibition. These findings suggest that PI4P is a regulator of chloroplast division in a PDV1- and DRP5B-dependent manner. PMID:25736058

  2. Salinity induces membrane structure and lipid changes in maize mesophyll and bundle sheath chloroplasts.

    PubMed

    Omoto, Eiji; Iwasaki, Yugo; Miyake, Hiroshi; Taniguchi, Mitsutaka

    2016-05-01

    The membranes of Zea mays (maize) mesophyll cell (MC) chloroplasts are more vulnerable to salinity stress than are those of bundle sheath cell (BSC) chloroplasts. To clarify the mechanism underlying this difference in salt sensitivity, we monitored changes in the glycerolipid and fatty acid compositions of both types of chloroplast upon exposure to salinity stress. The monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG) contents were higher in MC chloroplasts than in BSC chloroplasts, in both the presence and absence of salt treatment. Under salt conditions, the MGDG level in MC chloroplasts was significantly lower than under normal conditions, while it was unchanged in BSC chloroplasts. In both types of chloroplast, the contents of DGDG, phosphatidylglycerol and phosphatidylinositol remained at the same levels in control and salt-treated plants, whereas sulfoquinovosyldiacylglycerol and phosphatidylcholine were significantly lower and higher, respectively, upon salt treatment. In addition, the fatty acid composition and double bond index of individual lipid classes were changed by salt treatment in both BSC and MC chloroplasts, although these factors had no effect on glycerolipid content. These findings suggest that the difference in salt sensitivity of MC and BSC chloroplast membranes is related to differences in MGDG responses to salinity. Thus, we propose that the low MGDG content and the low sensitivity of MGDG to salinity in BSC chloroplasts render them more tolerant than MC chloroplasts to salinity stress. PMID:26555406

  3. Transcriptome analysis of ectopic chloroplast development in green curd cauliflower (Brassica oleracea L. var. botrytis)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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 ...

  4. Manipulating the chloroplast genome of Chlamydomonas: Present realities and future prospects

    SciTech Connect

    Boynton, J.; Gillham, N.; Hauser, C.; Heifetz, P.; Lers, A.; Newman, S.; Osmond, B.

    1992-01-01

    Biotechnology is being applied in vitro modification and stable reintroduction of chloroplast genes in Chlamydomonas reinhardtii and Nicotiana tabacum by homologous recombination. We are attempting the function analyses of plastid encoded proteins involved in photosynthesis, characterization of sequences which regulate expression of plastid genes at the transcriptional and translational levels, targeted disruption of chloroplast genes and molecular analysis of processes involved in chloroplast recombination.

  5. Manipulating the chloroplast genome of Chlamydomonas: Present realities and future prospects

    SciTech Connect

    Boynton, J.; Gillham, N.; Hauser, C.; Heifetz, P.; Lers, A.; Newman, S.; Osmond, B.

    1992-12-31

    Biotechnology is being applied in vitro modification and stable reintroduction of chloroplast genes in Chlamydomonas reinhardtii and Nicotiana tabacum by homologous recombination. We are attempting the function analyses of plastid encoded proteins involved in photosynthesis, characterization of sequences which regulate expression of plastid genes at the transcriptional and translational levels, targeted disruption of chloroplast genes and molecular analysis of processes involved in chloroplast recombination.

  6. Chloroplast Iron Transport Proteins - Function and Impact on Plant Physiology.

    PubMed

    López-Millán, Ana F; Duy, Daniela; Philippar, Katrin

    2016-01-01

    Chloroplasts originated about three billion years ago by endosymbiosis of an ancestor of today's cyanobacteria with a mitochondria-containing host cell. During evolution chloroplasts of higher plants established as the site for photosynthesis and thus became the basis for all life dependent on oxygen and carbohydrate supply. To fulfill this task, plastid organelles are loaded with the transition metals iron, copper, and manganese, which due to their redox properties are essential for photosynthetic electron transport. In consequence, chloroplasts for example represent the iron-richest system in plant cells. However, improvement of oxygenic photosynthesis in turn required adaptation of metal transport and homeostasis since metal-catalyzed generation of reactive oxygen species (ROS) causes oxidative damage. This is most acute in chloroplasts, where radicals and transition metals are side by side and ROS-production is a usual feature of photosynthetic electron transport. Thus, on the one hand when bound by proteins, chloroplast-intrinsic metals are a prerequisite for photoautotrophic life, but on the other hand become toxic when present in their highly reactive, radical generating, free ionic forms. In consequence, transport, storage and cofactor-assembly of metal ions in plastids have to be tightly controlled and are crucial throughout plant growth and development. In the recent years, proteins for iron transport have been isolated from chloroplast envelope membranes. Here, we discuss their putative functions and impact on cellular metal homeostasis as well as photosynthetic performance and plant metabolism. We further consider the potential of proteomic analyses to identify new players in the field. PMID:27014281

  7. Non-reductive modulation of chloroplast fructose-1,6-bisphosphatase by 2-Cys peroxiredoxin

    SciTech Connect

    Caporaletti, Daniel; D'Alessio, Ana C.; Rodriguez-Suarez, Roberto J.; Senn, Alejandro M.; Duek, Paula D.; Wolosiuk, Ricardo A. . E-mail: rwolosiuk@leloir.org.ar

    2007-04-13

    2-Cys peroxiredoxin (2-Cys Prx) is a large group of proteins that participate in cell proliferation, differentiation, apoptosis, and photosynthesis. In the prevailing view, this ubiquitous peroxidase poises the concentration of H{sub 2}O{sub 2} and, in so doing, regulates signal transduction pathways or protects macromolecules against oxidative damage. Here, we describe First purification of 2-Cys Prx from higher plants and subsequently we show that the native and the recombinant forms of rapeseed leaves stimulate the activity of chloroplast fructose-1,6-bisphosphatase (CFBPase), a key enzyme of the photosynthetic CO{sub 2} assimilation. The absence of reductants, the strict requirement of both fructose 1,6-bisphosphate and Ca{sup 2+}, and the response of single mutants C174S and C179S CFBPase bring forward clear differences with the well-known stimulation mediated by reduced thioredoxin via the regulatory 170's loop of CFBPase. Taken together, these findings provide an unprecedented insight into chloroplast enzyme regulation wherein both 2-Cys Prx and the 170's loop of CFBPase exhibit novel functions.

  8. Multiple feedbacks between chloroplast and whole plant in the context of plant adaptation and acclimation to the environment.

    PubMed

    Demmig-Adams, Barbara; Stewart, Jared J; Adams, William W

    2014-04-19

    This review focuses on feedback pathways that serve to match plant energy acquisition with plant energy utilization, and thereby aid in the optimization of chloroplast and whole-plant function in a given environment. First, the role of source-sink signalling in adjusting photosynthetic capacity (light harvesting, photochemistry and carbon fixation) to meet whole-plant carbohydrate demand is briefly reviewed. Contrasting overall outcomes, i.e. increased plant growth versus plant growth arrest, are described and related to respective contrasting environments that either do or do not present opportunities for plant growth. Next, new insights into chloroplast-generated oxidative signals, and their modulation by specific components of the chloroplast's photoprotective network, are reviewed with respect to their ability to block foliar phloem-loading complexes, and, thereby, affect both plant growth and plant biotic defences. Lastly, carbon export capacity is described as a newly identified tuning point that has been subjected to the evolution of differential responses in plant varieties (ecotypes) and species from different geographical origins with contrasting environmental challenges. PMID:24591724

  9. A Metabolic Control Analysis of the Glutamine Synthetase/Glutamate Synthase Cycle in Isolated Barley (Hordeum vulgare L.) Chloroplasts.

    PubMed Central

    Baron, A. C.; Tobin, T. H.; Wallsgrove, R. M.; Tobin, A. K.

    1994-01-01

    Ammonia assimilation in chloroplasts occurs via the glutamine synthetase/glutamate synthase (GS/GOGAT) cycle. To determine the extent to which these enzymes contribute to the control of ammonia assimilation, a metabolic control analysis was performed on isolated barley (Hordeum vulgare L.) leaf chloroplasts. Pathway flux was measured polarographically as ammonium-plus-2-oxoglutarate-plus-glutamine-dependent O2 evolution in illuminated chloroplasts. Enzyme activity was modulated by titration with specific, irreversible inhibitors of GS (phosphinothricin) and GOGAT (azaserine). Flux control coefficients (CJ0E0) were determined (a) by differentiation of best-fit hyperbolic curves of the data sets (flux versus enzyme activity), and (b) from estimates of the deviation indices (D/[prime]E0). Both analyses gave similar values for the coefficients. The control coefficient for GS was relatively high and the value did not change significantly with changes in 2-oxoglutarate concentration (C/0E0 = 0.58 at 5 mM 2-oxoglutarate and 0.40 at 20 mM 2-oxoglutarate). The control coefficient for GOGAT decreased with decreasing glutamine concentrations, from 0.76 at 20 mM glutamine to 0.19 at 10 mM glutamine. Thus, at high concentrations of glutamine, GOGAT exerts a major control over flux with a significant contribution also from GS. At lower concentrations of glutamine, however, GOGAT exerts far less control over pathway flux. PMID:12232211

  10. Systematic analysis of plant mitochondrial and chloroplast small RNAs suggests organelle-specific mRNA stabilization mechanisms

    PubMed Central

    Ruwe, Hannes; Wang, Gongwei; Gusewski, Sandra; Schmitz-Linneweber, Christian

    2016-01-01

    Land plant organellar genomes encode a small number of genes, many of which are essential for respiration and photosynthesis. Organellar gene expression is characterized by a multitude of RNA processing events that lead to stable, translatable transcripts. RNA binding proteins (RBPs), have been shown to generate and protect transcript termini and eventually induce the accumulation of short RNA footprints. We applied knowledge of such RBP-derived footprints to develop software (sRNA miner) that enables identification of RBP footprints, or other clusters of small RNAs, in organelles. We used this tool to determine mitochondrial and chloroplast cosRNAs (clustered organellar sRNAs) in Arabidopsis. We found that in mitochondria, cosRNAs coincide with transcript 3′-ends, but are largely absent from 5′-ends. In chloroplasts this bias is absent, suggesting a different mode of 5′ processing, possibly owing to different sets of RNases. Furthermore, we identified a large number of cosRNAs that represent silenced insertions of mitochondrial DNA in the nuclear genome of Arabidopsis. Steady-state RNA analyses demonstrate that cosRNAs display differential accumulation during development. Finally, we demonstrate that the chloroplast RBP PPR10 associates in vivo with its cognate cosRNA. A hypothetical role of cosRNAs as competitors of mRNAs for PPR proteins is discussed. PMID:27235415

  11. Multiple feedbacks between chloroplast and whole plant in the context of plant adaptation and acclimation to the environment

    PubMed Central

    Demmig-Adams, Barbara; Stewart, Jared J.; Adams, William W.

    2014-01-01

    This review focuses on feedback pathways that serve to match plant energy acquisition with plant energy utilization, and thereby aid in the optimization of chloroplast and whole-plant function in a given environment. First, the role of source–sink signalling in adjusting photosynthetic capacity (light harvesting, photochemistry and carbon fixation) to meet whole-plant carbohydrate demand is briefly reviewed. Contrasting overall outcomes, i.e. increased plant growth versus plant growth arrest, are described and related to respective contrasting environments that either do or do not present opportunities for plant growth. Next, new insights into chloroplast-generated oxidative signals, and their modulation by specific components of the chloroplast's photoprotective network, are reviewed with respect to their ability to block foliar phloem-loading complexes, and, thereby, affect both plant growth and plant biotic defences. Lastly, carbon export capacity is described as a newly identified tuning point that has been subjected to the evolution of differential responses in plant varieties (ecotypes) and species from different geographical origins with contrasting environmental challenges. PMID:24591724

  12. Systematic analysis of plant mitochondrial and chloroplast small RNAs suggests organelle-specific mRNA stabilization mechanisms.

    PubMed

    Ruwe, Hannes; Wang, Gongwei; Gusewski, Sandra; Schmitz-Linneweber, Christian

    2016-09-01

    Land plant organellar genomes encode a small number of genes, many of which are essential for respiration and photosynthesis. Organellar gene expression is characterized by a multitude of RNA processing events that lead to stable, translatable transcripts. RNA binding proteins (RBPs), have been shown to generate and protect transcript termini and eventually induce the accumulation of short RNA footprints. We applied knowledge of such RBP-derived footprints to develop software (sRNA miner) that enables identification of RBP footprints, or other clusters of small RNAs, in organelles. We used this tool to determine mitochondrial and chloroplast cosRNAs (clustered organellar sRNAs) in Arabidopsis. We found that in mitochondria, cosRNAs coincide with transcript 3'-ends, but are largely absent from 5'-ends. In chloroplasts this bias is absent, suggesting a different mode of 5' processing, possibly owing to different sets of RNases. Furthermore, we identified a large number of cosRNAs that represent silenced insertions of mitochondrial DNA in the nuclear genome of Arabidopsis. Steady-state RNA analyses demonstrate that cosRNAs display differential accumulation during development. Finally, we demonstrate that the chloroplast RBP PPR10 associates in vivo with its cognate cosRNA. A hypothetical role of cosRNAs as competitors of mRNAs for PPR proteins is discussed. PMID:27235415

  13. The complete chloroplast genome of Schrenkiella parvula (Brassicaceae).

    PubMed

    He, Qi; Hao, Guoqian; Wang, Xiaojuan; Bi, Hao; Li, Yuanshuo; Guo, Xinyi; Ma, Tao

    2016-09-01

    Schrenkiella parvula is an Arabidopsis-related model species used here for studying plant stress tolerance. In this study, the complete chloroplast genome sequence of S. parvula has been reported for the first time. The total length of the chloroplast genome was 153 979 bp, which had a typical quadripartite structure. The annotated plastid genome includes 87 protein-coding genes, 39 tRNA genes and 8 ribosomal RNA genes. The evolutionary relationships revealed by our phylogenetic analysis indicated that S. parvula is closer to the Brassiceae species when compared with Eutrema salsugineum. PMID:26260181

  14. Robust expression of a bioactive mammalian protein in chlamydomonas chloroplast

    DOEpatents

    Mayfield, Stephen P.

    2010-03-16

    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 or proteins/peptides, especially gut active proteins, without purification is disclosed.

  15. Robust expression of a bioactive mammalian protein in Chlamydomonas chloroplast

    DOEpatents

    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.

  16. The complete chloroplast genome sequence of Sapindus mukorossi.

    PubMed

    Yang, Bingxian; Li, Mengzhu; Ma, Ji; Fu, Zhengzheng; Xu, Xiaobao; Chen, Qinyi; Zhu, Wei; Tian, Jingkui

    2016-05-01

    The complete chloroplast genome sequence of Sapindus mukorossi, a critical Chinese medicine, was reported here. The total length of the chloroplast genome is 160,481 bp long with 37.7% overall GC content. A pair of IRs (inverted repeats) of 27,979 bp were separated by SSC (18,873 bp) and LSC (85,650 bp). It contains 78 protein-coding genes, 30 tRNA genes and four rRNA genes. Sixteen genes contain one or two introns. PMID:25317637

  17. Circadian oscillations of cytosolic and chloroplastic free calcium in plants

    NASA Technical Reports Server (NTRS)

    Johnson, C. H.; Knight, M. R.; Kondo, T.; Masson, P.; Sedbrook, J.; Haley, A.; Trewavas, A.

    1995-01-01

    Tobacco and Arabidopsis plants, expressing a transgene for the calcium-sensitive luminescent protein apoaequorin, revealed circadian oscillations in free cytosolic calcium that can be phase-shifted by light-dark signals. When apoaequorin was targeted to the chloroplast, circadian chloroplast calcium rhythms were likewise observed after transfer of the seedlings to constant darkness. Circadian oscillations in free calcium concentrations can be expected to control many calcium-dependent enzymes and processes accounting for circadian outputs. Regulation of calcium flux is therefore fundamental to the organization of circadian systems.

  18. Oxidation versus reductive detoxification of SO sub 2 by chloroplasts

    SciTech Connect

    Ghisi, R.; Dittrich, A.P.M.; Heber, U. )

    1990-03-01

    Intact chloroplasts isolated from spinach (Spinacia oleracea L. cv Yates) both oxidized and reduced added sulfite in the light. Oxidation was fast only when endogenous superoxide dismutase was inhibited by cyanide. It was largely suppressed by scavengers of oxygen radicals. After addition of O-acetylserine, chloroplasts reduced sulfite to cysteine and exhibited sulfite-dependent oxygen evolution. Cysteine synthesis from sulfite was faster than from sulfate. The results are discussed in relation to species-specific differences in the phytotoxicity of SO{sub 2}.

  19. A chloroplast retrograde signal regulates nuclear alternative splicing

    PubMed Central

    Petrillo, Ezequiel; Herz, Micaela A. Godoy; Fuchs, Armin; Reifer, Dominik; Fuller, John; Yanovsky, Marcelo J.; Simpson, Craig; Brown, John W. S.; Barta, Andrea; Kalyna, Maria; Kornblihtt, Alberto R.

    2015-01-01

    Light is a source of energy and also a regulator of plant physiological adaptations. We show here that light/dark conditions affect alternative splicing of a subset of Arabidopsis genes preferentially encoding proteins involved in RNA processing. The effect requires functional chloroplasts and is also observed in roots when the communication with the photosynthetic tissues is not interrupted, suggesting that a signaling molecule travels through the plant. Using photosynthetic electron transfer inhibitors with different mechanisms of action we deduce that the reduced pool of plastoquinones initiates a chloroplast retrograde signaling that regulates nuclear alternative splicing and is necessary for proper plant responses to varying light conditions. PMID:24763593

  20. A mammalian cytochrome fused to a chloroplast transit peptide is a functional haemoprotein and is imported into isolated chloroplasts.

    PubMed Central

    Liu, Y Y; Kaderbhai, N; Kaderbhai, M A

    2000-01-01

    The small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) is a major chloroplast stromal protein that is cytosolically synthesized as a precursor with an N-terminal extension, known as the transit sequence or transit peptide (Tp). The Tp is essential for the post-translational uptake of the precursor by the chloroplast. The Tp is thought to influence the conformation of the precursor protein and to facilitate polypeptide translocation across the chloroplast envelope barrier via a Tp-selective translocon. To address these issues we have devised a novel strategy to generate substrate amounts of a chloroplast targeting sequence as a fusion with the chromogenic globular domain of cytochrome b(5) (Cyt). The chimaeric protein is an ideal probe for investigating the conformation of a preprotein and events surrounding protein import into isolated chloroplasts. The Cyt of liver endoplasmic reticulum was fused at its N-terminus with the Tp of the small subunit of Rubisco of Pisum sativum (pea). To enhance its production by clearance from the cytoplasm of Escherichia coli, the chimaera was engineered by further N-terminal linkage of a prokaryotic secretory signal. Expression of this tripartite fusion resulted in mg quantities of the signal sequence-processed Tp-Cyt protein, which was eventually targeted to the membranes. The chromogenic nature of the chimaera and its localization to the bacterial membrane facilitated the biochemical isolation of the precursor in a soluble and functional form. The purified preprotein displayed spectral and enzymic properties that were indistinguishable from the native parental Cyt, implying an absence of observable influence of the Tp on the conformation of the haemoprotein. The chimaeric precursor was imported into the stroma of the isolated chloroplasts in a dose-dependent manner. Import was also strongly dependent upon exogenously supplied ATP. The stromally imported chimaeric precursor protein was processed to a size

  1. Isolation of dimorphic chloroplasts from the single-cell C4 species Bienertia sinuspersici

    PubMed Central

    2012-01-01

    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. PMID:22394490

  2. Ubiquitin facilitates a quality-control pathway that removes damaged chloroplasts

    PubMed Central

    Woodson, Jesse D.; Joens, Matthew S.; Sinson, Andrew B.; Gilkerson, Jonathan; Salomé, Patrice A.; Weigel, Detlef; Fitzpatrick, James A.; Chory, Joanne

    2016-01-01

    Energy production by chloroplasts and mitochondria causes constant oxidative damage. A functioning photosynthetic cell requires quality-control mechanisms to turn over and degrade chloroplasts damaged by reactive oxygen species (ROS). Here, we generated a conditionally lethal Arabidopsis mutant that accumulated excess protoporphyrin IX in the chloroplast and produced singlet oxygen. Damaged chloroplasts were subsequently ubiquitinated and selectively degraded. A genetic screen identified the plant U-box 4 (PUB4) E3 ubiquitin ligase as being necessary for this process. pub4-6 mutants had defects in stress adaptation and longevity. Thus, we have identified a signal that leads to the targeted removal of ROS-overproducing chloroplasts. PMID:26494759

  3. WHITE PANICLE1, a Val-tRNA Synthetase Regulating Chloroplast Ribosome Biogenesis in Rice, Is Essential for Early Chloroplast Development1[OPEN

    PubMed Central

    Wang, Chunming; Zheng, Ming; Lyu, Jia; Xu, Yang; Li, Xiaohui; Niu, Mei; Long, Wuhua; Wang, Di; Wang, Yihua; Wan, Jianmin

    2016-01-01

    Chloroplasts and mitochondria contain their own genomes and transcriptional and translational systems. Establishing these genetic systems is essential for plant growth and development. Here we characterized a mutant form of a Val-tRNA synthetase (OsValRS2) from Oryza sativa that is targeted to both chloroplasts and mitochondria. A single base change in OsValRS2 caused virescent to albino phenotypes in seedlings and white panicles at heading. We therefore named this mutant white panicle 1 (wp1). Chlorophyll autofluorescence observations and transmission electron microscopy analyses indicated that wp1 mutants are defective in early chloroplast development. RNA-seq analysis revealed that expression of nuclear-encoded photosynthetic genes is significantly repressed, while expression of many chloroplast-encoded genes also changed significantly in wp1 mutants. Western-blot analyses of chloroplast-encoded proteins showed that chloroplast protein levels were reduced in wp1 mutants, although mRNA levels of some genes were higher in wp1 than in wild type. We found that wp1 was impaired in chloroplast ribosome biogenesis. Taken together, our results show that OsValRS2 plays an essential role in chloroplast development and regulating chloroplast ribosome biogenesis. PMID:26839129

  4. DipM is required for peptidoglycan hydrolysis during chloroplast division

    PubMed Central

    2014-01-01

    Background Chloroplasts have evolved from a cyanobacterial endosymbiont and their continuity has been maintained over time by chloroplast division, a process which is performed by the constriction of a ring-like division complex at the division site. The division complex has retained certain components of the cyanobacterial division complex, which function inside the chloroplast. It also contains components developed by the host cell, which function outside of the chloroplast and are believed to generate constrictive force from the cytosolic side, at least in red algae and Viridiplantae. In contrast to the chloroplasts in these lineages, those in glaucophyte algae possess a peptidoglycan layer between the two envelope membranes, as do cyanobacteria. Results In this study, we show that chloroplast division in the glaucophyte C. paradoxa does not involve any known chloroplast division proteins of the host eukaryotic origin, but rather, peptidoglycan spitting and probably the outer envelope division process rely on peptidoglycan hydrolyzing activity at the division site by the DipM protein, as in cyanobacterial cell division. In addition, we found that DipM is required for normal chloroplast division in the moss Physcomitrella patens. Conclusions These results suggest that the regulation of peptidoglycan splitting was essential for chloroplast division in the early evolution of chloroplasts and this activity is likely still involved in chloroplast division in Viridiplantae. PMID:24602296

  5. Diversity of a ribonucleoprotein family in tobacco chloroplasts: two new chloroplast ribonucleoproteins and a phylogenetic tree of ten chloroplast RNA-binding domains.

    PubMed Central

    Ye, L H; Li, Y Q; Fukami-Kobayashi, K; Go, M; Konishi, T; Watanabe, A; Sugiura, M

    1991-01-01

    Two new ribonucleoproteins (RNPs) have been identified from a tobacco chloroplast lysate. These two proteins (cp29A and cp29B) are nuclear-encoded and have a less affinity to single-stranded DNA as compared with three other chloroplast RNPs (cp28, cp31 and cp33) previously isolated. DNA sequencing revealed that both contain two consensus sequence-type homologous RNA-binding domains (CS-RBDs) and a very acidic amino-terminal domain but shorter than that of cp28, cp31 and cp33. Comparison of cp29A and cp29B showed a 19 amino acid insertion in the region separating the two CS-RBDs in cp29B. This insertion results in three tandem repeats of a glycine-rich sequence of 10 amino acids, which is a novel feature in RNPs. The two proteins are encoded by different single nuclear genes and no alternatively spliced transcripts could be identified. We constructed a phylogenetic tree for the ten chloroplast CS-RBDs. These results suggest that there is a sizable RNP family in chloroplasts and the diversity was mainly generated through a series of gene duplications rather than through alternative pre-mRNA splicing. The gene for cp29B contains three introns. The first and second introns interrupt the first CS-RBD and the third intron does the second CS-RBD. The position of the first intron site is the same as that in the human hnRNP A1 protein gene. Images PMID:1721701

  6. Photobiotechnology: Algal hydrogen production and photoconductivity of metalized chloroplasts

    SciTech Connect

    Greenbaum, E.

    1991-01-01

    Sustained hydrogen photoevolution from Chlamydomonas reinhardtii and C. moewusii was measured under an anoxic, CO{sub 2}-containing atmosphere. It has been discovered that light intensity and temperature influence the partitioning of reductant between the hydrogen photoevolution pathway and the Calvin cycle. Under low incident light intensity (1-3 W m{sup {minus}2}) or low temperature (approx. O{degrees}C), the flow of photosynthetic reductant to the Calvin cycle was reduced, and reductant was partitioned to the hydrogen pathway as evidenced by sustained H{sub 2} photoevolution. Under saturating light (25 W m{sup {minus}2}) and moderate temperature 20 {plus minus} 5{degrees}C, the Calvin cycle became the absolute sink for reductant with the exception of a burst of H{sub 2} occurring at light on. A novel photobiophysical phenomenon was observed in isolated spinach chloroplasts that were metalized by precipitating colloidal platinum onto the surface of the thylakoid membranes. A two-point irradiation and detection system was constructed in which a continuous beam helium-neon laser ({lambda} = 632.8 nm) was used to irradiate the platinized chloroplasts at varying perpendicular distances from a single linear platinum electrode in pressure contact with the platinized chloroplasts. No external voltage bias was applied to the system. The key objective of the experiments reported in this report was to measure the relative photoconductivity of the chloroplast-metal composite matrix. 46 refs., 1 tab.

  7. Transport Across Chloroplast Membranes: Optimizing Photosynthesis for Adverse Environmental Conditions.

    PubMed

    Pottosin, Igor; Shabala, Sergey

    2016-03-01

    Chloroplasts are central to solar light harvesting and photosynthesis. Optimal chloroplast functioning is vitally dependent on a very intensive traffic of metabolites and ions between the cytosol and stroma, and should be attuned for adverse environmental conditions. This is achieved by an orchestrated regulation of a variety of transport systems located at chloroplast membranes such as porines, solute channels, ion-specific cation and anion channels, and various primary and secondary active transport systems. In this review we describe the molecular nature and functional properties of the inner and outer envelope and thylakoid membrane channels and transporters. We then discuss how their orchestrated regulation affects thylakoid structure, electron transport and excitation energy transfer, proton-motive force partition, ion homeostasis, stromal pH regulation, and volume regulation. We link the activity of key cation and anion transport systems with stress-specific signaling processes in chloroplasts, and discuss how these signals interact with the signals generated in other organelles to optimize the cell performance, with a special emphasis on Ca(2+) and reactive oxygen species signaling. PMID:26597501

  8. Chloroplast microsatellite markers for Artocarpus (Moraceae) developed from transcriptome sequences

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  9. Chloroplast EF-Tu and thermal aggregation of Rubisco activase

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  10. Characterization of a chloroplast inner envelope K+ channel.

    PubMed Central

    Mi, F; Peters, J S; Berkowitz, G A

    1994-01-01

    A K(+)-conducting protein of the chloroplast inner envelope was characterized as a K+ channel. Studies of this transport protein in the native membrane documented its sensitivity to K+ channel blockers. Further studies of native membranes demonstrated a sensitivity of K+ conductance to divalent cations such as Mg2+, which modulate ion conduction through interaction with negative surface charges on the inner-envelope membrane. Purified chloroplast inner-envelope vesicles were fused into an artificial planar lipid bilayer to facilitate recording of single-channel K+ currents. These single-channel K+ currents had a slope conductance of 160 picosiemens. Antibodies generated against the conserved amino acid sequence that serves as a selectivity filter in the pore of K+ channels immunoreacted with a 62-kD polypeptide derived from the chloroplast inner envelope. This polypeptide was fractionated using density gradient centrifugation. Comigration of this immunoreactive polypeptide and K+ channel activity in sucrose density gradients further suggested that this polypeptide is the protein facilitating K+ conductance across the chloroplast inner envelope. PMID:8058841

  11. Structure of "Arabidopsis" chloroplastic monothiol glutaredoxin AtGRXcp

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Monothiol glutaredoxins (Grxs) play important roles in maintaining redox homeostasis in living cells and are conserved across species. "Arabidopsis thaliana" monothiol glutaredoxin AtGRXcp, is critical for protection from oxidative stress in chloroplasts. The crystal structure of AtGRXcp has been de...

  12. The complete chloroplast genome sequence of medicinal plant Pinellia ternata.

    PubMed

    Han, Limin; Chen, Chen; Wang, Bin; Wang, Zhe-Zhi

    2016-07-01

    Pinellia ternata is an important medicinal plant used in the treatment of cough, to dispel phlegm, to calm vomiting and to terminate early pregnancy, as an anti-ulcer and anti-tumor medicine. In this study, we found that the complete chloroplast genome of Pinellia ternata was 164 013 bp in length, containing a pair of inverted repeats of 25 625 bp separated by a large single-copy region and a small single-copy region of 89 783 bp and 22 980 bp, respectively. The chloroplast genome encodes 132 predicted functional genes, including 87 protein-coding genes, eight ribosomal RNA genes, and 37 transfer RNA genes. The chloroplast DNA is GC-rich (36.7%). The phylogenetic analysis showed a strong sister relationship with Colocasia esculenta, which also strongly supports the position of Pinellia ternata. The complete chloroplast genome sequence of Pinellia ternata reported here has the potential to advance population and phylogenetic studies of this medicinal plant. PMID:26153849

  13. Extending the biosynthetic repertoires of cyanobacteria and chloroplasts.

    PubMed

    Nielsen, Agnieszka Zygadlo; Mellor, Silas Busck; Vavitsas, Konstantinos; Wlodarczyk, Artur Jacek; Gnanasekaran, Thiyagarajan; Perestrello Ramos H de Jesus, Maria; King, Brian Christopher; Bakowski, Kamil; Jensen, Poul Erik

    2016-07-01

    Chloroplasts 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 and complex, high-value bioactive molecules. Biosynthesis of all these compounds can be performed in photosynthetic organelles/organisms by heterologous expression of the appropriate pathways, but this requires optimization of carbon flux and reducing power, and a thorough understanding of regulatory pathways. Secretion or storage of the compounds produced can be exploited for the isolation or confinement of the desired compounds. In this review, we explore the use of chloroplasts and cyanobacteria as biosynthetic compartments and hosts, and we estimate the levels of production 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 derived from the photosynthetic light reactions, appears to be non-limiting, but redirection of the fixed carbon via precursor molecules presents a challenge. We also discuss the available synthetic biology tools and the need to expand the molecular toolbox to facilitate cellular reprogramming for increased production yields in both cyanobacteria and chloroplasts. PMID:27005523

  14. Chloroplast heterogeneity and historical admixture within the genus Malus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Premise of the study: We examined chloroplast DNA sequence variation in 412 samples representing 30 Malus species (including Malus x domestica Borkh.). Malus wild species are of particular interest for providing novel alleles and traits in apple breeding programs, yet the taxonomic status of these s...

  15. Efficiency of hydrogen photoproduction by chloroplast-bacterial hydrogenase systems.

    PubMed

    Krasnovsky, A A; Van Ni, C; Nikandrov, V V; Brin, G P

    1980-11-01

    A comparative study of H(2) photoproduction by chloroplasts and solubilized chlorophyll was performed in the presence of hydrogenase preparations of Clostridium butyricum. The photoproduction of H(2) by chloroplasts in the absence of exogenous electron donors, and with irreversibly oxidized dithiothreitol and cysteine, is thought to be limited by a cyclic transport of electrons wherein methylviologen short-circuits the electron transport in photosystem I. The efficiency of H(2) photoproduction by chloroplasts with ascorbate and NADPH is limited by a back reaction between light-reduced methylviologen and the oxidized electron donors. The use of a combination of electron donors (dithiothreitol and ascorbate), providing anaerobiosis without damage to chloroplasts, makes it possible to avoid consumption of reduced methylviologen for the reduction of oxidized electron donors and to exclude the short-circuiting of electron transfer. Under these conditions, photoproduction of H(2) was observed to occur with a rate of 350 to 400 micromoles H(2) per milligram chlorophyll per hour. In this case, the full electron-transferring capability of photosystem I (measured by irreversible photoreduction of methyl red or O(2)) is used to produce H(2). PMID:16661554

  16. Complete Chloroplast Genome Sequence of Phagomixotrophic Green Alga Cymbomonas tetramitiformis

    PubMed Central

    Paasch, Amber E.; Graham, Linda E.; Kim, Eunsoo

    2016-01-01

    We report here the complete chloroplast genome sequence of Cymbomonas tetramitiformis strain PLY262, which is a prasinophycean green alga that retains a phagomixotrophic mode of nutrition. The genome is 84,524 bp in length, with a G+C content of 37%, and contains 3 rRNAs, 26 tRNAs, and 76 protein-coding genes. PMID:27313295

  17. Choline oxidation by intact chloroplasts isolated directly from spinach leaves

    SciTech Connect

    Weigel, P.; Hanson, A.D.

    1986-04-01

    Illuminated chloroplasts derived from spinach leaf protoplasts synthesize betaine from choline via the intermediate betaine aldehyde (BAL) (PNAS 82:3678). Photosynthetically active chloroplasts isolated directly from spinach leaves oxidized (/sup 14/C)choline in the light at rates 10 times higher (25-80 nmol/mg chl b) than protoplast-derived chloroplasts. Up to 20% of the (/sup 14/C)choline supplied during a 30 min incubation was oxidized in the light; the main product was (/sup 14/C)BAL. Rates of (/sup 14/C)choline oxidation in darkness were only 5-30% of rates in light. Light-dependent (/sup 14/C)choline oxidation was abolished by DCMU and 5 mM DTT. Pre-illumination of the chloroplasts did not promote (/sup 14/C)choline oxidation in darkness. The uncouplers nigericin and CCCP at concentrations which eliminated CO/sub 2/-dependent O/sub 2/ evolution did not affect (/sup 14/C)choline oxidation in the light. They hypothesize that (/sup 14/C)choline oxidation is not dependent upon light activation of an enzymatic system or upon the electrochemical proton gradient but requires an oxidant generated in the light.

  18. Choline oxidation by intact spinach chloroplasts. [Spinacia oleracea L

    SciTech Connect

    Weigel, P.; Lerma, C.; Hanson, A.D.

    1988-01-01

    Plants synthesize betaine by a two-step oxidation of choline (choline ..-->.. betaine aldehyde ..-->.. betaine). Protoplast-derived chloroplasts of spinach (Spinacia oleracea L.) carry out both reactions, more rapidly in light than in darkness. We investigated the light-stimulated oxidation of choline, using spinach chloroplasts isolated directly from leaves. The rates of choline oxidation obtained (dark and light rates: 10-50 and 100-300 nanomoles per hour per milligram chlorophyll, respectively) were approximately 20-fold higher than for protoplast-derived chloroplasts. Betaine aldehyde was the main product. Choline oxidation in darkness and light was suppressed by hypoxia. Neither uncouplers not the Calvin cycle inhibitor glyceraldehyde greatly affected choline oxidation in the light, and maximal choline oxidation was attained far below light saturation of CO/sub 2/ fixation. The light stimulation of choline oxidation was abolished by the PSII inhibitors DCMU and dibromothymoquinone, and was partially restored by adding reduced diaminodurene, an electron donor to PSI. Both methyl viologen and phenazine methosulfate prevented choline oxidation. Adding dihydroxyacetone phosphate, which can generate NADPH in organello, doubled the dark rate of choline oxidation. These results indicate that choline oxidation in chloroplasts requires oxygen, and reducing power generated from PSI. Enzymic reactions consistent with these requirements are discussed.

  19. Complete Chloroplast Genome Sequence of Phagomixotrophic Green Alga Cymbomonas tetramitiformis.

    PubMed

    Satjarak, Anchittha; Paasch, Amber E; Graham, Linda E; Kim, Eunsoo

    2016-01-01

    We report here the complete chloroplast genome sequence of Cymbomonas tetramitiformis strain PLY262, which is a prasinophycean green alga that retains a phagomixotrophic mode of nutrition. The genome is 84,524 bp in length, with a G+C content of 37%, and contains 3 rRNAs, 26 tRNAs, and 76 protein-coding genes. PMID:27313295

  20. ULTRAVIOLET INACTIVATION OF CHLOROPLAST FORMATION IN SYNCHRONOUSLY DIVIDING EUGLENA GRACILIS.

    PubMed

    PETROPULOS, S F

    1964-07-24

    Ultraviolet inactivation of chloroplast formation was studied in synchronously dividing cultures of Euglena gracilis. Sensitivity to sublethal doses given at intervals throughout the cell cycle was greater just before cell division than during division. There was approximately a twofold difference in the doseresponse relationships for the periods of high and low sensitivity. PMID:14172598

  1. Senescence-Associated Vacuoles, a Specific Lytic Compartment for Degradation of Chloroplast Proteins?

    PubMed Central

    Carrión, Cristian A.; Martínez, Dana E.; Costa, M. Lorenza; Guiamet, Juan José

    2014-01-01

    Degradation of chloroplasts and chloroplast components is a distinctive feature of leaf senescence. In spite of its importance in the nutrient economy of plants, knowledge about the mechanism(s) involved in the breakdown of chloroplast proteins is incomplete. A novel class of vacuoles, “senescence-associated vacuoles” (SAVs), characterized by intense proteolytic activity appear during senescence in chloroplast-containing cells of leaves. Since SAVs contain some chloroplast proteins, they are candidate organelles to participate in chloroplast breakdown. In this review we discuss the characteristics of SAVs, and their possible involvement in the degradation of Rubisco, the most abundant chloroplast protein. Finally, SAVs are compared with other extra-plastidial protein degradation pathways operating in senescing leaves. PMID:27135516

  2. Evolution of the Cp-Actin-based Motility System of Chloroplasts in Green Plants.

    PubMed

    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. PMID:27200035

  3. Evolution of the Cp-Actin-based Motility System of Chloroplasts in Green Plants

    PubMed Central

    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. PMID:27200035

  4. Reinvestigation of the triplet-minus-singlet spectrum of chloroplasts

    NASA Astrophysics Data System (ADS)

    Jávorfi, T.; Garab, G.; Razi Naqvi, K.

    2000-01-01

    A comparison of the triplet-minus-singlet (TmS) absorption spectrum of spinach chloroplasts, recorded some thirty years ago, with the more recently published TmS spectrum of isolated Chl a/ b LHCII (light-harvesting complexes associated with photosystem II of higher plants) shows that the two spectra are very similar, which is to be expected, since only the carotenoid pigments contribute to each spectrum. Be that as it may, the comparison also reveals a dissimilarity: photoexcitation of the sample does, or does not, affect the absorbance in the Qy region (650-700 nm), depending on whether the sample is a suspension of chloroplasts or of isolated LHCII. The Qy-signal in the TmS spectrum of LHCII decays, it should be noted, at the same rate as the rest of the difference spectrum, and its most prominent feature is a negative peak. As the carotenoids do not absorb in the Qy region, the presence of a signal in this region calls for an explanation: van der Vos, Carbonera and Hoff, the first to find as well as fathom the phenomenon, attributed the Qy-signal to a change, in the absorption spectrum of a chlorophyll a (Chl a) molecule, brought about by the presence of triplet excitation on a neighbouring carotenoid (Car). The difference in the behaviours of chloroplasts and LHCII, if reproducible, would imply that the Car triplets which give rise to the TmS spectrum of chloroplasts do not influence the absorption spectra of their Chl a neighbours. With a view to reaching a firm conclusion about this vexed issue, spinach chloroplasts and thylakoids have been examined with the aid of the same kinetic spectrometer as that used for investigating LHCII; the TmS spectra of both chloroplasts and thylakoids contain prominent bleaching signals centred at 680 nm, and the triplet decay time in each case is comparable to that of the Chl a/ b LHCII triplets. Results pertaining to other closely related systems are recalled, and it is concluded that, so far as the overall appearance of the

  5. Glucose respiration in the intact chloroplast of Chlamydomonas reinhardtii

    SciTech Connect

    Changguo Chen; Gibbs, M. )

    1991-01-01

    Chloroplastic respiration was monitored by measuring {sup 14}CO{sub 2} from {sup 14}C glucose in the darkened Chlamydomonas reinhardtii F-60 chloroplast, The patterns of {sup 14}CO{sub 2} evolution from labeled glucose in the absence and presence of the inhibitors iodoacetamide, glycolate-2-phosphate, and phosphoenolypyruvate were those expected from the oxidative pentose phosphate cycle and glycolysis. The K{sub m} for glucose was 56 micromolar and for MgATP was 200 micromolar. Release of {sup 14}CO{sub 2} was inhibited by phloretin and inorganic phosphate. Comparing the inhibition of CO{sub 2} evolution generated by pH 7.5 with respect to pH 8.2 (optimum) in chloroplasts given C-1, C-2, and C-6 labeled glucose indicated that a suboptimum pH affects the recycling of the pentose phosphate intermediates to a greater extent than CO{sub 2} evolution from C-1 of glucose. Respiratory inhibition by pH 7.5 in the darkened chloroplast was alleviated by NH{sub 4}Cl and KCl (stromal alkalating agents), iodoacetamide (an inhibitor of glyceraldehyde 3-phosphate dehydrogenase), or phosphoenolypyruvate (an inhibitor of phosphofructokinase). It is concluded that the site which primarily mediates respiration in the darkened Chlamydomonas chloroplast is the fructose-1,6-bisphosphatase/phosphofructokinase junction. The respiratory pathways described here can account for the total oxidation of a hexose to Co{sub 2} and for interactions between carbohydrate metabolism and the oxyhydrogen reaction in algal cells adapted to a hydrogen metabolism.

  6. Possible function of VIPP1 in maintaining chloroplast membranes.

    PubMed

    Zhang, Lingang; Sakamoto, Wataru

    2015-09-01

    A protein designated as VIPP1 is found widely in organisms performing oxygenic photosynthesis, but its precise role in chloroplasts has remained somewhat mysterious. Based on its structural similarity, it presumably has evolved from bacterial Phage shock protein A (PspA) with a C-terminal extension of approximately 40 amino acids. Both VIPP1 and PspA are membrane-associated despite the lack of transmembrane helices. They form an extremely large homo-complex that consists of an oligomeric ring unit. Although PspA is known to respond to membrane stress and although it acts in maintaining proton motive force through membrane repair, the multiple function of VIPP1, such as vesicle budding from inner envelope to deliver lipids to thylakoids, maintenance of photosynthetic complexes in thylakoid membranes, biogenesis of Photosystem I, and protective role of inner envelope against osmotic stress, has been proposed. Whatever its precise function in chloroplasts, it is an important protein because depletion of VIPP1 in mutants severely affects photoautotrophic growth. Recent reports of the relevant literature describe that VIPP1 becomes highly mobile when chloroplasts receive hypotonic stress, and that VIPP1 is tightly bound to lipids, which implies a crucial role of VIPP1 in membrane repair through lipid transfer. This review presents a summary of our current knowledge related to VIPP1, particularly addressing the dynamic behavior of complexes against stress and its property of lipid binding. Those data altogether suggest that VIPP1 acts a priori in chloroplast membrane maintenance through its activity to transfer lipids rather than in thylakoid formation through vesicles. This article is part of a Special Issue titled: Chloroplast Biogenesis. PMID:25725437

  7. Differential responses of seabirds to inter-annual environmental change in the continental shelf and oceanic habitats of southeastern Bering Sea

    NASA Astrophysics Data System (ADS)

    Yamamoto, T.; Kokubun, N.; Kikuchi, D. M.; Sato, N.; Takahashi, A.; Will, A.; Kitaysky, A. S.; Watanuki, Y.

    2015-11-01

    Seasonal sea-ice cover has been decreasing in the southeastern Bering Sea shelf, which might affect ecosystem dynamics and availability of food resources to marine top predators breeding in the region. In this study, we investigated the foraging responses of two seabird species, surface-foraging red-legged kittiwakes Rissa brevirostris (hereafter, RLKI) and pursuit-diving foraging thick-billed murres Uria lomvia (TBMU) to the inter-annual change in environmental conditions. Between the study years, winter ice retreated earlier and summer water temperatures were warmer in 2014 compared to those in 2013. At-sea distributions of RLKI and TBMU breeding on St. George Island, the largest seabird colony in the region, were recorded using GPS loggers, and blood samples were taken to examine their physiological condition and isotopic foraging niche in a given year. RLKI foraging occurred mostly over the oceanic basin in both years. TBMU, however, foraged mostly over the shelf, but showed a relatively higher use of the shelf break and oceanic basin in the colder year, 2013. The foraging distances from the colony peaked at 250-300 km in 2013 and, bimodally, at 150-250 and 300-350 km in 2014 for RLKI, and tended to be farther in 2013 compared to those in 2014 for TBMU. Plasma levels of corticosterone did not differ between years in RLKI, but differed in TBMU, showing higher levels of physiological stress incurred by murres during the colder year, 2013. δ13N (a proxy of trophic level of prey) did not differ between the years in either RLKI or TBMU, while δ13C (a proxy of prey origin) were lower in 2014 than in 2013 in both species, suggesting possible differences in influx of oceanic prey items into foraging areas. These results suggest that the response of ecosystem dynamics to climate variability in the southeast Bering Sea may differ between the ocean basin and continental shelf regions, which, in turn, may generate differential responses in seabirds relying on those

  8. Genetic variation and species identification of Thai Boesenbergia (Zingiberaceae) analyzed by chloroplast DNA polymorphism.

    PubMed

    Techaprasan, Jiranan; Ngamriabsakul, Chatchai; Klinbunga, Sirawut; Chusacultanachai, Sudsanguan; Jenjittikul, Thaya

    2006-07-31

    Genetic variation and molecular phylogeny of 22 taxa representing 14 extant species and 3 unidentified taxa of Boesenbergia in Thailand and four outgroup species (Cornukaempferia aurantiflora, Hedychium biflorum, Kaempferia parviflora, and Scaphochlamys rubescens) were examined by sequencing of 3 chloroplast (cp) DNA regions (matK, psbA-trnH and petA-psbJ). Low interspecific genetic divergence (0.25-1.74%) were observed in these investigated taxa. The 50% majority-rule consensus tree constructed from combined chloroplast DNA sequences allocated Boesenbergia in this study into 3 different groups. Using psbA-1F/psbA-3R primers, an insertion of 491 bp was observed in B. petiolata. Restriction analysis of the amplicon (380-410 bp) from the remaining species with Rsa I further differentiated Boesenbergia to 2 groupings; I (B. basispicata, B. longiflora, B. longipes, B. plicata, B.pulcherrima, B. tenuispicata, B. thorelii, B. xiphostachya, Boesenbergia sp.1 and Boesenbergia sp.3; phylogenetic clade A) that possesses a Rsa I restriction site and II (B.curtisii, B. regalis, B. rotunda and Boesenbergia sp.2; phylogenetic clade B and B. siamensis; phylogenetic clade C) that lacks a restriction site of Rsa I. Single nucleotide polymorphism (SNP) and indels found can be unambiguously applied to authenticate specie-origin of all investigated samples and revealed that Boesenbergia sp.1, Boesenbergia sp.2 and B. pulcherrima (Mahidol University, Kanchanaburi), B. cf. pulcherrima1 (Prachuap Khiri Khan) and B. cf. pulcherrima2 (Thong Pha Phum, Kanchanaburi) are B. plicata, B. rotunda and B. pulcherrima, respectively. In addition, molecular data also suggested that Boesenbergia sp.3 should be further differentiated from B. longiflora and regarded as a newly unidentified Boesenbergia species. PMID:16889678

  9. Chloroplast Genome Sequence of the Moss Tortula ruralis: Gene Content and Structural Arrangement Relative to Other Green Plant Chloroplast Genomes

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Tortula ruralis, a widely distributed moss species in the family Pottiaceae, is increasingly being 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 Tortula ruralis, only the second publishe...

  10. Redirecting the Cyanobacterial Bicarbonate Transporters BicA and SbtA to the Chloroplast Envelope: Soluble and Membrane Cargos Need Different Chloroplast Targeting Signals in Plants

    PubMed Central

    Rolland, Vivien; Badger, Murray R.; Price, G. Dean

    2016-01-01

    Most major crops used for human consumption are C3 plants, which yields are limited by photosynthetic inefficiency. To circumvent this, it has been proposed to implement the cyanobacterial CO2-concentrating mechanism (CCM), principally consisting of bicarbonate transporters and carboxysomes, into plant chloroplasts. As it is currently not possible to recover homoplasmic transplastomic monocots, foreign genes must be introduced in these plants via nuclear transformation. Consequently, it is paramount to ensure that resulting proteins reach the appropriate sub-cellular compartment, which for cyanobacterial transporters BicA and SbtA, is the chloroplast inner-envelope membrane (IEM). At present, targeting signals to redirect large transmembrane proteins from non-chloroplastic organisms to plant chloroplast envelopes are unknown. The goal of this study was to identify such signals, using agrobacteria-mediated transient expression and confocal microscopy to determine the sub-cellular localization of ∼37 GFP-tagged chimeras. Initially, fragments of chloroplast proteins known to target soluble cargos to the stroma were tested for their ability to redirect BicA, but they proved ineffective. Next, different N-terminal regions from Arabidopsis IEM transporters were tested. We demonstrated that the N-terminus of AtHP59, AtPLGG1 or AtNTT1 (92–115 amino acids), containing a cleavable chloroplast transit peptide (cTP) and a membrane protein leader (MPL), was sufficient to redirect BicA or SbtA to the chloroplast envelope. This constitutes the first evidence that nuclear-encoded transmembrane proteins from non-chloroplastic organisms can be targeted to the envelope of plant chloroplasts; a finding which represents an important advance in chloroplast engineering by opening up the door to further manipulation of the chloroplastic envelope. PMID:26973659

  11. Redirecting the Cyanobacterial Bicarbonate Transporters BicA and SbtA to the Chloroplast Envelope: Soluble and Membrane Cargos Need Different Chloroplast Targeting Signals in Plants.

    PubMed

    Rolland, Vivien; Badger, Murray R; Price, G Dean

    2016-01-01

    Most major crops used for human consumption are C3 plants, which yields are limited by photosynthetic inefficiency. To circumvent this, it has been proposed to implement the cyanobacterial CO2-concentrating mechanism (CCM), principally consisting of bicarbonate transporters and carboxysomes, into plant chloroplasts. As it is currently not possible to recover homoplasmic transplastomic monocots, foreign genes must be introduced in these plants via nuclear transformation. Consequently, it is paramount to ensure that resulting proteins reach the appropriate sub-cellular compartment, which for cyanobacterial transporters BicA and SbtA, is the chloroplast inner-envelope membrane (IEM). At present, targeting signals to redirect large transmembrane proteins from non-chloroplastic organisms to plant chloroplast envelopes are unknown. The goal of this study was to identify such signals, using agrobacteria-mediated transient expression and confocal microscopy to determine the sub-cellular localization of ∼37 GFP-tagged chimeras. Initially, fragments of chloroplast proteins known to target soluble cargos to the stroma were tested for their ability to redirect BicA, but they proved ineffective. Next, different N-terminal regions from Arabidopsis IEM transporters were tested. We demonstrated that the N-terminus of AtHP59, AtPLGG1 or AtNTT1 (92-115 amino acids), containing a cleavable chloroplast transit peptide (cTP) and a membrane protein leader (MPL), was sufficient to redirect BicA or SbtA to the chloroplast envelope. This constitutes the first evidence that nuclear-encoded transmembrane proteins from non-chloroplastic organisms can be targeted to the envelope of plant chloroplasts; a finding which represents an important advance in chloroplast engineering by opening up the door to further manipulation of the chloroplastic envelope. PMID:26973659

  12. Pb-Induced Avoidance-Like Chloroplast Movements in Fronds of Lemna trisulca L.

    PubMed Central

    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

  13. Pb-induced avoidance-like chloroplast movements in fronds of Lemna trisulca L.

    PubMed

    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

  14. Myrionecta Rubra Population Genetic Diversity and Its Cryptophyte Chloroplast Specificity in Recurrent Red Tides in the Columbia River Estuary

    SciTech Connect

    Herfort, Lydie; Peterson, Tawnya D.; McCue, Lee Ann; Crump, Byron C.; Prahl, Fredrick G.; Baptista, Antonio M.; Campbell, Victoria; Warnick, Rachel; Selby, Mikaela; Roegner, G. Curtis; Zuber, Peter A.

    2011-01-04

    For at least a decade, annually recurring blooms of the photosynthetic ciliate, Myrionecta rubra have been observed in the Columbia River estuary in late summer. In an effort to understand the dynamics of these blooms, we investigated the genetic variability of M. rubra and its cryptophyte plastids within three large estuarine blooms formed in consecutive years (2007-2009), and conducted a broader spatial survey along the coasts of Oregon/Washington. Analysis of the ‘18S-28S’ sequences specific for Mesodiniidae uncovered at least 7 variants of M. rubra within the Columbia River coastal margin in spring and summer, but only one of these M. rubra variants was implicated in estuary bloom formation. Using a multigene approach, we show that the bloom-forming variant of M. rubra appears to harbor the same cryptophyte chloroplast in recurring blooms. Analyses of chloroplast 16S rRNA, cryptophyte RuBisCO and Photosystem II D2 genes together suggest that the plastid is derived from Teleaulax amphioxeia. Free-living cells of this species and of other cryptophytes were practically absent from the bloom patches in the estuary main channels based on 18S rDNA sequence analyses. The respectively low and high proportions of T. amphioxeia nuclei and chloroplasts signals found in the M. rubra bloom of the Columbia River estuary in successive years supports the notion of a transient association between T. amphioxeia and the bloom-forming M. rubra variant, with loss of cryptophyte nuclei. The genetic variability of M. rubra uncovered here is relevant to the controversy in the literature regarding the cryptophyte /M. rubra association.

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

    PubMed

    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. Learning the Languages of the Chloroplast: Retrograde Signaling and Beyond.

    PubMed

    Chan, Kai Xun; Phua, Su Yin; Crisp, Peter; McQuinn, Ryan; Pogson, Barry J

    2016-04-29

    The chloroplast can act as an environmental sensor, communicating with the cell during biogenesis and operation to change the expression of thousands of proteins. This process, termed retrograde signaling, regulates expression in response to developmental cues and stresses that affect photosynthesis and yield. Recent advances have identified many signals and pathways-including carotenoid derivatives, isoprenes, phosphoadenosines, tetrapyrroles, and heme, together with reactive oxygen species and proteins-that build a communication network to regulate gene expression, RNA turnover, and splicing. However, retrograde signaling pathways have been viewed largely as a means of bilateral communication between organelles and nuclei, ignoring their potential to interact with hormone signaling and the cell as a whole to regulate plant form and function. Here, we discuss new findings on the processes by which organelle communication is initiated, transmitted, and perceived, not only to regulate chloroplastic processes but also to intersect with cellular signaling and alter physiological responses. PMID:26735063

  17. The complete chloroplast genome sequence of Safflower (Carthamus tinctorius L.).

    PubMed

    Lu, Chaolong; Shen, Qi; Yang, Jun; Wang, Bo; Song, Chi

    2016-09-01

    Safflower (Carthamus tinctorius L.) is a traditional medical plants of Asia. In this study, the complete chloroplast genome of safflower was presented. The total genome size was 153,675 bp in length, containing a pair of inverted repeats (IRs) of 25,407 bp, separated by large single copy (LSC) and small single copy (SSC) of 83,606 bp and 19,156 bp, respectively. Overall GC content of the genome was 37.4%. The chloroplast genome harbored 127 annotated genes, including 89 protein coding genes, 30 tRNA genes and 8 rRNA genes. A total of 7 of these genes were duplicated in the inverted repeat regions. Twelve genes contained one intron. PMID:25740214

  18. The complete chloroplast genome of Torreya fargesii (Taxaceae).

    PubMed

    Tao, Ke; Gao, Lei; Li, Jia; Chen, Shanshan; Su, Yingjuan; Wang, Ting

    2016-09-01

    The complete chloroplast genome sequence of Torreya fargesii (Taxaceae), a relic plant endemic to China, is presented in this study. The genome is 137 075 bp in length, with 35.47% average GC content. One copy of the large inverted repeats is lost from this genome. The T. fargesii chloroplast genome encodes 118 unique genes, in which trnI-CAU, trnQ-UUG, trnN-GUU are duplicated. Protein-coding, tRNA and rRNA genes represent 54.7%, 1.9% and 3.4% of the genome, respectively. There are 17 intron-containing genes, of which 6 are tRNA genes. A maximum likelihood phylogenetic analysis revealed a strong sister relationship between Torreya and Amentotaxus. PMID:27158868

  19. The complete chloroplast genome of North American ginseng, Panax quinquefolius.

    PubMed

    Han, Zeng-Jie; Li, Wei; Liu, Yuan; Gao, Li-Zhi

    2016-09-01

    We report complete nucleotide sequence of the Panax quinquefolius chloroplast genome using next-generation sequencing technology. The genome size is 156 359 bp, including two inverted repeats (IRs) of 52 153 bp, separated by the large single-copy (LSC 86 184 bp) and small single-copy (SSC 18 081 bp) regions. This cp genome encodes 114 unigenes (80 protein-coding genes, four rRNA genes, and 30 tRNA genes), in which 18 are duplicated in the IR regions. Overall GC content of the genome is 38.08%. A phylogenomic analysis of the 10 complete chloroplast genomes from Araliaceae using Daucus carota from Apiaceae as outgroup showed that P. quinquefolius is closely related to the other two members of the genus Panax, P. ginseng and P. notoginseng. PMID:27158867

  20. The complete chloroplast genome sequence of Amentotaxus argotaenia (Taxaceae).

    PubMed

    Li, Jia; Gao, Lei; Tao, Ke; Su, Yingjuan; Wang, Ting

    2016-07-01

    The complete chloroplast genome sequence of Amentotaxus argotaenia was determined in this study. The genome is 136 657 bp in length and lacks one inverted repeat region. The overall GC content of the genome is 35.85% (protein-coding genes, 36.90%; tRNA genes, 53.31%; rRNA genes, 52.99%; introns, 36.10%; intergenic spacers, 31.03%). The A. argotaenia chloroplast genome contains 118 unique genes, including 83 protein-coding genes, 31 tRNA genes, and four rRNA genes. Ten protein-coding genes and six tRNA genes have one intron, while ycf3 contains two. The coding regions occupy 60.27% of the genome length and the gene density is estimated to be 0.88 genes/kb. A maximum likelihood phylogenetic analysis suggested that Amentotaxus is sister to Taxus within the Taxaceae family. PMID:26119122

  1. The whole chloroplast genome of shrub willows (Salix suchowensis).

    PubMed

    Wu, Zhiqiang

    2016-05-01

    The whole chloroplast genome of Salix suchowensis was determined in this study using next generation sequencing data. The total genome size was 155,214 bp in length, containing a pair of inverted repeats (IRs) of 27,459 bp, which were separated by large single copy (LSC) and small single copy (SSC) of 84,077 bp and 16,219 bp, respectively. The overall GC contents of the chloroplast genome were 36.73%. One hundred and ten unique genes were annotated, including 76 protein coding genes, 30 tRNA genes and 4 rRNA genes. Among these, 18 are duplicated in the inverted repeat regions, 14 genes contained 1 intron, and 3 genes (rps12, clpP and ycf3) comprised of 2 introns. PMID:25418623

  2. Toxic effects of copper on photosystem II of spinach chloroplasts

    SciTech Connect

    Hsu, Bandar; Lee, Jeeyau )

    1988-05-01

    The room temperature fluorescence induction of chloroplasts was utilized as a probe to locate the site of inhibition on PSII by copper. It was found that, while the initial fluorescence yield was hardly affected, the variable fluorescence yield was lowered without significant change in its kinetics. Addition of DCMU, or abolishing oxygen evolution capability by Tris treatment, did not alter this basic inhibition pattern. Copper was also found to lower the fluorescence yield of chloroplasts treated with linolenic acid which inhibited the secondary electron transport on both oxidizing and reducing sides of PSII. The data indicate that copper adversely affects the primary change separation at the PSII reaction center. We suggest that the inhibition is due to creation of a lesion close to the reaction center, leading to increased dissipation of incoming excitation energy to heat.

  3. Chloroplast and nuclear microsatellite analysis of Aegilops cylindrica.

    PubMed

    Gandhi, Harish T; Vales, M Isabel; Watson, Christy J W; Mallory-Smith, Carol A; Mori, Naoki; Rehman, Maqsood; Zemetra, Robert S; Riera-Lizarazu, Oscar

    2005-08-01

    Aegilops cylindrica Host (2n = 4x = 28, genome CCDD) is an allotetraploid formed by hybridization between the diploid species Ae. tauschii Coss. (2n = 2x = 14, genome DD) and Ae. markgrafii (Greuter) Hammer (2n = 2x = 14, genome CC). Previous research has shown that Ae. tauschii contributed its cytoplasm to Ae. cylindrica. However, our analysis with chloroplast microsatellite markers showed that 1 of the 36 Ae. cylindrica accessions studied, TK 116 (PI 486249), had a plastome derived from Ae. markgrafii rather than Ae. tauschii. Thus, Ae. markgrafii has also contributed its cytoplasm to Ae. cylindrica. Our analysis of chloroplast and nuclear microsatellite markers also suggests that D-type plastome and the D genome in Ae. cylindrica were closely related to, and were probably derived from, the tauschii gene pool of Ae. tauschii. A determination of the likely source of the C genome and the C-type plastome in Ae. cylindrica was not possible. PMID:15986256

  4. The complete chloroplast genome sequence of Fagopyrum cymosum.

    PubMed

    Yang, Jun; Lu, Chaolong; Shen, Qi; Yan, Yuying; Xu, Changjiang; Song, Chi

    2016-07-01

    Fagopyrum cymosum is a traditional medicinal plant. In this study, the complete chloroplast genome of Fagopyrum cymosum is presented. The total genome size is 160,546 bp in length, containing a pair of inverted repeats (IRs) of 32,598 bp, separated by large single copy (LSC) and small single copy (SSC) of 84,237 bp and 11,014 bp, respectively. Overall GC contents of the genome were 36.9%. The chloroplast genome harbors 126 annotated genes, including 91 protein coding genes, 29 tRNA genes, and six rRNA genes. Eighteen genes contain one or two introns. Phylogenetic analyses indicated a clear evolutionary relationship among species of Caryophyllales. PMID:26119127

  5. Chloroplast Structure and Function Is Altered in the NCS2 Maize Mitochondrial Mutant 1

    PubMed Central

    Roussell, Deborah L.; Thompson, Deborah L.; Pallardy, Steve G.; Miles, Donald; Newton, Kathleen J.

    1991-01-01

    The nonchromosomal stripe 2 (NCS2) mutant of maize (Zea mays L.) has a DNA rearrangement in the mitochondrial genome that segregates with the abnormal growth phenotype. Yet, the NCS2 characteristic phenotype includes striped sectors of pale-green tissue on the leaves. This suggests a chloroplast abnormality. To characterize the chloroplasts present in the mutant sectors, we examined the chloroplast structure by electron microscopy, chloroplast function by radiolabeled carbon dioxide fixation and fluorescence induction kinetics, and thylakoid protein composition by polyacrylamide gel electrophoresis. The data from these analyses suggest abnormal or prematurely arrested chloroplast development. Deleterious effects of the NCS2 mutant mitochondria upon the cells of the leaf include structural and functional alterations in the both the bundle sheath and mesophyll chloroplasts. ImagesFigure 1Figure 2Figure 3Figure 5Figure 6 PMID:16668157

  6. Oxygen Evolution and the Permeability of the Outer Envelope of Isolated Whole Chloroplasts 1

    PubMed Central

    Robinson, J. Michael; Stocking, C. R.

    1968-01-01

    A rapid oxygraph method of studying the permeability of the envelope of isolated chloroplasts was used. The outer envelope of aqueously isolated whole spinach (Spinacia oleracea L.) chloroplasts in buffer is readily permeable to 3-phosphoglyceric acid, which induces an immediate light dependent oxygen evolution. This light dependent oxygen evolution was completely eliminated by swelling these plastids in an osmotically dilute solution. Exogenous adenosine diphosphate, but not inorganic phosphate, strongly stimulated this oxygen evolution. This indicated that the chloroplast envelope is relatively permeable to adenosine diphosphate. Oxygen evolution and swelling studies indicated that the chloroplast envelope is relatively impermeable to NADP and to ferredoxin. A method is described whereby the percent of whole chloroplasts present in a chloroplast preparation may be rapidly estimated. PMID:16656943

  7. Chloroplast Distribution in Arabidopsis thaliana (L.) Depends on Light Conditions during Growth.

    PubMed Central

    Trojan, A.; Gabrys, H.

    1996-01-01

    Chloroplasts of Arabidopsis thaliana move in response to blue light. Sensitivity to light and the range of fluence rates to which the chloroplasts respond were found to be comparable to those of other higher plants studied. We investigated typical chloroplast distributions in Arabidopsis grown under three different light conditions:standard-light conditions, similar to natural light intensities; weak-light intensities, close to the compensation point of photosynthesis; and strong-light intensities, close to the saturation of the light-response curve of photosynthesis. We observed a striking difference in chloroplast arrangement in darkness between plants grown under weak- and strong-light conditions. There was a slight difference after weak-light pretreatment, and the arrangements of chloroplasts after strong-light pretreatment in both plant groups were very similar. These results support the ecological significance of chloroplast movements. PMID:12226297

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

    SciTech Connect

    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.

  9. Structural organization of photosynthetic apparatus in agranal chloroplasts of maize.

    PubMed

    Romanowska, Elzbieta; Kargul, Joanna; Powikrowska, Marta; Finazzi, Giovanni; Nield, Jon; Drozak, Anna; Pokorska, Berenika

    2008-09-19

    We investigated the organization of photosystem II (PSII) in agranal bundle sheath thylakoids from a C(4) plant maize. Using blue native/SDS-PAGE and single particle analysis, we show for the first time that PSII in the bundle sheath (BS) chloroplasts exists in a dimeric form and forms light-harvesting complex II (LHCII).PSII supercomplexes. We also demonstrate that a similar set of photosynthetic membrane complexes exists in mesophyll and agranal BS chloroplasts, including intact LHCI.PSI supercomplexes, PSI monomers, PSII core dimers, PSII monomers devoid of CP43, LHCII trimers, LHCII monomers, ATP synthase, and cytochrome b(6)f complex. Fluorescence functional measurements clearly indicate that BS chloroplasts contain PSII complexes that are capable of performing charge separation and are efficiently sensitized by the associated LHCII. We identified a fraction of LHCII present within BS thylakoids that is weakly energetically coupled to the PSII reaction center; however, the majority of BS LHCII is shown to be tightly connected to PSII. Overall, we demonstrate that organization of the photosynthetic apparatus in BS agranal chloroplasts of a model C(4) plant is clearly distinct from that of the stroma lamellae of the C(3) plants. In particular, supramolecular organization of the dimeric LHCII.PSII in the BS thylakoids strongly suggests that PSII in the BS agranal membranes may donate electrons to PSI. We propose that the residual PSII activity may supply electrons to poise cyclic electron flow around PSI and prevent PSI overoxidation, which is essential for the CO(2) fixation in BS cells, and hence, may optimize ATP production within this compartment. PMID:18632664

  10. Wage Equity and Female Faculty Job-Satisfaction: The Role of Wage Differentials in a Job Satisfaction Causal Model. ASHE Annual Meeting Paper.

    ERIC Educational Resources Information Center

    Hagedorn, Linda Serra

    This study examined the role of female/male wage differentials in a model of job satisfaction. It is based on data from 5,021 respondents to the 1989 Carnegie Foundation for the Advancement of Teaching national faculty survey. The model considers the interrelated effects of the calculated wage differential, stress, social perceptions of students,…

  11. Chloroplast Retrograde Regulation of Heat Stress Responses in Plants.

    PubMed

    Sun, Ai-Zhen; Guo, Fang-Qing

    2016-01-01

    It is well known that intracellular signaling from chloroplast to nucleus plays a vital role in stress responses to survive environmental perturbations. The chloroplasts were proposed as sensors to heat stress since components of the photosynthetic apparatus housed in the chloroplast are the major targets of thermal damage in plants. Thus, communicating subcellular perturbations to the nucleus is critical during exposure to extreme environmental conditions such as heat stress. By coordinating expression of stress specific nuclear genes essential for adaptive responses to hostile environment, plants optimize different cell functions and activate acclimation responses through retrograde signaling pathways. The efficient communication between plastids and the nucleus is highly required for such diverse metabolic and biosynthetic functions during adaptation processes to environmental stresses. In recent years, several putative retrograde signals released from plastids that regulate nuclear genes have been identified and signaling pathways have been proposed. In this review, we provide an update on retrograde signals derived from tetrapyrroles, carotenoids, reactive oxygen species (ROS) and organellar gene expression (OGE) in the context of heat stress responses and address their roles in retrograde regulation of heat-responsive gene expression, systemic acquired acclimation, and cellular coordination in plants. PMID:27066042

  12. [Antenna replacement in the evolutionary origin of chloroplasts].

    PubMed

    Stadnichuk, I N; Tropin, I V

    2014-01-01

    Endosymbiotic origin of chloroplasts from unicellular cyanobacteria is presently beyond doubt. Oxygenic photosynthesis is based on coordinated action of two photosystems (PS), PS I and PS II, cooperating with several variants of the pigment antenna. In cyanobacteria, red algae, and glaucophytes, phycobilisomes (PBS) act as antennae, while in terrestrial plants, as well as most macro- and microalgae antennae are formed by chlorophyll a/b- and chlorophyll a/c-containing proteins. Advantages and disadvantages of the PBS antenna compared to other light-gathering complexes form the basis for adaptive variations of the antenna in the course of development of eukaryotic photosynthesis. During the evolution of the "green" and "chromophyte" lineages of the chloroplasts, PBS, in spite of their optimal features of light absorption,were replaced by chlorophyll a/b- and chlorophyll a/c-containing light-gathering complexes. Development of the cell wall associated with limited motility and with tissue formation in photosynthetic eukaryotes were the factors responsible for the antenna shift. The subsequent redistribution of cell resources in favor of cellulose biosynthesis required increased for CO2 consumption, higher PS II levels, and greater number and density of the thylakoids in the chloroplasts, got incompatible with the energy-consuming and overly large PBS antenna. PMID:25844449

  13. Pea amyloplast DNA is qualitatively similar to pea chloroplast DNA

    NASA Technical Reports Server (NTRS)

    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.

  14. Chloroplast Retrograde Regulation of Heat Stress Responses in Plants

    PubMed Central

    Sun, Ai-Zhen; Guo, Fang-Qing

    2016-01-01

    It is well known that intracellular signaling from chloroplast to nucleus plays a vital role in stress responses to survive environmental perturbations. The chloroplasts were proposed as sensors to heat stress since components of the photosynthetic apparatus housed in the chloroplast are the major targets of thermal damage in plants. Thus, communicating subcellular perturbations to the nucleus is critical during exposure to extreme environmental conditions such as heat stress. By coordinating expression of stress specific nuclear genes essential for adaptive responses to hostile environment, plants optimize different cell functions and activate acclimation responses through retrograde signaling pathways. The efficient communication between plastids and the nucleus is highly required for such diverse metabolic and biosynthetic functions during adaptation processes to environmental stresses. In recent years, several putative retrograde signals released from plastids that regulate nuclear genes have been identified and signaling pathways have been proposed. In this review, we provide an update on retrograde signals derived from tetrapyrroles, carotenoids, reactive oxygen species (ROS) and organellar gene expression (OGE) in the context of heat stress responses and address their roles in retrograde regulation of heat-responsive gene expression, systemic acquired acclimation, and cellular coordination in plants. PMID:27066042

  15. Chloroplast quality control - balancing energy production and stress.

    PubMed

    Woodson, Jesse D

    2016-10-01

    Contents 36 I. 36 II. 37 III. 37 IV. 38 V. 39 VI. 40 VII. 40 40 References 40 SUMMARY: All organisms require the ability to sense their surroundings and adapt. Such capabilities allow them to thrive in a wide range of habitats. This is especially true for plants, which are sessile and have to be genetically equipped to withstand every change in their environment. Plants and other eukaryotes use their energy-producing organelles (i.e. mitochondria and chloroplasts) as such sensors. In response to a changing cellular or external environment, these organelles can emit 'retrograde' signals that alter gene expression and/or cell physiology. This signaling is important in plants, fungi, and animals and impacts diverse cellular functions including photosynthesis, energy production/storage, stress responses, growth, cell death, ageing, and tumor progression. Originally, chloroplast retrograde signals in plants were known to lead to the reprogramming of nuclear transcription. New research, however, has pointed to additional posttranslational mechanisms that lead to chloroplast regulation and turnover in response to stress. Such mechanisms involve singlet oxygen, ubiquitination, the 26S proteasome, and cellular degradation machinery. PMID:27533783

  16. Non-canonical transit peptide for import into the chloroplast.

    PubMed

    Miras, Stéphane; Salvi, Daniel; Ferro, Myriam; Grunwald, Didier; Garin, Jérôme; Joyard, Jacques; Rolland, Norbert

    2002-12-01

    The large majority of plastid proteins are nuclear-encoded and, thus, must be imported within these organelles. Unlike most of the outer envelope proteins, targeting of proteins to all other plastid compartments (inner envelope membrane, stroma, and thylakoid) is strictly dependent on the presence of a cleavable transit sequence in the precursor N-terminal region. In this paper, we describe the identification of a new envelope protein component (ceQORH) and demonstrate that its subcellular localization is limited to the inner membrane of the chloroplast envelope. Immunopurification, microsequencing of the natural envelope protein and cloning of the corresponding full-length cDNA demonstrated that this protein is not processed in the N-terminal region during its targeting to the inner envelope membrane. Transient expression experiments in plant cells were performed with truncated forms of the ceQORH protein fused to the green fluorescent protein. These experiments suggest that neither the N-terminal nor the C-terminal are essential for chloroplastic localization of the ceQORH protein. These observations are discussed in the frame of the endosymbiotic theory of chloroplast evolution and suggest that a domain of the ceQORH bacterial ancestor may have evolved so as to exclude the general requirement of an N-terminal plastid transit sequence. PMID:12368288

  17. Lipid fixation during preparation of chloroplasts for electron microscopy.

    PubMed

    Ongun, A; Thomson, W W; Mudd, J B

    1968-07-01

    Reaction of osmium tetroxide with isolated spinach chloroplasts fixed completely the glycolipids, phosphatidyl glycerol, and phosphatidyl choline. Under the same reaction conditions only 30% of the chlorophyll was fixed. Reaction of potassium permanganate with isolated spinach chloroplasts fixed more than 90% of the glycolipids, phosphatidyl glycerol, and phosphatidyl choline, provided the reaction period was long enough. Potassium permanganate also fixed the chlorophyll. Reaction of osmium tetroxide and potassium permanganate with isolated (14)C-lipids from Chlorella pyrenoidosa fixed 59% and 66% of the radioactivity, respectively. The lipids that were not fixed included sterols and pigments. Electron micrographs show that chloroplasts extracted with chloroform-methanol after fixation in osmium tetroxide or potassium permanganate differ from those dehydrated with acetone mainly in that in the former, osmiophilic globules have been removed and there seems to be some fusion of the boundary membranes and grana membranes. These effects may be due to the extraction of unfixed, neutral lipids such as sterols and quinones. PMID:4177832

  18. Crosstalk between chloroplast thioredoxin systems in regulation of photosynthesis.

    PubMed

    Nikkanen, Lauri; Toivola, Jouni; Rintamäki, Eevi

    2016-08-01

    Thioredoxins (TRXs) mediate light-dependent activation of primary photosynthetic reactions in plant chloroplasts by reducing disulphide bridges in redox-regulated enzymes. Of the two plastid TRX systems, the ferredoxin-TRX system consists of ferredoxin-thioredoxin reductase (FTR) and multiple TRXs, while the NADPH-dependent thioredoxin reductase (NTRC) contains a complete TRX system in a single polypeptide. Using Arabidopsis plants overexpressing or lacking a functional NTRC, we have investigated the redundancy and interaction between the NTRC and Fd-TRX systems in regulation of photosynthesis in vivo. Overexpression of NTRC raised the CO2 fixation rate and lowered non-photochemical quenching and acceptor side limitation of PSI in low light conditions by enhancing the activation of chloroplast ATP synthase and TRX-regulated enzymes in Calvin-Benson cycle (CBC). Overexpression of NTRC with an inactivated NTR or TRX domain partly recovered the phenotype of knockout plants, suggesting crosstalk between the plastid TRX systems. NTRC interacted in planta with fructose-1,6-bisphosphatase, phosphoribulokinase and CF1 γ subunit of the ATP synthase and with several chloroplast TRXs. These findings indicate that NTRC-mediated regulation of the CBC and ATP synthesis occurs both directly and through interaction with the ferredoxin-TRX system and is crucial when availability of light is limiting photosynthesis. PMID:26831830

  19. Posttranslational modifications of FERREDOXIN-NADP+ OXIDOREDUCTASE in Arabidopsis chloroplasts.

    PubMed

    Lehtimäki, Nina; Koskela, Minna M; Dahlström, Käthe M; Pakula, Eveliina; Lintala, Minna; Scholz, Martin; Hippler, Michael; Hanke, Guy T; Rokka, Anne; Battchikova, Natalia; Salminen, Tiina A; Mulo, Paula

    2014-12-01

    Rapid responses of chloroplast metabolism and adjustments to photosynthetic machinery are of utmost importance for plants' survival in a fluctuating environment. These changes may be achieved through posttranslational modifications of proteins, which are known to affect the activity, interactions, and localization of proteins. Recent studies have accumulated evidence about the crucial role of a multitude of modifications, including acetylation, methylation, and glycosylation, in the regulation of chloroplast proteins. Both of the Arabidopsis (Arabidopsis thaliana) leaf-type FERREDOXIN-NADP(+) OXIDOREDUCTASE (FNR) isoforms, the key enzymes linking the light reactions of photosynthesis to carbon assimilation, exist as two distinct forms with different isoelectric points. We show that both AtFNR isoforms contain multiple alternative amino termini and undergo light-responsive addition of an acetyl group to the α-amino group of the amino-terminal amino acid of proteins, which causes the change in isoelectric point. Both isoforms were also found to contain acetylation of a conserved lysine residue near the active site, while no evidence for in vivo phosphorylation or glycosylation was detected. The dynamic, multilayer regulation of AtFNR exemplifies the complex regulatory network systems controlling chloroplast proteins by a range of posttranslational modifications, which continues to emerge as a novel area within photosynthesis research. PMID:25301888

  20. Method of producing metallized chloroplasts and use thereof in the photochemical production of hydrogen and oxygen

    DOEpatents

    Greenbaum, Elias

    1987-01-01

    The invention is primarily a metallized chloroplast composition for use in a photosynthetic reaction. A catalytic metal is precipitated on a chloroplast membrane at the location where a catalyzed reduction reaction occurs. This metallized chloroplast is stabilized by depositing it on a support medium such as fiber so that it can be easily handled. A possible application of this invention is the splitting of water to form hydrogen and oxygen that can be used as a renewable energy source.

  1. Effect of growth temperature on chloroplast structure and activity in barley.

    PubMed

    Smillie, R M; Critchley, C; Bain, J M; Nott, R

    1978-08-01

    Seedlings of barley (Hordeum vulgare L. cv. Abyssinian) were grown at constant temperature and light intensity and the properties and structure of chloroplasts in the primary leaf were examined. Seventeen growth temperatures ranging from 2 to 37 C were employed. Three major effects of the growth temperature were seen. (a) At very low and high growth temperatures chloroplast biogenesis was inhibited. This occurred in plants grown at temperatures above 32 C while growth at 2 C resulted in a mixed population of pale yellow, pale green, and green plants. (b) Chloroplasts were produced at all other temperatures tested but growth temperatures within a few degrees of those inhibitory to chloroplast development resulted in chloroplasts with abnormal properties and structure. Chloroplasts in the green plants grown at 2 and 5 C showed a number of structural peculiarities, including a characteristic crimping of granal thylakoids. Photoreductive activity, measured using ferricyanide as the Hill oxidant in the presence of gramicidin D, was high, but this activity in chloroplasts isolated from plants grown at 2 C showed thermal inactivation at temperatures 5 degrees lower than was the case with plants grown at higher temperatures. High growth temperatures (30 to 32 C) yielded chloroplasts with reduced photoreductive activity and a tendency toward the formation of large grana and disorientation of the lamellar systems with respect to one another. Chloroplasts of the most affected plants (grown at 32 C) frequently contained a very large elongated granum, with narrow intrathylakoid spaces. (c) Photoreductive activity was not constant at intermediate growth temperatures but steadily declined with decreasing growth temperatures between 27 and 11 C. Some alterations in chloroplast structure were also observed.The changes in chloroplast activity and structure indicate that acclimation to temperature takes place over the entire temperature range in which chloroplast development is

  2. Production of Recombinant Proteins in the Chloroplast of the Green Alga Chlamydomonas reinhardtii.

    PubMed

    Guzmán-Zapata, Daniel; Macedo-Osorio, Karla Soledad; Almaraz-Delgado, Alma Lorena; Durán-Figueroa, Noé; Badillo-Corona, Jesus Agustín

    2016-01-01

    Chloroplast transformation in the green algae Chlamydomonas reinhardtii can be used for the production of valuable recombinant proteins. Here, we describe chloroplast transformation of C. reinhardtii followed by protein detection. Genes of interest integrate stably by homologous recombination into the chloroplast genome following introduction by particle bombardment. Genes are inherited and expressed in lines recovered after selection in the presence of an antibiotic. Recombinant proteins can be detected by conventional techniques like immunoblotting and purified from liquid cultures. PMID:26614282

  3. The origin of populations of Arabidopsis thaliana in China, based on the chloroplast DNA sequences

    PubMed Central

    2010-01-01

    Background In the studies incorporating worldwide sampling of A. thaliana populations, the samples from East Asia, especially from China, were very scattered; and the studies focused on global patterns of cpDNA genetic variation among accessions of A. thaliana are very few. In this study, chloroplast DNA sequence variability was used to infer phylogenetic relationships among Arabidopsis thaliana accessions from around the world, with the emphasis on samples from China. Results A data set comprising 77 accessions of A. thaliana, including 19 field-collected Chinese accessions together with three related species (A. arenosa, A. suecica, and Olimarabidopsis cabulica) as the out-group, was compiled. The analysis of the nucleotide sequences showed that the 77 accessions of A. thaliana were partitioned into two major differentiated haplotype classes (MDHCs). The estimated divergence time of the two MDHCs was about 0.39 mya. Forty-nine haplotypes were detected among the 77 accessions, which exhibited nucleotide diversity (π) of 0.00169. The Chinese populations along the Yangtze River were characterized by five haplotypes, and the two accessions collected from the middle range of the Altai Mountains in China shared six specific variable sites. Conclusions The dimorphism in the chloroplast DNA could be due to founder effects during late Pleistocene glaciations and interglacial periods, although introgression cannot be ruled out. The Chinese populations along the Yangtze River may have dispersed eastwards to their present-day locations from the Himalayas. These populations originated from a common ancestor, and a rapid demographic expansion began approximately 90,000 years ago. Two accessions collected from the middle range of the Altai Mountains in China may have survived in a local refugium during late Pleistocene glaciations. The natural populations from China with specific genetic characteristics enriched the gene pools of global A. thaliana collections. PMID:20141622

  4. Regulation of chloroplast number and DNA synthesis in higher plants. Final report, August 1995--August 1996

    SciTech Connect

    Mullet, J.E.

    1997-06-17

    The long term objective of this research is to understand the process of chloroplast development and its coordination with leaf development in higher plants. This is important because the photosynthetic capacity of plants is directly related to leaf and chloroplast development. This research focused on obtaining a detailed description of leaf development and the early steps in chloroplast development including activation of plastid DNA synthesis, changes in plastid DNA copy number, activation of chloroplast transcription and increases in plastid number per cell. The research focused on the isolation of the plastid DNA polymerase, and identification of genetic mutants which are altered in their accumulation of plastid DNA and plastid number per cell.

  5. Different effects of eubacterial and eukaryotic DNA topoisomerase II inhibitors on chloroplasts ofEuglena gracilis

    NASA Astrophysics Data System (ADS)

    Krajčovič, Juraj; Ebringer, Libor

    1990-03-01

    Inhibitors of eubacterial and eukaryotic DNA topoisomerases type II exhibited different effects on chloroplasts of the flagellateEuglena gracilis. Antibacterial agents (cinoxacin, nalidixic and oxolinic acids, ciprofloxacin, enoxacin, norfloxacin and ofloxacin) from the group of quinolones and coumarins (coumermycin A1, clorobiocin and novobiocin) — all inhibitors of prokaryotic DNA topoisomerase II — were very potent eliminators of chloroplasts fromE. gracilis. In contrast, antitumor drugs (adriamycin, etoposide, teniposide and mitoxantrone) — antagonists of the eukaryotic counterpart — did not affect these semiautonomous photosynthetic organelles. These findings point out again the close evolutionary relationships between eubacteria and chloroplasts and are in agreement with the hypothesis of an endosymbiotic origin of chloroplasts.

  6. Purification and cDNA isolation of chloroplastic phosphoglycerate kinase from Chlamydomonas reinhardtii.

    PubMed Central

    Kitayama, M; Togasaki, R K

    1995-01-01

    Chloroplastic phosphoglycerate kinase (PGK) was purified to homogeneity from a soluble fraction of chloroplasts of a cell-wall-deficient mutant strain of Chlamydomonas reinhardtii (cw-15) using ammonium sulfate fractionation, Reactive Blue-72 column chromatography, and native polyacrylamide gel electrophoresis. PGK activity was attributed to a single polypeptide with a molecular mass of 42 kD. Relative purity and identity of the isolated enzyme was confirmed by N-terminal amino acid sequence determination. Antiserum against this enzyme was raised and a western blot analysis of whole-cell lysate from cw-15 cells using this anti-chloroplastic PGK serum detected a single polypeptide with a molecular mass of 42 kD. The cDNA clone corresponding to the Chlamydomonas chloroplastic PGK was isolated from a Chlamydomonas cDNA expression library using the anti-PGK serum. The cDNA sequence was determined and apparently codes for the entire precursor peptide, which consists of 461 codons. The results from Southern and northern blot analyses suggest that the chloroplastic PGK gene exists as a single copy in the nuclear genome of C. reinhardtii and is expressed as a 1.8-kb transcript. The C. reinhardtii chloroplastic PGK cDNA has 71 and 66% homology with wheat chloroplastic PGK and spinach chloroplastic PGK, respectively. Based on the deduced amino acid sequence, the chloroplastic PGK of C. reinhardtii has more similarity to plant PGKs than to other PGKs, having both prokaryotic and eukaryotic features. PMID:7724671

  7. Sonication-based isolation and enrichment of Chlorella protothecoides chloroplasts for illumina genome sequencing

    SciTech Connect

    Angelova, Angelina; Park, Sang-Hycuk; Kyndt, John; Fitzsimmons, Kevin; Brown, Judith K

    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.

  8. Using stable isotopes to characterize differential depth of water uptake based on environmental conditions in perennial biofuel and traditional annual crops

    NASA Astrophysics Data System (ADS)

    Miller, J. N.; Nystrom, R.; Bernacchi, C.

    2013-12-01

    Global climate change related to fossil fuel consumption coupled with the necessity for secure, cost-effective, and renewable domestic energy is continuing to drive the development of a bioenergy industry. Numerous second-generation biofuel crops have been identified that hold promise as sustainable feedstocks for the industry, including perennial grasses that utilize the highly water and energy efficient C4 photosynthetic pathway. Among the perennial grasses, miscanthus (Miscanthus × giganteus) and switchgrass (Panicum virgatum) stand out as having high biomass, minimal maintenance, low nutrient input requirements, and positive environmental benefits. These grasses are able to withstand a wide range of growing season temperatures and precipitation regimes, particularly in reference to the annual row crops that they are likely to replace. During the drought of 2012 traditional row crops suffered major reductions in yield whereas the perennial grasses retained relatively high biomass yields. We hypothesize that this is due to the ability of the perennial grasses to access water from deeper soil water relative to the annual row crops. To test this hypothesis, we use isotopic techniques to determine the soil depth from which the various species obtain water. Data from summer 2013 suggests that the perennial grasses preferentially use surface water when available but can extract water from depths that the annual row crops are unable to reach. These results indicate that perennial grasses, with deeper roots, will likely sustain growth under conditions when annual row crops are unable.

  9. Phylogeography of Thlaspi arvense (Brassicaceae) in China Inferred from Chloroplast and Nuclear DNA Sequences and Ecological Niche Modeling

    PubMed Central

    An, Miao; Zeng, Liyan; Zhang, Ticao; Zhong, Yang

    2015-01-01

    Thlaspi arvense is a well-known annual farmland weed with worldwide distribution, which can be found from sea level to above 4000 m high on the Qinghai-Tibetan Plateau (QTP). In this paper, a phylogeographic history of T. arvense including 19 populations from China was inferred by using three chloroplast (cp) DNA segments (trnL-trnF, rpl32-trnL and rps16) and one nuclear (n) DNA segment (Fe-regulated transporter-like protein, ZIP). A total of 11 chloroplast haplotypes and six nuclear alleles were identified, and haplotypes unique to the QTP were recognized (C4, C5, C7 and N4). On the basis of molecular dating, haplotypes C4, C5 and C7 have separated from others around 1.58 Ma for cpDNA, which corresponds to the QTP uplift. In addition, this article suggests that the T. arvense populations in China are a mixture of diverged subpopulations as inferred by hT/vT test (hT ≤ vT, cpDNA) and positive Tajima’s D values (1.87, 0.05 < p < 0.10 for cpDNA and 3.37, p < 0.01 for nDNA). Multimodality mismatch distribution curves and a relatively large shared area of suitable environmental conditions between the Last Glacial Maximum (LGM) as well as the present time recognized by MaxEnt software reject the sudden expansion population model. PMID:26110380

  10. Light-Dependent Isoprene Emission (Characterization of a Thylakoid-Bound Isoprene Synthase in Salix discolor Chloroplasts).

    PubMed Central

    Wildermuth, M. C.; Fall, R.

    1996-01-01

    Isoprene synthase is an enzyme that is responsible for the production of the volatile C5 hydrocarbon, isoprene, in plant leaves. Isoprene formation in numerous C3 plants is interesting because (a) large quantities of isoprene are emitted, 5 x 1014 g of C annually, (b) a plant may release 1 to 8% of its fixed C as isoprene, and (c) the function of plant isoprene production is unknown. Because of the dependence of foliar isoprene emission on light, the existence of a plastidic isoprene synthase has been postulated. To pursue this idea, a method to isolate chloroplasts from Salix discolor was developed and shows a plastidic isoprene synthase that is tightly bound to the thylakoid membrane and accessible to trypsin inactivation. The thylakoid-bound isoprene synthase has catalytic properties similar to known soluble isoprene synthases; however, the relationship between these enzymes is unknown. The discovery of a thylakoid-bound isoprene synthase with a stromal-facing domain places it in the chloroplast, where it may be subject to numerous direct and indirect light-mediated effects. Implications for the light-dependent regulation of foliar isoprene production and its function are presented. PMID:12226383

  11. Characterization of Coffea chloroplast microsatellites and evidence for the recent divergence of C. arabica and C. eugenioides chloroplast genomes.

    PubMed

    Tesfaye, Kassahun; Borsch, Thomas; Govers, Kim; Bekele, Endashaw

    2007-12-01

    Comparative sequencing of >7 kb of highly variable chloroplast genome regions (atpB-rbcL, trnS-trnG, rpl22-rps19, and rps19-rpl2 spacers; introns in atpF, trnG, trnK, and rpl16) with microsatellites known from other angiosperms was carried out in Coffea. Samples comprised 8 diploid species of Coffea, 5 individuals of tetraploid C. arabica representing geographically distant wild populations from Ethiopia, 2 commercial cultivars of C. arabica, and Psilanthus leroyi and Ixora coccinea as outgroups. Phylogeny reconstruction using maximum parsimony and Bayesian inference resulted in congruent topologies with high support for C. arabica and C. eugenioides being sisters. Partitioned analyses showed that all regions except the atpB-rbcL spacer resolved this sister-group, although this was often unsupported. The large sequence data set further shows that chloroplast genomes of C. arabica and C. eugenioides each possess apomorphies, indicating that not C. eugenioides but an ancestor or close relative of C. eugenioides is the maternal parent of C. arabica. Seven variable chloroplast microsatellites were characterized in Coffea. Most microsatellites are poly(A/T) stretches, whereas one in the trnS-trnG spacer has an (AT)n motif. Most strikingly, all individuals of C. arabica possess identical sequences, suggesting a single chloroplast haplotype. This can be explained by a recent origin of C. arabica in a unique allopolyploidization event, or by severe bottleneck effects in the evolutionary history of the species. Reconstruction of the evolution of microstructural mutations shows much higher levels of homoplasy in microsatellite loci than in other parts of spacers and introns. Microsatellites are inferred to evolve by insertion and deletion of 1 to 3 motif copies in one step. PMID:18059539

  12. Chloroplast-encoded serotonin N-acetyltransferase in the red alga Pyropia yezoensis: gene transition to the nucleus from chloroplasts.

    PubMed

    Byeon, Yeong; Yool Lee, Hyoung; Choi, Dong-Woog; Back, Kyoungwhan

    2015-02-01

    Melatonin biosynthesis involves the N-acetylation of arylalkylamines such as serotonin, which is catalysed by serotonin N-acetyltransferase (SNAT), the penultimate enzyme of melatonin biosynthesis in both animals and plants. Here, we report the functional characterization of a putative N-acetyltransferase gene in the chloroplast genome of the alga laver (Pyropia yezoensis, formerly known as Porphyra yezoensis) with homology to the rice SNAT gene. To confirm that the putative Pyropia yezoensis SNAT (PySNAT) gene encodes an SNAT, we cloned the full-length chloroplastidic PySNAT gene by PCR and purified the recombinant PySNAT protein from Escherichia coli. PySNAT was 174 aa and had 50% amino acid identity with cyanobacteria SNAT. Purified recombinant PySNAT showed a peak activity at 55 °C with a K m of 467 µM and V max of 28 nmol min-1 mg(-1) of protein. Unlike other plant SNATs, PySNAT localized to the cytoplasm due to a lack of N-terminal chloroplast transit peptides. Melatonin was present at 0.16ng g(-1) of fresh mass but increased during heat stress. Phylogenetic analysis of the sequence suggested that PySNAT has evolved from the cyanobacteria SNAT gene via endosymbiotic gene transfer. Additionally, the chloroplast transit peptides of plant SNATs were acquired 1500 million years ago, concurrent with the appearance of green algae. PMID:25183745

  13. Chloroplast-encoded serotonin N-acetyltransferase in the red alga Pyropia yezoensis: gene transition to the nucleus from chloroplasts

    PubMed Central

    Byeon, Yeong; Yool Lee, Hyoung; Choi, Dong-Woog; Back, Kyoungwhan

    2015-01-01

    Melatonin biosynthesis involves the N-acetylation of arylalkylamines such as serotonin, which is catalysed by serotonin N-acetyltransferase (SNAT), the penultimate enzyme of melatonin biosynthesis in both animals and plants. Here, we report the functional characterization of a putative N-acetyltransferase gene in the chloroplast genome of the alga laver (Pyropia yezoensis, formerly known as Porphyra yezoensis) with homology to the rice SNAT gene. To confirm that the putative Pyropia yezoensis SNAT (PySNAT) gene encodes an SNAT, we cloned the full-length chloroplastidic PySNAT gene by PCR and purified the recombinant PySNAT protein from Escherichia coli. PySNAT was 174 aa and had 50% amino acid identity with cyanobacteria SNAT. Purified recombinant PySNAT showed a peak activity at 55 °C with a K m of 467 µM and V max of 28 nmol min–1 mg–1 of protein. Unlike other plant SNATs, PySNAT localized to the cytoplasm due to a lack of N-terminal chloroplast transit peptides. Melatonin was present at 0.16ng g–1 of fresh mass but increased during heat stress. Phylogenetic analysis of the sequence suggested that PySNAT has evolved from the cyanobacteria SNAT gene via endosymbiotic gene transfer. Additionally, the chloroplast transit peptides of plant SNATs were acquired 1500 million years ago, concurrent with the appearance of green algae. PMID:25183745

  14. Mollusc-Algal Chloroplast Endosymbiosis. Photosynthesis, Thylakoid Protein Maintenance, and Chloroplast Gene Expression Continue for Many Months in the Absence of the Algal Nucleus1

    PubMed Central

    Green, Brian J.; Li, Wei-Ye; Manhart, James R.; Fox, Theodore C.; Summer, Elizabeth J.; Kennedy, Robert A.; Pierce, Sidney K.; Rumpho, Mary E.

    2000-01-01

    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 CO2 fixation for at least 9 months if provided with only light and a source of CO2. 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. PMID:10982447

  15. Whole genome sequencing of enriched chloroplast DNA using the Illumina GAII platform

    PubMed Central

    2010-01-01

    Background Complete chloroplast genome sequences provide a valuable source of molecular markers for studies in molecular ecology and evolution of plants. To obtain complete genome sequences, recent studies have made use of the polymerase chain reaction to amplify overlapping fragments from conserved gene loci. However, this approach is time consuming and can be more difficult to implement where gene organisation differs among plants. An alternative approach is to first isolate chloroplasts and then use the capacity of high-throughput sequencing to obtain complete genome sequences. We report our findings from studies of the latter approach, which used a simple chloroplast isolation procedure, multiply-primed rolling circle amplification of chloroplast DNA, Illumina Genome Analyzer II sequencing, and de novo assembly of paired-end sequence reads. Results A modified rapid chloroplast isolation protocol was used to obtain plant DNA that was enriched for chloroplast DNA, but nevertheless contained nuclear and mitochondrial DNA. Multiply-primed rolling circle amplification of this mixed template produced sufficient quantities of chloroplast DNA, even when the amount of starting material was small, and improved the template quality for Illumina Genome Analyzer II (hereafter Illumina GAII) sequencing. We demonstrate, using independent samples of karaka (Corynocarpus laevigatus), that there is high fidelity in the sequence obtained from this template. Although less than 20% of our sequenced reads could be mapped to chloroplast genome, it was relatively easy to assemble complete chloroplast genome sequences from the mixture of nuclear, mitochondrial and chloroplast reads. Conclusions We report successful whole genome sequencing of chloroplast DNA from karaka, obtained efficiently and with high fidelity. PMID:20920211

  16. The Unicellular Green Alga Chlamydomonas reinhardtii as an Experimental System to Study Chloroplast RNA Metabolism

    NASA Astrophysics Data System (ADS)

    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. Heat-induced Accumulation of Chloroplast Protein Synthesis Elongation Factor, EF-TU, in Winter Wheat

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Chloroplast protein synthesis elongation factor, EF-Tu, has been implicated in heat tolerance in maize (Zea mays L.). Chloroplast EF-Tu is highly conserved, and it is possible that this protein may be of importance to heat tolerance in other species including wheat (Triticum aestivum L.). In this ...

  18. The Arabidopsis YELLOW STRIPE LIKE4 and 6 transporters control iron release from the chloroplast.

    PubMed

    Divol, Fanchon; Couch, Daniel; Conéjéro, Geneviève; Roschzttardtz, Hannetz; Mari, Stéphane; Curie, Catherine

    2013-03-01

    In most plant cell types, the chloroplast represents the largest sink for iron, which is both essential for chloroplast metabolism and prone to cause oxidative damage. Here, we show that to buffer the potentially harmful effects of iron, besides ferritins for storage, the chloroplast is equipped with specific iron transporters that respond to iron toxicity by removing iron from the chloroplast. We describe two transporters of the YELLOW STRIPE1-LIKE family from Arabidopsis thaliana, YSL4 and YSL6, which are likely to fulfill this function. Knocking out both YSL4 and YSL6 greatly reduces the plant's ability to cope with excess iron. Biochemical and immunolocalization analyses showed that YSL6 resides in the chloroplast envelope. Elemental analysis and histochemical staining indicate that iron is trapped in the chloroplasts of the ysl4 ysl6 double mutants, which also accumulate ferritins. Also, vacuolar iron remobilization and NRAMP3/4 expression are inhibited. Furthermore, ubiquitous expression of YSL4 or YSL6 dramatically reduces plant tolerance to iron deficiency and decreases chloroplastic iron content. These data demonstrate a fundamental role for YSL4 and YSL6 in managing chloroplastic iron. YSL4 and YSL6 expression patterns support their physiological role in detoxifying iron during plastid dedifferentiation occurring in embryogenesis and senescence. PMID:23512854

  19. Induction and Segregation of Chloroplast Mutations in Vegetative Cell Cultures of Chlamydomonas Reinhardtii

    PubMed Central

    Lee, Robert W.; Haughn, George W.

    1980-01-01

    The single chloroplast of the alga Chlamydomonas reinhardtii contains at least 100 copies of the chloroplast chromosome. It is not known how the chloroplast (or cell) becomes homoplasmic for a mutation that arises in one of these copies. Under suitable selection conditions, clones with chloroplast mutations for streptomycin resistance induced by methyl methanesulfonate can be recovered with direct plating after mutagenesis. Using an adaptation of the Luria-Delbrück fluctuation test, mutagenized cultures grown on nonselective liquid medium for seven to nine doublings show negligible proliferation of cells capable of forming such mutant colonies. In contrast, cells among the same cultures with reduced nuclear mutations conferring streptomycin resistance reveal considerable clonal propagation prior to plating on selection medium. Reconstruction growth-rate experiments show no reduced growth of cells with chloroplast mutations relative to either wild-type cells or to those with nuclear mutations. We propose that newly arising chloroplast mutations and their copies are usually transmitted to only one daughter cell for several cell generations by reductional divisions of the chloroplast genome. In the absence of recombination and mixing, such a reductional partition of chloroplast alleles would readily permit the formation of homoplasmic lines without the need for selection. PMID:17249064

  20. Chloroplast Proteases: Updates on Proteolysis within and across Suborganellar Compartments1[OPEN

    PubMed Central

    Nishimura, Kenji

    2016-01-01

    Chloroplasts originated from the endosymbiosis of ancestral cyanobacteria and maintain transcription and translation machineries for around 100 proteins. Most endosymbiont genes, however, have been transferred to the host nucleus, and the majority of the chloroplast proteome is composed of nucleus-encoded proteins that are biosynthesized in the cytosol and then imported into chloroplasts. How chloroplasts and the nucleus communicate to control the plastid proteome remains an important question. Protein-degrading machineries play key roles in chloroplast proteome biogenesis, remodeling, and maintenance. Research in the past few decades has revealed more than 20 chloroplast proteases, which are localized to specific suborganellar locations. In particular, two energy-dependent processive proteases of bacterial origin, Clp and FtsH, are central to protein homeostasis. Processing endopeptidases such as stromal processing peptidase and thylakoidal processing peptidase are involved in the maturation of precursor proteins imported into chloroplasts by cleaving off the amino-terminal transit peptides. Presequence peptidases and organellar oligopeptidase subsequently degrade the cleaved targeting peptides. Recent findings have indicated that not only intraplastidic but also extraplastidic processive protein-degrading systems participate in the regulation and quality control of protein translocation across the envelopes. In this review, we summarize current knowledge of the major chloroplast proteases in terms of type, suborganellar localization, and diversification. We present details of these degradation processes as case studies according to suborganellar compartment (envelope, stroma, and thylakoids). Key questions and future directions in this field are discussed. PMID:27288365

  1. Participation of β-carotene in reactivation of PSI of heptane-extracted spinach chloroplasts.

    PubMed

    Tukendorf, A; Subczynski, W K; Baszynski, T

    1981-09-01

    A carotenoid requirement for photosystem I activity in spinach chloroplasts using extraction-reconstitution technique has been investigated. The transfer of electron from N,N,N',N'-tetramethyl-p-phenylene diamine through the chloroplast photosystem to methyl viologen dye or to NADP(+) was used as an assay of photosystem I activity. Extraction of lyophilized spinach chloroplasts with heptane at near 0°C removed almost all β-carotene and reduced photochemical activities associated with photosystem I to a low level (about 15% of the original activity). Reconstitution of the extracted chloroplasts with β-carotene completely restored photosystem I activity. The maximum rate of methyl viologen photoreduction in reconstituted chloroplasts occurred at an β-carotene/chlorophyll molar ratio of 0.5. Cyclic phosphorylation mediated by phenazine methosulphate was partially restored. Xanthophylls (lutein, neoxanthin, violaxanthin), as components of chloroplast membranes, were not able to replace β-carotene in reconstitution of chloroplasts and had essentially no effect on restoring photoreactions. On the basis of the P700/total chlorophyll ratio it can be assumed that extraction of lyophilized chloroplasts with heptane do not affect photosystem I reaction centre. Therefore it is possible that β-carotene, removed during heptane extraction and belonging mainly to the antenna pigment pool of photosystem I, is effective in the restoration of photosystem I activity. PMID:24470228

  2. Carbonic anhydrase activity in isolated chloroplasts of chlamydomonas reinhardtii

    SciTech Connect

    Katzman, G.; Togasaki, R.K. ); Marcus, Y. ); Moroney, J.V. )

    1989-04-01

    In a new assay of carbonic anhydrase, NaH{sup 14}CO{sub 3} solution at the bottom of a sealed vessel releases {sup 14}CO{sub 3} which diffuses to the top of the vessel to be assimilated by actively photosynthesizing Chlamydomonas cells. The assay is initiated by illuminating cells and stopped by turning the light off and killing the cells with acid. Enzyme activity was estimated from acid stable radioactivity above the uncatalyzed background level. With bovine carbonic anhydrase, 1.5 Wilbur Anderson Unit (WAU) can be consistantly measured at 5-6 fold above background. Sonicated whole cells of air adapted wild type (+)gave 741.1 {plus minus} 12.4 WAU/mg chl. Intact washed cells of mixotrophically grown wall-less mutant CWD(-) and a high CO2 requiring wall-less double mutant CIA-3/CW15 (-) gave 7.1 {plus minus} 1.9 and 2.8 {plus minus} 7.8 WAU/mg chl respectively. Chloroplasts isolated from CWD and CIA-3/CW15 and subsequently disrupted gave 64.0 {plus minus} 14.7 and 2.8 {plus minus} 3.2 WAU/mg chl respectively. Chloroplast sonicate from another wall-less mutant CW15(-) gave activity comparable to CWD. Thus on a chlorophyll basis, enzyme activity in chloroplasts from mixotrophically grown cells is about 1/10th of the level found in air adapted wild type cells. CIA-3 seems to lack this activity.

  3. Photoinduction of cyclosis-mediated interactions between distant chloroplasts.

    PubMed

    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. PMID:25615586

  4. Studies on the reconstitution of o(2)-evolution of chloroplasts.

    PubMed

    Sayre, R T; Cheniae, G M

    1982-05-01

    Extraction of spinach (Spinacia oleracea L.) chloroplasts with cholate-asolectin in the absence of Mg(2+) results in the rapid and selective inactivation of O(2) evolution and a partial (30 to 40%) loss of photosystem II (PSII) donor activity without extraction of thylakoid bound Mn ( approximately 5 to 6 Mn per 400 Chlorophyll). Inclusion of ethylene glycol in the extractions inhibits loss of O(2) evolution and results in quantitative and qualitative differences in proteins solubilized but does not significantly inhibit the partial loss of PSII donor activity. Similarly, in two stage experiments (extraction followed by addition of organic solvent and solubilized thylakoid protein), O(2) evolution (V and V(max)) of extracted chloroplasts is enhanced approximately 2.5- to 8-fold. However, PSII donor activity remains unaffected. This reversal of cholate inactivation of O(2) evolution can be induced by solvents including ethanol, methanol, 2-propanol, and dimethyl sulfoxide. Such enhancements of O(2) evolution specifically required cholate-solubilized proteins, which are insensitive to NH(2)OH and are only moderately heat-labile. NH(2)OH extraction of chloroplasts prior to cholate-asolectin extraction abolishes reconstitutability of O(2) evolution. Thus, the protein(s) affecting reconstitution is unlike those of the O(2).Mn enzyme. The specific activity of the protein fraction effecting reconstitution of O(2) evolution is greatest in fractions depleted of the reported Mn-containing, 65-kilodalton, and the Fe-heme, 232-kilodalton (58-kilodalton monomer), proteins. Divalent ( approximately 3 millimolar) and monovalent ( approximately 30 millimolar) cations do not affect reconstitution of PSII donor activity but do affect reconstitution of O(2) evolution by decreasing the protein(s) concentration required for reconstitution of O(2) evolution in nonfractionated, cholate-asolectin extractions. The data indicate a reconstitution of the PSII segment linking the PSII

  5. The complete chloroplast genome sequence of Dendrobium officinale.

    PubMed

    Yang, Pei; Zhou, Hong; Qian, Jun; Xu, Haibin; Shao, Qingsong; Li, Yonghua; Yao, Hui

    2016-01-01

    The complete chloroplast sequence of Dendrobium officinale, an endangered and economically important traditional Chinese medicine, was reported and characterized. The genome size is 152,018 bp, with 37.5% GC content. A pair of inverted repeats (IRs) of 26,284 bp are separated by a large single-copy region (LSC, 84,944 bp) and a small single-copy region (SSC, 14,506 bp). The complete cp DNA contains 83 protein-coding genes, 39 tRNA genes and 8 rRNA genes. Fourteen genes contained one or two introns. PMID:25103425

  6. Proteomic comparison of etioplast and chloroplast protein complexes.

    PubMed

    Plöscher, Matthias; Reisinger, Veronika; Eichacker, Lutz A

    2011-08-12

    Angiosperms grown in darkness develop etioplasts during skotomorphogenesis. It is well known that etioplasts accumulate large quantities of protochlorophyllideoxidoreductase, are devoid of chlorophyll and are the site to assemble the photosynthetic machinery during photomorphogenesis. Proteomic investigation of the membrane protein complexes by Native PAGE, in combination with CyDye labelling and mass spectrometric analysis revealed that etioplasts and chloroplasts share a number of membrane protein complexes characteristic for electron transport, chlorophyll and protein synthesis as well as fatty acid biosynthesis. The complex regulatory function in both developmental states is discussed. PMID:21440687

  7. The complete chloroplast genome sequence of Hibiscus syriacus.

    PubMed

    Kwon, Hae-Yun; Kim, Joon-Hyeok; Kim, Sea-Hyun; Park, Ji-Min; Lee, Hyoshin

    2016-09-01

    The complete chloroplast genome sequence of Hibiscus syriacus L. is presented in this study. The genome is composed of 161 019 bp in length, with a typical circular structure containing a pair of inverted repeats of 25 745 bp of length separated by a large single-copy region and a small single-copy region of 89 698 bp and 19 831 bp of length, respectively. The overall GC content is 36.8%. One hundred and fourteen genes were annotated, including 81 protein-coding genes, 4 ribosomal RNA genes and 29 transfer RNA genes. PMID:26357910

  8. The complete chloroplast genome sequence of Chloranthus japonicus.

    PubMed

    Sun, Jing; Zhang, Gang; Li, Yimin; Chen, Ying; Zhang, Xiaofei; Tang, Zhishu; Wu, Haifeng

    2016-09-01

    The complete chloroplast genome of Chloranthus japonicus, an important traditional Chinese herbal medicine, was sequenced and characterized in this study. The genome size is 158,640 bp in length with 38.9% GC content. Two inverted repeats of 26,149 bp are separated by a large single-copy region (87,724 bp) and a small single-copy region (18,618 bp). The genome contains 131 individual genes, including 86 protein-coding genes, 37 tRNA genes and 8 rRNA genes. Eighteen genes contain one or two introns. PMID:25707409

  9. The complete chloroplast genome sequence of Curcuma flaviflora (Curcuma).

    PubMed

    Zhang, Yan; Deng, Jiabin; Li, Yangyi; Gao, Gang; Ding, Chunbang; Zhang, Li; Zhou, Yonghong; Yang, Ruiwu

    2016-09-01

    The complete chloroplast (cp) genome of Curcuma flaviflora, a medicinal plant in Southeast Asia, was sequenced. The genome size was 160 478 bp in length, with 36.3% GC content. A pair of inverted repeats (IRs) of 26 946 bp were separated by a large single copy (LSC) of 88 008 bp and a small single copy (SSC) of 18 578 bp, respectively. The cp genome contained 132 annotated genes, including 79 protein coding genes, 30 tRNA genes, and four rRNA genes. And 19 of these genes were duplicated in inverted repeat regions. PMID:26367332

  10. GOBASE—a database of mitochondrial and chloroplast information

    PubMed Central

    O'Brien, Emmet A.; Badidi, Elarbi; Barbasiewicz, Ania; deSousa, Cristina; Lang, B. Franz; Burger, Gertraud

    2003-01-01

    GOBASE is a relational database containing integrated sequence, RNA secondary structure and biochemical and taxonomic information about organelles. GOBASE release 6 (summer 2002) contains over 130 000 mitochondrial sequences, an increase of 37% over the previous release, and more than 30 000 chloroplast sequences in a new auxiliary database. To handle this flood of new data, we have designed and implemented GOpop, a Java system for population and verification of the database. We have also implemented a more powerful and flexible user interface using the PHP programming language. http://megasun.bch.umontreal.ca/gobase/gobase.html. PMID:12519975

  11. A Nucleus-Encoded Chloroplast Phosphoprotein Governs Expression of the Photosystem I Subunit PsaC in Chlamydomonas reinhardtii.

    PubMed

    Douchi, Damien; Qu, Yujiao; Longoni, Paolo; Legendre-Lefebvre, Linnka; Johnson, Xenie; Schmitz-Linneweber, Christian; Goldschmidt-Clermont, Michel

    2016-05-01

    The nucleo-cytoplasmic compartment exerts anterograde control on chloroplast gene expression through numerous proteins that intervene at posttranscriptional steps. Here, we show that the maturation of psaC mutant (mac1) of Chlamydomonas reinhardtii is defective in photosystem I and fails to accumulate psaC mRNA. The MAC1 locus encodes a member of the Half-A-Tetratricopeptide (HAT) family of super-helical repeat proteins, some of which are involved in RNA transactions. The Mac1 protein localizes to the chloroplast in the soluble fraction. MAC1 acts through the 5' untranslated region of psaC transcripts and is required for their stability. Small RNAs that map to the 5'end of psaC RNA in the wild type but not in the mac1 mutant are inferred to represent footprints of MAC1-dependent protein binding, and Mac1 expressed in bacteria binds RNA in vitro. A coordinate response to iron deficiency, which leads to dismantling of the photosynthetic electron transfer chain and in particular of photosystem I, also causes a decrease of Mac1. Overexpression of Mac1 leads to a parallel increase in psaC mRNA but not in PsaC protein, suggesting that Mac1 may be limiting for psaC mRNA accumulation but that other processes regulate protein accumulation. Furthermore, Mac 1 is differentially phosphorylated in response to iron availability and to conditions that alter the redox balance of the electron transfer chain. PMID:27113776

  12. Genetic structure based on nuclear and chloroplast microsatellite loci of Solanum lycocarpum A. St. Hil. (Solanaceae) in Central Brazil.

    PubMed

    Martins, K; Chaves, L J; Vencovsky, R; Kageyama, P Y

    2011-01-01

    Solanum lycocarpum (Solanaceae) is a woody species found in the Brazilian Cerrado. The flowers are pollinated by Xylocopa spp bees, and seeds are dispersed by mammals with distinct home range sizes. As a consequence, relative contributions of pollen and seeds to overall gene flow can vary according to different spatial scales. We studied the genetic structure of four natural populations of S. lycocarpum separated by 19 to 128 km, including individuals located along dirt roads that interlink three of the populations. A total of 294 individuals were genotyped with five nuclear and six chloroplast microsatellite markers. Significant spatial genetic structure was found in the total set of individuals; the Sp statistic was 0.0086. Population differentiation based on the six chloroplast microsatellite markers (θ(pC) = 0.042) was small and similar to that based on the five nuclear microsatellite markers (θ(p) = 0.054). For this set of populations, pollen and seed flow did not differ significantly from one another (pollen-to-seed flow ratio = 1.22). Capability for long distance seed dispersion and colonization of anthropogenic sites contributes to the ability of S. lycocarpum to maintain genetic diversity. Seed dispersion along dirt roads may be critical in preserving S. lycocarpum genetic diversity in fragmented landscapes. PMID:21523656

  13. Variability of chloroplast DNA and nuclear ribosomal DNA in cassava (Manihot esculenta Crantz) and its wild relatives.

    PubMed

    Fregene, M A; Vargas, J; Ikea, J; Angel, F; Tohme, J; Asiedu, R A; Akoroda, M O; Roca, W M

    1994-11-01

    Chloroplast DNA (cp) and nuclear ribosomal DNA (rDNA) variation was investigated in 45 accessions of cultivated and wild Manihot species. Ten independent mutations, 8 point mutations and 2 length mutations were identified, using eight restriction enzymes and 12 heterologous cpDNA probes from mungbean. Restriction fragment length polymorphism analysis defined nine distinct chloroplast types, three of which were found among the cultivated accessions and six among the wild species. Cladistic analysis of the cpDNA data using parsimony yielded a hypothetical phylogeny of lineages among the cpDNAs of cassava and its wild relatives that is congruent with morphological evolutionary differentiation in the genus. The results of our survey of cpDNA, together with rDNA restriction site change at the intergenic spacer region and rDNA repeat unit length variation (using rDNA cloned fragments from taro as probe), suggest that cassava might have arisen from the domestication of wild tuberous accessions of some Manihot species, followed by intensive selection. M. esculenta subspp flabellifolia is probably a wild progenitor. Introgressive hybridization with wild forms and pressures to adapt to the widely varying climates and topography in which cassava is found might have enhanced the crop's present day variability. PMID:24178017

  14. Chloroplast Transformation of Platymonas (Tetraselmis) subcordiformis with the bar Gene as Selectable Marker

    PubMed Central

    Cui, Yulin; Qin, Song; Jiang, Peng

    2014-01-01

    The objective of this research was to establish a chloroplast transformation technique for Platymonas (Tetraselmis) subcordiformis. Employing the gfp gene as a reporter and the bar gene as a selectable marker, transformation vectors of P. subcordiformis chloroplast were constructed with endogenous fragments rrn16S–trnI (left) and trnA–rrn23S (right) as a recombination site of the chloroplast genome. The plasmids were transferred into P. subcordiformis via particle bombardment. Confocal laser scanning microscopy indicated that the green fluorescence protein was localized in the chloroplast of P. subcordiformis, confirming the activity of the Chlamydomonas reinhardtii promoter. Cells transformed with the bar gene were selected using the herbicide Basta. Resistant colonies were analyzed by PCR and Southern blotting, and the results indicated that the bar gene was successfully integrated into the chloroplast genome via homologous recombination. The technique will improve genetic engineering of this alga. PMID:24911932

  15. The evolution of blue-greens and the origins of chloroplasts

    NASA Technical Reports Server (NTRS)

    Schwartz, R. M.; Dayhoff, M. O.

    1981-01-01

    All of the available molecular data support the theory that the chloroplasts of eukaryote cells were originally free-living blue-greens. Of great interest is what the relationships are between contemporary types of blue-greens and eukaryote chloroplasts and whether the chloroplasts of the various eukaryotes are the result of one or more than one symbiosis. By combining information from phylogenetic trees based on cytochrome c6 and 2Fe-2S ferredoxin sequences, it is shown that the chloroplasts of a number of eukaryote algae as well as the protist Euglena are polyphyletic; the chloroplasts of green algae and the higher plants may be the result of a single symbiosis.

  16. Circadian Rhythms of Chloroplast Orientation and Photosynthetic Capacity in Ulva123

    PubMed Central

    Britz, Steven J.; Briggs, Winslow R.

    1976-01-01

    Ulva lactuca L. var. latissima (L.) Decandolle and var. rigida (C. Agardh) Le Jolis and U. mutabilis Foyn have a circadian rhythm of chloroplast orientation which results in large changes in the light-absorption properties of the thallus. During the day, the chloroplasts cover the outer face of the cells and absorbance is high. At night, the chloroplasts are along the side walls and absorbance is low. Enteromorpha linza (L.) J. Agardh, E. intestinalis (L.) Link, E. sp., and Monostroma grevillei (Thuret) Wittrock, members of the Ulvales, were not observed to have this rhythmic movement. Chloroplasts, when in the face position, could not be induced to move to the sides by high intensity light up to 80,000 lux. Unrelated to chloroplast position per se and light-absorption efficiency, there is a rhythm of photosynthetic capacity which peaks just before midday and which continues in constant darkness. Images PMID:16659613

  17. Chloroplast transformation of Platymonas (Tetraselmis) subcordiformis with the bar gene as selectable marker.

    PubMed

    Cui, Yulin; Qin, Song; Jiang, Peng

    2014-01-01

    The objective of this research was to establish a chloroplast transformation technique for Platymonas (Tetraselmis) subcordiformis. Employing the gfp gene as a reporter and the bar gene as a selectable marker, transformation vectors of P. subcordiformis chloroplast were constructed with endogenous fragments rrn16S-trnI (left) and trnA-rrn23S (right) as a recombination site of the chloroplast genome. The plasmids were transferred into P. subcordiformis via particle bombardment. Confocal laser scanning microscopy indicated that the green fluorescence protein was localized in the chloroplast of P. subcordiformis, confirming the activity of the Chlamydomonas reinhardtii promoter. Cells transformed with the bar gene were selected using the herbicide Basta. Resistant colonies were analyzed by PCR and Southern blotting, and the results indicated that the bar gene was successfully integrated into the chloroplast genome via homologous recombination. The technique will improve genetic engineering of this alga. PMID:24911932

  18. The complete chloroplast genome sequence of Dendropanax morbifera (Léveillé).

    PubMed

    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. PMID:26153746

  19. Chloroplasts extend stromules independently and in response to internal redox signals.

    PubMed

    Brunkard, Jacob O; Runkel, Anne M; Zambryski, Patricia C

    2015-08-11

    A fundamental mystery of plant cell biology is the occurrence of "stromules," stroma-filled tubular extensions from plastids (such as chloroplasts) that are universally observed in plants but whose functions are, in effect, completely unknown. One prevalent hypothesis is that stromules exchange signals or metabolites between plastids and other subcellular compartments, and that stromules are induced during stress. Until now, no signaling mechanisms originating within the plastid have been identified that regulate stromule activity, a critical missing link in this hypothesis. Using confocal and superresolution 3D microscopy, we have shown that stromules form in response to light-sensitive redox signals within the chloroplast. Stromule frequency increased during the day or after treatment with chemicals that produce reactive oxygen species specifically in the chloroplast. Silencing expression of the chloroplast NADPH-dependent thioredoxin reductase, a central hub in chloroplast redox signaling pathways, increased chloroplast stromule frequency, whereas silencing expression of nuclear genes related to plastid genome expression and tetrapyrrole biosynthesis had no impact on stromules. Leucoplasts, which are not photosynthetic, also made more stromules in the daytime. Leucoplasts did not respond to the same redox signaling pathway but instead increased stromule formation when exposed to sucrose, a major product of photosynthesis, although sucrose has no impact on chloroplast stromule frequency. Thus, different types of plastids make stromules in response to distinct signals. Finally, isolated chloroplasts could make stromules independently after extraction from the cytoplasm, suggesting that chloroplast-associated factors are sufficient to generate stromules. These discoveries demonstrate that chloroplasts are remarkably autonomous organelles that alter their stromule frequency in reaction to internal signal transduction pathways. PMID:26150490

  20. Chloroplast NDH: A different enzyme with a structure similar to that of respiratory NADH dehydrogenase.

    PubMed

    Shikanai, Toshiharu

    2016-07-01

    Eleven genes encoding chloroplast NADH dehydrogenase-like (NDH) complex have been discovered in plastid genomes on the basis of their homology with genes encoding respiratory complex I. Despite this structural similarity, chloroplast NDH and its evolutionary origin NDH-1 in cyanobacteria accept electrons from ferredoxin (Fd), indicating that chloroplast NDH is an Fd-dependent plastoquinone (PQ) reductase rather than an NAD(P)H dehydrogenase. In Arabidopsis thaliana, chloroplast NDH interacts with photosystem I (PSI); this interaction is needed to stabilize NDH, especially under high light. On the basis of these distinct characters of chloroplast and cyanobacterial NDH, it can be distinguished as a photosynthetic NDH from respiratory complex I. In fact, chloroplast NDH forms part of the machinery of photosynthesis by mediating the minor pathway of PSI cyclic electron transport. Along with the antimycin A-sensitive main pathway of PSI cyclic electron transport, chloroplast NDH compensates the ATP/NADPH production ratio in the light reactions of photosynthesis. In this review, I revisit the original concept of chloroplast NDH on the basis of its similarity to respiratory complex I and thus introduce current progress in the field to researchers focusing on respiratory complex I. I summarize recent progress on the basis of structure and function. Finally, I introduce the results of our examination of the process of assembly of chloroplast NDH. Although the process requires many plant-specific non-subunit factors, the core processes of assembly are conserved between chloroplast NDH and respiratory complex I. This article is part of a Special Issue entitled Respiratory complex I, edited by Volker Zickermann and Ulrich Brandt. PMID:26519774

  1. Chloroplast redox homeostasis is essential for lateral root formation in Arabidopsis

    PubMed Central

    Ferrández, Julia; González, Maricruz; Cejudo, Francisco Javier

    2012-01-01

    Redox regulation based on dithiol-disulphide interchange is an essential component of the control of chloroplast metabolism. In contrast to heterotrophic organisms, and non-photosynthetic plant tissues, chloroplast redox regulation relies on ferredoxin (Fd) reduced by the photosynthetic electron transport chain, thus being highly dependent on light. The finding of the NADPH-dependent thioredoxin reductase C (NTRC), a chloroplast-localized NTR with a joint thioredoxin domain, showed that NADPH is also used as source of reducing power for chloroplast redox homeostasis. Recently we have found that NTRC is also in plastids of non-photosynthetic tissues. Because these non-green plastids lack photochemical reactions, their redox homeostasis depends exclusively on NADPH produced from sugars and, thus, NTRC may play an essential role maintaining the redox homeostasis in these plastids. The fact that redox regulation occurs in any type of plastids raises the possibility that the functions of chloroplasts and non-green plastids, such as amyloplasts, are integrated to harmonize the growth of the different organs of the plant. To address this question, we generated Arabidopsis plants the redox homeostasis of which is recovered exclusively in chloroplasts, by leaf-specific expression of NTRC in the ntrc mutant, or exclusively in amyloplasts, by root-specific expression of NTRC. The analysis of these plants suggests that chloroplasts exert a pivotal role on plant growth, as expected because chloroplasts constitute the major source of nutrients and energy, derived from photosynthesis, for growth of heterotrophic tissues. However, NTRC deficiency causes impairment of auxin synthesis and lateral root formation. Interestingly, recovery of redox homeostasis of chloroplasts, but not of amyloplasts, was sufficient to restore wild type levels of lateral roots, showing the important signaling function of chloroplasts for the development of heterotrophic organs. PMID:22899086

  2. Mesophyll Chloroplast Investment in C3, C4 and C2 Species of the Genus Flaveria.

    PubMed

    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. PMID:26985020

  3. The complete chloroplast genome provides insight into the evolution and polymorphism of Panax ginseng

    PubMed Central

    Zhao, Yongbing; Yin, Jinlong; Guo, Haiyan; Zhang, Yuyu; Xiao, Wen; Sun, Chen; Wu, Jiayan; Qu, Xiaobo; Yu, Jun; Wang, Xumin; Xiao, Jingfa

    2015-01-01

    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 strains

  4. Copper-Deficiency in Brassica napus Induces Copper Remobilization, Molybdenum Accumulation and Modification of the Expression of Chloroplastic Proteins

    PubMed Central

    Billard, Vincent; Ourry, Alain; Maillard, Anne; Garnica, Maria; Coquet, Laurent; Jouenne, Thierry; Cruz, Florence; Garcia-Mina, José-Maria; Yvin, Jean-Claude; Etienne, Philippe

    2014-01-01

    During the last 40 years, crop breeding has strongly increased yields but has had adverse effects on the content of micronutrients, such as Fe, Mg, Zn and Cu, in edible products despite their sufficient supply in most soils. This suggests that micronutrient remobilization to edible tissues has been negatively selected. As a consequence, the aim of this work was to quantify the remobilization of Cu in leaves of Brassica napus L. during Cu deficiency and to identify the main metabolic processes that were affected so that improvements can be achieved in the future. While Cu deficiency reduced oilseed rape growth by less than 19% compared to control plants, Cu content in old leaves decreased by 61.4%, thus demonstrating a remobilization process between leaves. Cu deficiency also triggered an increase in Cu transporter expression in roots (COPT2) and leaves (HMA1), and more surprisingly, the induction of the MOT1 gene encoding a molybdenum transporter associated with a strong increase in molybdenum (Mo) uptake. Proteomic analysis of leaves revealed 33 proteins differentially regulated by Cu deficiency, among which more than half were located in chloroplasts. Eleven differentially expressed proteins are known to require Cu for their synthesis and/or activity. Enzymes that were located directly upstream or downstream of Cu-dependent enzymes were also differentially expressed. The overall results are then discussed in relation to remobilization of Cu, the interaction between Mo and Cu that occurs through the synthesis pathway of Mo cofactor, and finally their putative regulation within the Calvin cycle and the chloroplastic electron transport chain. PMID:25333918

  5. Copper-deficiency in Brassica napus induces copper remobilization, molybdenum accumulation and modification of the expression of chloroplastic proteins.

    PubMed

    Billard, Vincent; Ourry, Alain; Maillard, Anne; Garnica, Maria; Coquet, Laurent; Jouenne, Thierry; Cruz, Florence; Garcia-Mina, José-Maria; Yvin, Jean-Claude; Etienne, Philippe

    2014-01-01

    During the last 40 years, crop breeding has strongly increased yields but has had adverse effects on the content of micronutrients, such as Fe, Mg, Zn and Cu, in edible products despite their sufficient supply in most soils. This suggests that micronutrient remobilization to edible tissues has been negatively selected. As a consequence, the aim of this work was to quantify the remobilization of Cu in leaves of Brassica napus L. during Cu deficiency and to identify the main metabolic processes that were affected so that improvements can be achieved in the future. While Cu deficiency reduced oilseed rape growth by less than 19% compared to control plants, Cu content in old leaves decreased by 61.4%, thus demonstrating a remobilization process between leaves. Cu deficiency also triggered an increase in Cu transporter expression in roots (COPT2) and leaves (HMA1), and more surprisingly, the induction of the MOT1 gene encoding a molybdenum transporter associated with a strong increase in molybdenum (Mo) uptake. Proteomic analysis of leaves revealed 33 proteins differentially regulated by Cu deficiency, among which more than half were located in chloroplasts. Eleven differentially expressed proteins are known to require Cu for their synthesis and/or activity. Enzymes that were located directly upstream or downstream of Cu-dependent enzymes were also differentially expressed. The overall results are then discussed in relation to remobilization of Cu, the interaction between Mo and Cu that occurs through the synthesis pathway of Mo cofactor, and finally their putative regulation within the Calvin cycle and the chloroplastic electron transport chain. PMID:25333918

  6. Protein synthesis in chloroplasts. Characteristics and products of protein synthesis in vitro in etioplasts and developing chloroplasts from pea leaves.

    PubMed Central

    Siddell, S G; Ellis, R J

    1975-01-01

    The function of plastid ribosomes in pea (Pisum sativum L.) was investigated by characterizing the products of protein synthesis in vitro in plastids isolated at different stages during the transition from etioplast to chloroplast. Etioplasts and plastids isolated after 24, 48 and 96h of greening in continuous white light, use added ATP to incorporate labelled amino acids into protein. Plastids isolated from greening leaves can also use light as the source of energy for protein synthesis. The labelled polypeptides synthesized in isolated plastids were analysed by electrophoresis in sodium dodecyl sulphate-ureapolyacrylamide gels. Six polypeptides are synthesized in etioplasts with ATP as energy source. Only one of these polypeptides is present in a 150 000g supernatant fraction. This polypeptide has been identified as the large subunit of Fraction I protein (3-phospho-D-glycerate carboxylyase EC 4.1.1.39) by comparing the tryptic 'map' of its L-(35S)methionine-labelled peptides with the tryptic 'map' of large subunit peptides from Fraction I labelled with L-(35S)methionine in vivo. The same gel pattern of six polypeptides is seen when plastids isolated from greening leaves are incubated with either added ATP or light as the energy source. However, the rates of synthesis of particular polypeptides are different in plastids isolated at different stages of the etioplast to chloroplast transition. The results support the idea that plastid ribosomes synthesize only a small number of proteins, and that the number and molecular weight of these proteins does not alter during the formation of chloroplasts from etioplasts. Images PLATE 1 PMID:1147911

  7. Chloroplast SRP54 Was Recruited for Posttranslational Protein Transport via Complex Formation with Chloroplast SRP43 during Land Plant Evolution.

    PubMed

    Dünschede, Beatrix; Träger, Chantal; Schröder, Christine Vera; Ziehe, Dominik; Walter, Björn; Funke, Silke; Hofmann, Eckhard; Schünemann, Danja

    2015-05-22

    In bacteria, membrane proteins are targeted cotranslationally via a signal recognition particle (SRP). During the evolution of higher plant chloroplasts from cyanobacteria, the SRP pathway underwent striking adaptations that enable the posttranslational transport of the abundant light-harvesting chlorophyll-a/b-binding proteins (LHCPs). The conserved 54-kDa SRP subunit in higher plant chloroplasts (cpSRP54) is not bound to an SRP RNA, an essential SRP component in bacteria, but forms a stable heterodimer with the chloroplast-specific cpSRP43. This heterodimeric cpSRP recognizes LHCP and delivers it to the thylakoid membrane whereby cpSRP43 plays a central role. This study shows that the cpSRP system in the green alga Chlamydomonas reinhardtii differs significantly from that of higher plants as cpSRP43 is not complexed to cpSRP54 in Chlamydomonas and cpSRP54 is not involved in LHCP recognition. This divergence is attributed to altered residues within the cpSRP54 tail and the second chromodomain of cpSRP43 that are crucial for the formation of the binding interface in Arabidopsis. These changes are highly conserved among chlorophytes, whereas all land plants contain cpSRP proteins with typical interaction motifs. These data demonstrate that the coevolution of LHCPs and cpSRP43 occurred independently of complex formation with cpSRP54 and that the interaction between cpSRP54 and cpSRP43 evolved later during the transition from chlorophytes to land plants. Furthermore, our data show that in higher plants a heterodimeric form of cpSRP is required for the formation of a low molecular weight transit complex with LHCP. PMID:25833951

  8. Two types of ammonium uncoupling in pea chloroplasts.

    PubMed

    Opanasenko, V K; Vasyukhina, L A; Naydov, I A

    2010-06-01

    The effect of ammonium on ATP synthesis, electron transfer, and light-induced uptake of hydrogen ions in pea chloroplasts was studied. It is shown that the dependence of these reactions on ammonium concentration could be due to effects of two different uncoupling processes. The first process is induced by low ammonium concentrations (<0.2 mM); the second one is observed in the NH(4)Cl concentration interval of 0.5-5.0 mM. The first type of uncoupling is stimulated by palmitic acid or by N,N'-dicyclohexylcarbodiimide, while the second is stimulated by chloroplast thylakoid swelling caused by energy-dependent osmotic gradients. In the presence of the fluorescent dye sulforhodamine B, which does not penetrate through the cell membrane, this swelling causes the dye to enter the lumens. It is supposed that ammonium activates two different routes of cation leakage from the lumen. The first route involves channel proteins, while the second is a mechanosensitive pore that opens in response to osmotic gradients. PMID:20636271

  9. Purification and properties of a pea chloroplast DNA polymerase

    PubMed Central

    McKown, R. L.; Tewari, K. K.

    1984-01-01

    A DNA polymerase has been purified >3,000-fold from the chloroplasts of pea plants by chromatography on DEAE-cellulose, phosphocellulose, single-stranded DNA-agarose, and sedimentation in a glycerol gradient. Electrophoretic analysis on polyacrylamide gels in the presence of sodium dodecyl sulfate indicates that the final fraction contained a single discernible protein band of 90,000 daltons. Gel filtration on Sephacryl S-200 and glycerol gradient sedimentation under nondenaturing conditions demonstrate that the chloroplast DNA polymerase has a native molecular mass of approximately 87,000 daltons. The purified polymerase lacks any associated nuclease activity. The enzyme activity is inhibited by N-ethylmaleimide (74% at 1.0 mM) and ethidium bromide (90% at 0.23 mM) and is resistant to aphidicolin. The purified enzyme is totally dependent on the presence of added DNA, has an absolute requirement for Mg2+ (12 mM optimal), is stimulated by K+ (120 mM optimal), and requires all four deoxynucleoside triphosphates for maximum activity. Native DNA which has been degraded to a limited extent with DNase I is the most efficient template. Images PMID:16593454

  10. Chloroplast protein synthesis: thylakoid bound polysomes synthesize thylakoid proteins

    SciTech Connect

    Hurewitz, J.; Jagendorf, A.T.

    1986-04-01

    Previous work indicated more polysomes bound to pea thylakoids in light than in the dark, in vivo. With isolated intact chloroplasts incubated in darkness, 24 to 74% more RNA was thylakoid-bound at pH 8.3 than at pH 7. Thus the major effect of light in vivo may be due to higher stroma pH. In isolated pea chloroplasts, initiation inhibitors (pactamycin and kanamycin) decreased the extent of RNA binding, and elongation inhibitors (lincomycin and streptomycin) increased it. Thus translation initiation and termination probably control the cycling of bound ribosomes. While only 3 to 6% of total RNA is in bound polysomes the incorporation of /sup 3/H-Leu into thylakoids was proportional to the amount of this bound RNA. When Micrococcal nuclease-treated thylakoids were added to labeled runoff translation products of stroma ribosomes, less than 1% of the label adhered to the added membranes; but 37% of the labeled products made by thylakoid polysomes were bound. These data support the concept that stroma ribosomes are recruited into thylakoid proteins.

  11. The complete chloroplast genome sequence of Dioscorea zingiberensis (Dioscoreceae).

    PubMed

    Zhou, Wen; Chen, Chen; Hua, Wen-Ping; Wang, Zhe-Zhi

    2016-07-01

    Dioscorea zingiberensis (Dioscoreceae) is an important medicinal plant endemic to China. Here, its chloroplast genome sequence is reconstructed from the whole-genome Illumina sequencing data. The circular genome is 153,970 bp in length, and comprises a pair of inverted repeat (IR) regions of 25,491 bp each, a large single-copy (LSC) region of 83,950 bp and a small single-copy (SSC) region of 19,038 bp. The chloroplast genome contains 132 genes, including 86 protein-coding genes (79 PCG species), 8 ribosomal RNA genes (four rRNA species) and 38 transfer RNA genes (30 tRNA species). Out of these genes, 10 harbor a single intron, and 7 contain a couple of introns. The overall A + T content of the whole genome is 62.8%, while the corresponding values of the LSC, SSC and IR regions are 64.9%, 68.8% and 57.0%, respectively. PMID:26066025

  12. Ion Homeostasis in Chloroplasts under Salinity and Mineral Deficiency 1

    PubMed Central

    Schröppel-Meier, Gabriele; Kaiser, Werner M.

    1988-01-01

    Spinach (Spinacia oleracea var “Yates”) plants in hydroponic culture were exposed to stepwise increased concentrations of NaCl or NaNO3 up to a final concentration of 300 millimoles per liter, at constant Ca2+-concentration. Leaf cell sap and extracts from aqueously isolated spinach chloroplasts were analyzed for mineral cations, anions, amino acids, sugars, and quarternary ammonium compounds. Total osmolality of leaf sap and photosynthetic capacity of leaves were also measured. For comparison, leaf sap from salt-treated pea plants was also analyzed. Spinach plants under NaCl or NaNO3 salinity took up large amounts of sodium (up to 400 millimoles per liter); nitrate as the accompanying anion was taken up less (up to 90 millimoles per liter) than chloride (up to 450 millimoles per liter). Under chloride salinity, nitrate content in leaves decreased drastically, but total amino acid concentrations remained constant. This response was much more pronounced (and occurred at lower salt concentrations) in leaves from the glycophyte (pea, Pisum sativum var “Kleine Rheinländerin”) than from moderately salt-tolerant spinach. In spinach, sodium chloride or nitrate taken up into leaves was largely sequestered in the vacuoles; both salts induced synthesis of quarternary ammonium compounds, which were accumulated mainly in chloroplasts (and cytosol). This prevented impairment of metabolism, as indicated by an unchanged photosynthetic capacity of leaves. PMID:16666232

  13. The complete chloroplast genome of Cinnamomum kanehirae Hayata (Lauraceae).

    PubMed

    Wu, Chia-Chen; Ho, Cheng-Kuen; Chang, Shu-Hwa

    2016-07-01

    The complete chloroplast genome of Cinnamomum kanehirae (Hayata), the first to be completely sequenced of Lauraceae family, is presented in this study. The total genome size is 152,700 bp, with a typical circular structure including a pair of inverted repeats (IRa/b) of 20,107 bp of length separated by a large single-copy region (LSC) and a small single-copy region (SSC) of 93,642 bp and 18,844 bp of length, respectively. The overall GC content of the genome is 39.1%. The nucleotide sequence shows 91% identities with Liriodendron tulipifera in the Magnoliaceae. In total, 123 annotated genes consisted of 79 coding genes, eight rRNA genes, and 36 tRNA genes. Among all 79 coding genes, seven genes (rpoC1, atpF, rpl2, ndhB, ndhA, rps16, and rpl2) contain one intron, while two genes (ycf3 and clpP) contain two introns. The maximum likelihood phylogenetic analysis revealed that C. kanehirae chloroplast genome is closely related to Calycanthus fertilis within Laurales order. PMID:26053940

  14. Euglena mitochondria and chloroplasts form tyrosine-O-sulfate

    SciTech Connect

    Saidha, T.; Hanfstingl, U.; Schiff, J.A. )

    1989-04-01

    Mitochondria from light-grown wild-type Euglena gracilis var. bacillaris Cori or dark-grown mutant W{sub 10}BSmL incubated with {sup 35}SO{sub 4}{sup 2{minus}} and ATP, or with {sup 14}C-tyrosine, non-radioactive sulfate and ATP accumulate a labeled compound in the medium. Since this compound shows exact coelectrophoresis with tyrosine-O-sulfate (TOS) at pH 2.0, 5.8 or 8.0., yields sulfate and tyrosine on acid hydrolysis, and treatment with aryl sulfatase from Aerobacter aerogenes yields sulfate and tyrosine but no tyrosine methyl ester, it is identified as TOS. No TOS is found outside purified developing chloroplasts incubated with {sup 35}SO{sub 4}{sup 2{minus}} and ATP, but both chloroplasts and mitochondria form to {sup 35}S externally when incubated with adenosine 3{prime} phosphate 5{prime}phospho({sup 35}S) sulfate (PAP{sup 35}S). Since no tyrosine need be added, tyrosine is provided from endogenous sources. Although TOS is found in the free pool of Euglena cells it cannot be detected in proteins of cells or mucus ruling our sulfation of tyrosine of protein or incorporation of TOS into proteins. The system forming TOS is membrane-bound and may be involved in tyrosine transport.

  15. [Effects of exogenous silicon on active oxygen scavenging systems in chloroplasts of cucumber (Cucumis sativus L.) seedlings under salt stress].

    PubMed

    Qian, Qiong-Qiu; Zai, Wen-San; Zhu, Zhu-Jun; Yu, Jing-Quan

    2006-02-01

    With K(2)SiO(4) (1.0 mmol/L) treatment, the effects of Si on the distribution of Na(+), K(+) to chloroplasts and antioxidant system of cucumber leaves under 50 mmol/L NaCl stress were studied. The results showed that there was a selective transport of K(+) into the chloroplasts so that Na(+) content of chloroplasts was lower under Si treatment (Table 1); H(2)O(2) and MDA contents in chloroplasts were significantly decreased (Fig.1), and the activities of SOD, APX, GR and DHAR were increased simultaneity (Fig.2), and AsA, GSH contents were also increased in chloroplasts of salt-stressed cucumber by additional Si treatment (Fig.3). It may be concluded that Si could decrease absorption of Na(+) and increase ability of active oxygen scavenging in chloroplasts, therefore the injury of chloroplast membrane under salinity stress in cucumber was alleviated. PMID:16477139

  16. Repression of Essential Chloroplast Genes Reveals New Signaling Pathways and Regulatory Feedback Loops in Chlamydomonas[W

    PubMed Central

    Ramundo, Silvia; Rahire, Michèle; Schaad, Olivier; Rochaix, Jean-David

    2013-01-01

    Although reverse genetics has been used to elucidate the function of numerous chloroplast proteins, the characterization of essential plastid genes and their role in chloroplast biogenesis and cell survival has not yet been achieved. Therefore, we developed a robust repressible chloroplast gene expression system in the unicellular alga Chlamydomonas reinhardtii based mainly on a vitamin-repressible riboswitch, and we used this system to study the role of two essential chloroplast genes: ribosomal protein S12 (rps12), encoding a plastid ribosomal protein, and rpoA, encoding the α-subunit of chloroplast bacterial-like RNA polymerase. Repression of either of these two genes leads to the arrest of cell growth, and it induces a response that involves changes in expression of nuclear genes implicated in chloroplast biogenesis, protein turnover, and stress. This response also leads to the overaccumulation of several plastid transcripts and reveals the existence of multiple negative regulatory feedback loops in the chloroplast gene circuitry. PMID:23292734

  17. Low-molecular-weight (4.5S) ribonucleic acid in higher-plant chloroplast ribosomes.

    PubMed Central

    Whitfeld, P R; Leaver, C J; Bottomley, W; Atchison, B

    1978-01-01

    A species of RNA that migrates on 10% (w/v) polyacrylamide gels between 5S and 4S RNA was detected in spinach chloroplasts. This RNA (referred to as 4.5 S RNA) was present in amounts equimolar to the 5S RNA and its molecular weight was estimated to be approx. 33 000. Fractionation of the chloroplast components showed that the 4.5S RNA was associated with the 50 S ribosomal subunit and that it could be removed by washing the ribosomes with a buffer containing 0.01 M-EDTA and 0.5 M-KCl. It did not appear to be a cleavage product of the labile 23 S RNA of spinach chloroplast ribosomes. When 125I-labelled 4.5 S RNA was hybridized to fragments of spinach chloroplast DNA produced by SmaI restriction endonuclease, a single fragment (mol.wt. 1.15 times 10(6)) became labelled. The same DNA fragment also hybridized to chloroplast 5 S RNA and part of the 23 S RNA. It was concluded that the coding sequence for 4.5 S RNA was part of, or immediately adjacent to, the rRNA-gene region in chloroplast DNA . A comparable RNA species was observed in chloroplasts of tobacco and pea leaves. Images Fig. 8. PMID:743229

  18. Comparative Analysis of the Complete Chloroplast Genomes of Five Quercus Species

    PubMed Central

    Yang, Yanci; Zhou, Tao; Duan, Dong; Yang, Jia; Feng, Li; Zhao, Guifang

    2016-01-01

    Quercus is considered economically and ecologically one of the most important genera in the Northern Hemisphere. Oaks are taxonomically perplexing because of shared interspecific morphological traits and intraspecific morphological variation, which are mainly attributed to hybridization. Universal plastid markers cannot provide a sufficient number of variable sites to explore the phylogeny of this genus, and chloroplast genome-scale data have proven to be useful in resolving intractable phylogenetic relationships. In this study, the complete chloroplast genomes of four Quercus species were sequenced, and one published chloroplast genome of Quercus baronii was retrieved for comparative analyses. The five chloroplast genomes ranged from 161,072 bp (Q. baronii) to 161,237 bp (Q. dolicholepis) in length, and their gene organization and order, and GC content, were similar to those of other Fagaceae species. We analyzed nucleotide substitutions, indels, and repeats in the chloroplast genomes, and found 19 relatively highly variable regions that will potentially provide plastid markers for further taxonomic and phylogenetic studies within Quercus. We observed that four genes (ndhA, ndhK, petA, and ycf1) were subject to positive selection. The phylogenetic relationships of the Quercus species inferred from the chloroplast genomes obtained moderate-to-high support, indicating that chloroplast genome data may be useful in resolving relationships in this genus. PMID:27446185

  19. Is chloroplast movement in tobacco plants influenced systemically after local illumination or burning stress?

    PubMed

    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. PMID:19017116

  20. The arc mutants of Arabidopsis with fewer large chloroplasts have a lower mesophyll conductance.

    PubMed

    Weise, Sean E; Carr, David J; Bourke, Ashley M; Hanson, David T; Swarthout, Debbie; Sharkey, Thomas D

    2015-04-01

    Photosynthetic cells of most land plant lineages have numerous small chloroplasts even though most algae, and even the early diverging land plant group the hornworts, tend to have one or a few large chloroplasts. One constraint that small chloroplasts could improve is the resistance to CO2 diffusion from the atmosphere to the chloroplast stroma. We examined the mesophyll conductance (inverse of the diffusion resistance) of mutant Arabidopsis thaliana plants with one or only a few large chloroplasts per cell. The accumulation and replication of chloroplasts (arc) mutants of A. thaliana were studied by model fitting to gas exchange data and (13)CO2 discrimination during carbon fixation. The two methods generally agreed, but the value of the CO2 compensation point of Rubisco (Γ *) used in the model had a large impact on the estimated photosynthetic parameters, including mesophyll conductance. We found that having only a few large chloroplasts per cell resulted in a 25-50 % reduction in the mesophyll conductance at ambient CO2. PMID:25733184

  1. Protection of Chloroplast Membranes by VIPP1 Rescues Aberrant Seedling Development in Arabidopsis nyc1 Mutant

    PubMed Central

    Zhang, Lingang; Kusaba, Makoto; Tanaka, Ayumi; Sakamoto, Wataru

    2016-01-01

    Chlorophylls (Chl) in photosynthetic apparatuses, along with other macromolecules in chloroplasts, are known to undergo degradation during leaf senescence. Several enzymes involved in Chl degradation, by which detoxification of Chl is safely implemented, have been identified. Chl degradation also occurs during embryogenesis and seedling development. Some genes encoding Chl degradation enzymes such as Chl b reductase (CBR) function during these developmental stages. Arabidopsis mutants lacking CBR (NYC1 and NOL) have been reported to exhibit reduced seed storability, compromised germination, and cotyledon development. In this study, we examined aberrant cotyledon development and found that NYC1 is solely responsible for this phenotype. We inferred that oxidative damage of chloroplast membranes caused the aberrant cotyledon. To test the inference, we attempted to trans-complement nyc1 mutant with overexpressing VIPP1 protein that is unrelated to Chl degradation but which supports chloroplast membrane integrity. VIPP1 expression actually complemented the aberrant cotyledon of nyc1, whereas stay-green phenotype during leaf senescence remained. The swollen chloroplasts observed in unfixed cotyledons of nyc1, which are characteristics of chloroplasts receiving envelope membrane damage, were recovered by overexpressing VIPP1. These results suggest that chloroplast membranes are a target for oxidative damage caused by the impairment in Chl degradation. Trans-complementation of nyc1 with VIPP1 also suggests that VIPP1 is useful for protecting chloroplasts against oxidative stress. PMID:27200011

  2. A high-throughput method for detection of DNA in chloroplasts using flow cytometry

    PubMed Central

    Rowan, Beth A; Oldenburg, Delene J; Bendich, Arnold J

    2007-01-01

    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. PMID:17381841

  3. Chloroplast gene arrangement variation within a closely related group of green algae (Trebouxiophyceae, Chlorophyta).

    PubMed

    Letsch, Molly R; Lewis, Louise A

    2012-09-01

    The 22 published chloroplast genomes of green algae, representing sparse taxonomic sampling of diverse lineages that span over one billion years of evolution, each possess a unique gene arrangement. In contrast, many of the >190 published embryophyte (land plant) chloroplast genomes have relatively conserved architectures. To determine the phylogenetic depth at which chloroplast gene rearrangements occur in green algae, a 1.5-4 kb segment of the chloroplast genome was compared across nine species in three closely related genera of Trebouxiophyceae (Chlorophyta). In total, four distinct gene arrangements were obtained for the three genera Elliptochloris, Hemichloris, and Coccomyxa. In Elliptochloris, three distinct chloroplast gene arrangements were detected, one of which is shared with members of its sister genus Hemichloris. Both species of Coccomyxa examined share the fourth arrangement of this genome region, one characterized by very long spacers. Next, the order of genes found in this segment of the chloroplast genome was compared across green algae and land plants. As taxonomic ranks are not equivalent among different groups of organisms, the maximum molecular divergence among taxa sharing a common gene arrangement in this genome segment was compared. Well-supported clades possessing a single gene order had similar phylogenetic depth in green algae and embryophytes. When the dominant gene order of this chloroplast segment in embryophytes was assumed to be ancestral for land plants, the maximum molecular divergence was found to be over two times greater in embryophytes than in trebouxiophyte green algae. This study greatly expands information about chloroplast genome variation in green algae, is the first to demonstrate such variation among congeneric green algae, and further illustrates the fluidity of green algal chloroplast genome architecture in comparison to that of many embryophytes. PMID:22659018

  4. A Brassica napus Lipase Locates at the Membrane Contact Sites Involved in Chloroplast Development

    PubMed Central

    Tan, Xiaoli; Wang, Qiuye; Tian, Baoxia; Zhang, Henan; Lu, Daoli; Zhou, Jia

    2011-01-01

    Background Fatty acids synthesized in chloroplast are transported to endoplasmic reticulum (ER) for triacylglycerols (TAGs) resembling. The development of chloroplast also requires lipids trafficking from ER to chloroplast. The membrane contact sites (MCSs) between ER and chloroplast has been demonstrated to be involved for the trafficking of lipids and proteins. Lipids trafficking between ER and chloroplast is often accompanied by lipids interconversion. However, it is rarely known how lipids interconversion happens during their trafficking. Methodology/Principal Findings We cloned a lipase gene from Brassica napus L., designated as BnCLIP1. Green fluorescence protein (GFP)-tagged BnCLIP1 was shown to locate at the MCSs between ER and chloroplasts in tobacco leaves. Heterogeneous expression of BnCLIP1 in Saccharomyces cerevisiae (pep4) reduced the total amount of fatty acid. Gas chromatography-mass spectrometry (GC-MS) analysis revealed that the truncated BnCLIP1 had a substrate preference for C16:0 lipids in Saccharomyces cerevisiae (pep4). To probe the physiological function of BnCLIP1, two Brassica napus lines with different oil-content were introduced to investigate the transcript patterns of BnCLIP1 during seed development. Intriguingly, the transcript level of BnCLIP1 was found to be immediately up-regulated during the natural seed senescence of both lines; the transcription response of BnCLIP1 in the high oil-content seeds was faster than the lower ones, suggesting a potential role of BnCLIP1 in affecting seed oil synthesis via regulating chloroplast integrity. Further researches showed that chemical disruption of leaf chloroplast also activated the transcription of BnCLIP1. Conclusions/Significance The findings of this study show that BnCLIP1 encodes a lipase, localizes at the MCSs and involves in chloroplast development. PMID:22046373

  5. Unbiased estimation of chloroplast number in mesophyll cells: advantage of a genuine three-dimensional approach

    PubMed Central

    Kubínová, Zuzana

    2014-01-01

    Chloroplast number per cell is a frequently examined quantitative anatomical parameter, often estimated by counting chloroplast profiles in two-dimensional (2D) sections of mesophyll cells. However, a mesophyll cell is a three-dimensional (3D) structure and this has to be taken into account when quantifying its internal structure. We compared 2D and 3D approaches to chloroplast counting from different points of view: (i) in practical measurements of mesophyll cells of Norway spruce needles, (ii) in a 3D model of a mesophyll cell with chloroplasts, and (iii) using a theoretical analysis. We applied, for the first time, the stereological method of an optical disector based on counting chloroplasts in stacks of spruce needle optical cross-sections acquired by confocal laser-scanning microscopy. This estimate was compared with counting chloroplast profiles in 2D sections from the same stacks of sections. Comparing practical measurements of mesophyll cells, calculations performed in a 3D model of a cell with chloroplasts as well as a theoretical analysis showed that the 2D approach yielded biased results, while the underestimation could be up to 10-fold. We proved that the frequently used method for counting chloroplasts in a mesophyll cell by counting their profiles in 2D sections did not give correct results. We concluded that the present disector method can be efficiently used for unbiased estimation of chloroplast number per mesophyll cell. This should be the method of choice, especially in coniferous needles and leaves with mesophyll cells with lignified cell walls where maceration methods are difficult or impossible to use. PMID:24336344

  6. Relationships of wild and domesticated rices (Oryza AA genome species) based upon whole chloroplast genome sequences.

    PubMed

    Wambugu, Peterson W; Brozynska, Marta; Furtado, Agnelo; Waters, Daniel L; Henry, Robert J

    2015-01-01

    Rice is the most important crop in the world, acting as the staple food for over half of the world's population. The evolutionary relationships of cultivated rice and its wild relatives have remained contentious and inconclusive. Here we report on the use of whole chloroplast sequences to elucidate the evolutionary and phylogenetic relationships in the AA genome Oryza species, representing the primary gene pool of rice. This is the first study that has produced a well resolved and strongly supported phylogeny of the AA genome species. The pan tropical distribution of these rice relatives was found to be explained by long distance dispersal within the last million years. The analysis resulted in a clustering pattern that showed strong geographical differentiation. The species were defined in two primary clades with a South American/African clade with two species, O glumaepatula and O longistaminata, distinguished from all other species. The largest clade was comprised of an Australian clade including newly identified taxa and the African and Asian clades. This refined knowledge of the relationships between cultivated rice and the related wild species provides a strong foundation for more targeted use of wild genetic resources in rice improvement and efforts to ensure their conservation. PMID:26355750

  7. Diversification, Biogeographic Pattern, and Demographic History of Taiwanese Scutellaria Species Inferred from Nuclear and Chloroplast DNA

    PubMed Central

    Liao, Pei-Chun

    2012-01-01

    The ragged topography created by orogenesis generates diversified habitats for plants in Taiwan. In addition to colonization from nearby mainland China, high species diversity and endemism of plants is also present in Taiwan. Five of the seven Scutellaria species (Lamiaceae) in Taiwan, for example, are endemic to the island. Hypotheses of multiple sources or in situ radiation have arisen to explain the high endemism of Taiwanese species. In this study, phylogenetic analyses using both nuclear and chloroplast markers revealed the multiple sources of Taiwanese Scutellaria species and confirmed the rapid and recent speciation of endemic species, especially those of the “indica group” composed of S. indica, S. austrotaiwanensis, S. tashiroi, and S. playfairii. The common ancestors of the indica group colonized first in northern Taiwan and dispersed regionally southward and eastward. Climate changes during glacial/interglacial cycles led to gradual colonization and variance events in the ancestors of these species, resulting in the present distribution and genetic differentiation of extant populations. Population decline was also detected in S. indica, which might reflect a bottleneck effect from the glacials. In contrast, the recently speciated endemic members of the indica group have not had enough time to accumulate much genetic variation and are thus genetically insensitive to demographic fluctuations, but the extant lineages were spatially expanded in the coalescent process. This study integrated phylogenetic and population genetic analyses to illustrate the evolutionary history of Taiwanese Scutellaria of high endemism and may be indicative of the diversification mechanism of plants on continental islands. PMID:23226402

  8. Chloroplast DNA phylogeography reveals colonization history of a Neotropical tree, Cedrela odorata L., in Mesoamerica.

    PubMed

    Cavers, S; Navarro, C; Lowe, A J

    2003-06-01

    Spanish Cedar (Cedrela odorata L.) is a globally important timber species which has been severely exploited in Mesoamerica for over 200 years. Using polymerase chain reaction-restriction fragment length polymorphisms, its chloroplast (cp) DNA phylogeography was studied in Mesoamerica with samples from 29 populations in six countries. Five haplotypes were characterized, phylogenetically grouped into three lineages (Northern, Central and Southern). Spatial analysis of ordered genetic distance confirmed deviation from a pattern of isolation by distance. The geographically proximate Northern and Central cpDNA lineages were genetically the most differentiated, with the Southern lineage appearing between them on a minimum spanning tree. However, populations possessing Southern lineage haplotypes occupy distinct moist habitats, in contrast to populations possessing Northern and Central lineage haplotypes which occupy drier and more seasonal habitats. Given the known colonization of the proto-Mesoamerican peninsula by South American flora and fauna prior to the formation of the Isthmus of Panama, it seems most likely that the observed population structure in C. odorata results from repeated colonization of Mesoamerica from South American source populations. Such a model would imply an ancient, pre-Isthmian colonization of a dry-adapted type (possessing the Northern lineage or a prototype thereof), with a secondary colonization via the land bridge. Following this, a more recent (possibly post-Pleistocene) expansion of moist-adapted types possessing the Southern lineage from the south fits the known vegetation history of the region. PMID:12755874

  9. Relationships of wild and domesticated rices (Oryza AA genome species) based upon whole chloroplast genome sequences

    PubMed Central

    Wambugu, Peterson W.; Brozynska, Marta; Furtado, Agnelo; Waters, Daniel L.; Henry, Robert J.

    2015-01-01

    Rice is the most important crop in the world, acting as the staple food for over half of the world’s population. The evolutionary relationships of cultivated rice and its wild relatives have remained contentious and inconclusive. Here we report on the use of whole chloroplast sequences to elucidate the evolutionary and phylogenetic relationships in the AA genome Oryza species, representing the primary gene pool of rice. This is the first study that has produced a well resolved and strongly supported phylogeny of the AA genome species. The pan tropical distribution of these rice relatives was found to be explained by long distance dispersal within the last million years. The analysis resulted in a clustering pattern that showed strong geographical differentiation. The species were defined in two primary clades with a South American/African clade with two species, O glumaepatula and O longistaminata, distinguished from all other species. The largest clade was comprised of an Australian clade including newly identified taxa and the African and Asian clades. This refined knowledge of the relationships between cultivated rice and the related wild species provides a strong foundation for more targeted use of wild genetic resources in rice improvement and efforts to ensure their conservation. PMID:26355750

  10. Chloroplast redox imbalance governs phenotypic plasticity: the "grand design of photosynthesis" revisited.

    PubMed

    Hüner, Norman P A; Bode, Rainer; Dahal, Keshav; Hollis, Lauren; Rosso, Dominic; Krol, Marianna; Ivanov, Alexander G

    2012-01-01

    Sunlight, the ultimate energy source for life on our planet, enters the biosphere as a direct consequence of the evolution of photoautotrophy. Photoautotrophs must balance the light energy absorbed and trapped through extremely fast, temperature-insensitive photochemistry with energy consumed through much slower, temperature-dependent biochemistry and metabolism. The attainment of such a balance in cellular energy flow between chloroplasts, mitochondria and the cytosol is called photostasis. Photoautotrophs sense cellular energy imbalances through modulation of excitation pressure which is a measure of the relative redox state of Q(A), the first stable quinone electron acceptor of photosystem II reaction centers. High excitation pressure constitutes a potential stress condition that can be caused either by exposure to an irradiance that exceeds the capacity of C, N, and S assimilation to utilize the electrons generated from the absorbed energy or by low temperature or any stress that decreases the capacity of the metabolic pathways downstream of photochemistry to utilize photosynthetically generated reductants. The similarities and differences in the phenotypic responses between cyanobacteria, green algae, crop plants, and variegation mutants of Arabidopsis thaliana as a function of cold acclimation and photoacclimation are reconciled in terms of differential responses to excitation pressure and the predisposition of photoautotrophs to maintain photostasis. The various acclimation strategies associated with green algae and cyanobacteria versus winter cereals and A. thaliana are discussed in terms of retrograde regulation and the "grand design of photosynthesis" originally proposed by Arnon (1982). PMID:23230444

  11. Chloroplast redox imbalance governs phenotypic plasticity: the “grand design of photosynthesis” revisited

    PubMed Central

    Hüner, Norman P. A.; Bode, Rainer; Dahal, Keshav; Hollis, Lauren; Rosso, Dominic; Krol, Marianna; Ivanov, Alexander G.

    2012-01-01

    Sunlight, the ultimate energy source for life on our planet, enters the biosphere as a direct consequence of the evolution of photoautotrophy. Photoautotrophs must balance the light energy absorbed and trapped through extremely fast, temperature-insensitive photochemistry with energy consumed through much slower, temperature-dependent biochemistry and metabolism. The attainment of such a balance in cellular energy flow between chloroplasts, mitochondria and the cytosol is called photostasis. Photoautotrophs sense cellular energy imbalances through modulation of excitation pressure which is a measure of the relative redox state of QA, the first stable quinone electron acceptor of photosystem II reaction centers. High excitation pressure constitutes a potential stress condition that can be caused either by exposure to an irradiance that exceeds the capacity of C, N, and S assimilation to utilize the electrons generated from the absorbed energy or by low temperature or any stress that decreases the capacity of the metabolic pathways downstream of photochemistry to utilize photosynthetically generated reductants. The similarities and differences in the phenotypic responses between cyanobacteria, green algae, crop plants, and variegation mutants of Arabidopsis thaliana as a function of cold acclimation and photoacclimation are reconciled in terms of differential responses to excitation pressure and the predisposition of photoautotrophs to maintain photostasis. The various acclimation strategies associated with green algae and cyanobacteria versus winter cereals and A. thaliana are discussed in terms of retrograde regulation and the “grand design of photosynthesis” originally proposed by Arnon (1982). PMID:23230444

  12. Once upon a Time - Chloroplast Protein Import Research from Infancy to Future Challenges.

    PubMed

    Bölter, Bettina; Soll, Jürgen

    2016-06-01

    Protein import into chloroplasts has been a focus of research for several decades. The first publications dealing with this fascinating topic appeared in the 1970s. From the initial realization that many plastid proteins are being encoded for in the nucleus and require transport into their target organelle to the identification of import components in the cytosol, chloroplast envelopes, and stroma, as well as elucidation of some mechanistic details, more fascinating aspects are still being unraveled. With this overview, we present a survey of the beginnings of chloroplast protein import research, the first steps on this winding road, and end with a glimpse into the future. PMID:27142186

  13. Structure and Function of Tomato Leaf Chloroplasts During Ammonium Toxicity 1

    PubMed Central

    Puritch, George S.; Barker, Allen V.

    1967-01-01

    Ammonium toxicity resulted in morphological modifications of tomato leaf chloroplasts. The chloroplasts, which are normally flattened around the protoplast periphery, became ellipsoidally rounded and dispersed through the protoplasm. The first apparent effect of plastid degradation was development of many vesicles from the fretwork. Later the grana lamellae swelled, and some disappeared. Eventually, distinct grana could not be detected. Ammonium accumulation, chlorophyll loss, and photosynthetic decrease occurred simultaneously. Initial changes in these processes preceded the detection of modifications of fine structure; however, each continued with further breakdown of the chloroplasts. Images PMID:16656644

  14. Inactivation of Maize Leaf Phosphoenolpyruvate Carboxylase by the Binding to Chloroplast Membranes 1

    PubMed Central

    Wu, Min-Xian; Wedding, Randolph T.

    1992-01-01

    Phosphoenolpyruvate carboxylase (PEPC) purified from maize (Zea mays L.) leaves associates with maize leaf chloroplast membrane in vitro. The binding of PEPC to the membrane results in enzyme inactivation. A protein isolated from a maize leaf chloroplast membrane preparation inactivated PEPC. Treatment with membrane preparation or with partially purified inactivating protein accelerates PEPC inactivation at low temperature (4°C). Interaction of PEPC with chloroplast membrane or inactivating protein may inactivate the enzyme by influencing dissociation of the enzyme active tetramer. PMID:16652972

  15. Increasing tomato fruit quality by enhancing fruit chloroplast function. A double-edged sword?

    PubMed

    Cocaliadis, Maria Florencia; Fernández-Muñoz, Rafael; Pons, Clara; Orzaez, Diego; Granell, Antonio

    2014-08-01

    Fruits are generally regarded as photosynthate sinks as they rely on energy provided by sugars transported from leaves to carry out the highly demanding processes of development and ripening; eventually these imported photosynthates also contribute to the fruit organoleptic properties. Three recent reports have revealed, however, that transcriptional factors enhancing chloroplast development in fruit may result in higher contents not only of tomato fruit-specialized metabolites but also of sugars. In addition to suggesting new ways to improve fruit quality by fortifying fruit chloroplasts and plastids, these results prompted us to re-evaluate the importance of the contribution of chloroplasts/photosynthesis to fruit development and ripening. PMID:24723405

  16. The complete chloroplast genome sequence of Chikusichloa aquatica (Poaceae: Oryzeae).

    PubMed

    Zhang, Jie; Zhang, Dan; Shi, Chao; Gao, Ju; Gao, Li-Zhi

    2016-07-01

    The complete chloroplast sequence of the Chikusichloa aquatica was determined in this study. The genome consists of 136 563 bp containing a pair of inverted repeats (IRs) of 20 837 bp, which was separated by a large single-copy region and a small single-copy region of 82 315 bp and 33 411 bp, respectively. The C. aquatica cp genome encodes 111 functional genes (71 protein-coding genes, four rRNA genes, and 36 tRNA genes): 92 are unique, while 19 are duplicated in the IR regions. The genic regions account for 58.9% of whole cp genome, and the GC content of the plastome is 39.0%. A phylogenomic analysis showed that C. aquatica is closely related to Rhynchoryza subulata that belongs to the tribe Oryzeae. PMID:26190082

  17. The complete chloroplast genome sequence of Dianthus superbus var. longicalycinus.

    PubMed

    Gurusamy, Raman; Lee, Do-Hyung; Park, SeonJoo

    2016-05-01

    The complete chloroplast genome (cpDNA) sequence of Dianthus superbus var. longicalycinus is an economically important traditional Chinese medicine was reported and characterized. The cpDNA of Dianthus superbus var. longicalycinus is 149,539 bp, with 36.3% GC content. A pair of inverted repeats (IRs) of 24,803 bp is separated by a large single-copy region (LSC, 82,805 bp) and a small single-copy region (SSC, 17,128 bp). It encodes 85 protein-coding genes, 36 tRNA genes and 8 rRNA genes. Of 129 individual genes, 13 genes encoded one intron and three genes have two introns. PMID:25354144

  18. [Study of Chloroplast DNA Polymorphism in the Sunflower (Helianthus L.)].

    PubMed

    Markina, N V; Usatov, A V; Logacheva, M D; Azarin, K V; Gorbachenko, C F; Kornienko, I V; Gavrilova, V A; Tihobaeva, V E

    2015-08-01

    The polymorphism of microsatellite loci of chloroplast genome in six Helianthus species and 46 lines of cultivated sunflower H. annuus (17 CMS lines and 29 Rf-lines) were studied. The differences between species are confined to four SSR loci. Within cultivated forms of the sunflower H. annuus, the polymorphism is absent. A comparative analysis was performed on sequences of the cpDNA inbred line 3629, line 398941 of the wild sunflower, and the American line HA383 H. annuus. As a result, 52 polymorphic loci represented by 27 SSR and 25 SNP were found; they can be used for genotyping of H. annuus samples, including cultural varieties: twelve polymorphic positions, of which eight are SSR and four are SNP. PMID:26601486

  19. Shotgun proteomics analysis on maize chloroplast thylakoid membrane.

    PubMed

    Liu, Xiao-Yu; Wu, Ya-Dan; Shen, Zhi-Ying; Shen, Zhuo; Li, Hua-Hua; Yu, Xiao-Mei; Yan, Xiu-Feng; Guo, Chang-Hong; Wang, Bai-Chen

    2011-01-01

    In this study we initiated a proteomic investigation of the maize thylakoid membrane by using a shotgun proteomic approach based on LC-MS(E). A total of 34 maize thylakoid membrane proteins were identified, the majority of which are primarily involved in photosynthesis, including the light-reaction and carbon assimilation. It is noteworthy that all of the core subunits of the Photosystem II were identified in our search. Proteins involved in other processes, such as iron storage, were also detected in our study. The quantity of each identified protein was also determined. Of interest, we discovered that the amount of the three ATP synthase subunits were not equivalent, suggesting that these proteins perform other functions in addition to ATP synthesis. To our knowledge this is the first extensive proteomic investigation of the maize thylakoid membrane, and will likely enable further study of maize photosynthesis and chloroplast development. PMID:21196305

  20. The complete chloroplast genome of Sinopodophyllum hexandrum (Berberidaceae).

    PubMed

    Li, Huie; Guo, Qiqiang

    2016-07-01

    The complete chloroplast (cp) genome of the Sinopodophyllum hexandrum (Berberidaceae) was determined in this study. The circular genome is 157,940 bp in size, and comprises a pair of inverted repeat (IR) regions of 26,077 bp each, a large single-copy (LSC) region of 86,460 bp and a small single-copy (SSC) region of 19,326 bp. The GC content of the whole cp genome was 38.5%. A total of 133 genes were identified, including 88 protein-coding genes, 37 tRNA genes and eight rRNA genes. The whole cp genome consists of 114 unique genes, and 19 genes are duplicated in the IR regions. The phylogenetic analysis revealed that S. hexandrum is closely related to Nandina domestica within the family Berberidaceae. PMID:26704891

  1. The complete chloroplast genome of salt cress (Eutrema salsugineum).

    PubMed

    Guo, Xinyi; Hao, Guoqian; Ma, Tao

    2016-07-01

    The complete chloroplast (cp) sequence of the salt cress (Eutrema salsugineum), a plant well-adapted to salt stress, was presented in this study. The circular molecule is 153,407 bp in length and exhibit a typical quadripartite structure containing an 83,894 bp large single copy (LSC) region, a 17,607 bp small single copy (SSC) region, and the two 25,953 bp inverted repeats (IRs). The salt cress cp genome contains 135 known genes, including 87 protein-coding genes, 8 ribosomal RNA genes, and 40 tRNA genes; 21 of these are located in the inverted repeat region. As expected, phylogenetic analysis support the idea that E. salsugineum is sister to Brassiceae species within the Brassicaceae family. PMID:26114321

  2. The complete chloroplast genomes of Cannabis sativa and Humulus lupulus.

    PubMed

    Vergara, Daniela; White, Kristin H; Keepers, Kyle G; Kane, Nolan C

    2016-09-01

    Cannabis and Humulus are sister genera comprising the entirety of the Cannabaceae sensu stricto, including C. sativa L. (marijuana, hemp), and H. lupulus L. (hops) as two economically important crops. These two plants have been used by humans for many purposes including as a fiber, food, medicine, or inebriant in the case of C. sativa, and as a flavoring component in beer brewing in the case of H. lupulus. In this study, we report the complete chloroplast genomes for two distinct hemp varieties of C. sativa, Italian "Carmagnola" and Russian "Dagestani", and one Czech variety of H. lupulus "Saazer". Both C. sativa genomes are 153 871 bp in length, while the H. lupulus genome is 153 751 bp. The genomes from the two C. sativa varieties differ in 16 single nucleotide polymorphisms (SNPs), while the H. lupulus genome differs in 1722 SNPs from both C. sativa cultivars. PMID:26329384

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

    SciTech Connect

    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.

  4. The complete chloroplast genome sequence of Anoectochilus emeiensis.

    PubMed

    Zhu, Shuying; Niu, Zhitao; Yan, Wenjin; Xue, Qingyun; Ding, Xiaoyu

    2016-09-01

    The complete chloroplast (cp) genome sequence of Anoectochilus emeiensis, an extremely endangered medical plant with important economic value, was determined and characterized. The genome size was 152 650 bp, containing a pair of inverted repeats (IRs) (26 319 bp) which were separated by a large single copy (LSC) (82 670 bp) and a small single copy (SSC) (17 342 bp). The cpDNA of A. emeiensis contained 113 unique genes, including 79 protein coding genes, 30 tRNA genes and 4 rRNA genes. Among them, 18 genes contained one or two introns. The overall AT content of the genome was 63.1%. PMID:26403535

  5. The whole chloroplast genomes of two Eutrema species (Brassicaceae).

    PubMed

    Hao, Guoqian; Bi, Hao; Li, Yuanshuo; He, Qi; Ma, Yazhen; Guo, Xinyi; Ma, Tao

    2016-09-01

    In this study, we determined the complete chloroplast genomes from two crucifer species of the Eutrema genus. The sizes of the two cp genomes were 153 948 bp (E. yunnanense) and 153 876 bp (E. heterophyllum). Both genomes have the typical quadripartite structure consisting of a large single copy region, a small single copy region and two inverted repeats. Gene contents and their relative positions of the 132 individual genes (87 protein-coding genes, eight rRNA, and 37 tRNA genes) of either genome were identical to each other. Phylogenetic analysis supports the idea that the currently recognized Eutrema genus is monophyletic and that E. salsugineum and Schrenkiella parvula evolved salt tolerance independently. PMID:26329763

  6. The complete chloroplast genome sequence of Euonymus japonicus (Celastraceae).

    PubMed

    Choi, Kyoung Su; Park, SeonJoo

    2016-09-01

    The complete chloroplast (cp) genome sequence of the Euonymus japonicus, the first sequenced of the genus Euonymus, was reported in this study. The total length was 157 637 bp, containing a pair of 26 678 bp inverted repeat region (IR), which were separated by small single copy (SSC) region and large single copy (LSC) region of 18 340 bp and 85 941 bp, respectively. This genome contains 107 unique genes, including 74 coding genes, four rRNA genes, and 29 tRNA genes. Seventeen genes contain intron of E. japonicus, of which three genes (clpP, ycf3, and rps12) include two introns. The maximum likelihood (ML) phylogenetic analysis revealed that E. japonicus was closely related to Manihot and Populus. PMID:26407184

  7. Protein phosphorylation in chloroplasts - a survey of phosphorylation targets.

    PubMed

    Baginsky, Sacha

    2016-06-01

    The development of new software tools, improved mass spectrometry equipment, a suite of optimized scan types, and better-quality phosphopeptide affinity capture have paved the way for an explosion of mass spectrometry data on phosphopeptides. Because phosphoproteomics achieves good sensitivity, most studies use complete cell extracts for phosphopeptide enrichment and identification without prior enrichment of proteins or subcellular compartments. As a consequence, the phosphoproteome of cell organelles often comes as a by-product from large-scale studies and is commonly assembled from these in meta-analyses. This review aims at providing some guidance on the limitations of meta-analyses that combine data from analyses with different scopes, reports on the current status of knowledge on chloroplast phosphorylation targets, provides initial insights into phosphorylation site conservation in different plant species, and highlights emerging information on the integration of gene expression with metabolism and photosynthesis by means of protein phosphorylation. PMID:26969742

  8. Chloroplast RNA editing going extreme: more than 3400 events of C-to-U editing in the chloroplast transcriptome of the lycophyte Selaginella uncinata

    PubMed Central

    Oldenkott, Bastian; Yamaguchi, Kazuo; Tsuji-Tsukinoki, Sumika; Knie, Nils

    2014-01-01

    RNA editing in chloroplasts and mitochondria of land plants differs significantly in abundance. For example, some 200–500 sites of cytidine-to-uridine RNA editing exist in flowering plant mitochondria as opposed to only 30–50 such C-to-U editing events in their chloroplasts. In contrast, we predicted significantly more chloroplast RNA editing for the protein-coding genes in the available complete plastome sequences of two species of the spike moss genus Selaginella (Lycopodiophyta). To evaluate these predictions we investigated the Selaginella uncinata chloroplast transcriptome. Our exhaustive cDNA studies identified the extraordinary number of 3415 RNA-editing events, exclusively of the C-to-U type, in the 74 mRNAs encoding intact reading frames in the S. uncinata chloroplast. We find the overwhelming majority (61%) of the 428 silent editing events leaving codon meanings unaltered directly neighboring other editing events, possibly suggesting a sterically more flexible RNA-editing deaminase activity in Selaginella. No evidence of RNA editing was found for tRNAs or rRNAs but we identified a total of 74 editing sites in cDNA sequences of four group II introns (petBi6g2, petDi8g2, ycf3i124g2, and ycf3i354g2) retained in partially matured transcripts, which strongly contribute to improved base-pairing in the intron secondary structures as a likely prerequisite for their splicing. PMID:25142065

  9. Metallothionein expression in chloroplasts enhances mercury accumulation and phytoremediation capability

    PubMed Central

    Ruiz, Oscar N.; Alvarez, Derry; Torres, Cesar; Roman, Laura; Daniell, Henry

    2015-01-01

    Summary Genetic engineering to enhance mercury phytoremediation has been accomplished by expression of the merAB genes that protects the cell by converting Hg[II] into Hg[0] which volatilizes from the cell. A drawback of this approach is that toxic Hg is released back into the environment. A better phytoremediation strategy would be to accumulate mercury inside plants for subsequent retrieval. We report here the development of a transplastomic approach to express the mouse metallothionein gene (mt1) and accumulate mercury in high concentrations within plant cells. Real-time PCR analysis showed that up to 1284 copies of the mt1 gene were found per cell when compared with 1326 copies of the 16S rrn gene, thereby attaining homoplasmy. Past studies in chloroplast transformation used qualitative Southern blots to evaluate indirectly transgene copy number, whereas we used real-time PCR for the first time to establish homoplasmy and estimate transgene copy number and transcript levels. The mt1 transcript levels were very high with 183 000 copies per ng of RNA or 41% the abundance of the 16S rrn transcripts. The transplastomic lines were resistant up to 20 μm mercury and maintained high chlorophyll content and biomass. Although the transgenic plants accumulated high concentrations of mercury in all tissues, leaves accumulated up to 106 ng, indicating active phytoremediation and translocation of mercury. Such accumulation of mercury in plant tissues facilitates proper disposal or recycling. This study reports, for the first time, the use of metallothioniens in plants for mercury phytoremediation. Chloroplast genetic engineering approach is useful to express metal-scavenging proteins for phytoremediation. PMID:21518240

  10. Arabidopsis thaliana DNA gyrase is targeted to chloroplasts and mitochondria

    PubMed Central

    Wall, Melisa K.; Mitchenall, Lesley A.; Maxwell, Anthony

    2004-01-01

    DNA gyrase is the bacterial DNA topoisomerase (topo) that supercoils DNA by using the free energy of ATP hydrolysis. The enzyme, an A2B2 tetramer encoded by the gyrA and gyrB genes, catalyses topological changes in DNA during replication and transcription, and is the only topo that is able to introduce negative supercoils. Gyrase is essential in bacteria and apparently absent from eukaryotes and is, consequently, an important target for antibacterial agents (e.g., quinolones and coumarins). We have identified four putative gyrase genes in the model plant Arabidopsis thaliana; one gyrA and three gyrB homologues. DNA gyrase protein A (GyrA) has a dual translational initiation site targeting the mature protein to both chloroplasts and mitochondria, and there are individual targeting sequences for two of the DNA gyrase protein B (GyrB) homologues. N-terminal fusions of the organellar targeting sequences to GFPs support the hypothesis that one enzyme is targeted to the chloroplast and another to the mitochondrion, which correlates with supercoiling activity in isolated organelles. Treatment of seedlings and cultured cells with gyrase-specific drugs leads to growth inhibition. Knockout of A. thaliana gyrA is embryo-lethal whereas knockouts in the gyrB genes lead to seedling-lethal phenotypes or severely stunted growth and development. The A. thaliana genes have been cloned in Escherichia coli and found to complement gyrase temperature-sensitive strains. This report confirms the existence of DNA gyrase in eukaryotes and has important implications for drug targeting, organelle replication, and the evolution of topos in plants. PMID:15136745

  11. Thylakoid Polypeptides of Light and Dark Aged Chloroplasts 12

    PubMed Central

    dos Santos, Cesar P.; Hall, David O.

    1982-01-01

    Spinach (Spinacia oleracea) chloroplasts were aged at 4°C under red light and in the dark. The electron transport activity was monitored together with the thylakoid polypeptide patterns in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The light-induced decay of photosystem II (PSII) activity (half-life, about 4 hours) was correlated with a decrease in polypeptides with apparent molecular weights of 36, 48, and 50 kilodaltons. There was very little decay of photosystem I (PSI) activity until after 8 hours illumination. Prior freezing of the chloroplasts enhanced the decrease in PSI activity which was correlated with chlorophyll-protein complex I (CPI) disappearance and an increase in a polypeptide with apparent molecular weight of 60 kilodalton. No variations were detected in the light-harvesting chlorophyll a/b protein. In the dark, the decay of PSII started at 4 to 6 hours and showed a half life of about 30 hours. PSI activity decay (half life about 6 days) occurred simultaneously with the disappearance of CPI. The use of bovine serum albumin (30 mg/mg of chlorophyll) in the light-induced decay experiments increased the stability of PSII more than 2-fold; in the dark experiments, the stability of both photosystems was also more than doubled and the stability of the CPI complex was considerably improved. Comparative electrophoresis of the purified proteins indicated no changes in the cytochrome f band or in the subunits of the ATPase coupling factor during the light-induced decay experiments. Heating of purified PSI particles prior to electrophoresis showed that the 60 kilodaltons polypeptide increased with the disappearance of CPI. Images Fig. 5 Fig. 7 Fig. 8 PMID:16662578

  12. Events Surrounding the Early Development of Euglena Chloroplasts

    PubMed Central

    Vaisberg, Abraham J.; Schiff, Jerome A.; Li, Lynn; Freedman, Zachary

    1976-01-01

    d(−)threo-Chloramphenicol blocks chlorophyll and plastid protein synthesis in Euglena. During chloroplast development in white light, but not in red, the cells escape from chloramphenicol inhibition and chlorophyll formation is restored. Concomitantly, chloramphenicol is reduced. Reduction of chloramphenicol in an enzyme extract from Euglena requires NADPH and ferredoxin for maximal activity. Methyl viologen replaces ferredoxin, and when chemically reduced, ferredoxin or methyl viologen reduces chloramphenicol directly. This suggests that the enzyme involved is ferredoxin-NADP reductase. In agreement, crude extracts from wild type and W3BUL, a mutant lacking detectable plastids and plastid DNA, when separated on acrylamide gels, show a single band which reduces methyl viologen with NADPH, and its mobility is similar in wild type and in mutant W3BUL. The reductase is inducible by light and increases 3-fold in wild type in white or red light and 1.5-fold in W3BUL in white light. DCMU does not block chloramphenicol reduction in vivo indicating that electrons originate from sources other than photosynthetic electron transport. We infer that chloramphenicol is reduced by ferredoxin which receives electrons via ferredoxin-NADP reductase. The limiting step is not the enzyme but the source of reducing power which can be supplied from the cytoplasm, probably under control of the blue light receptor. Ferredoxin and ferredoxin NADP reductase appear to be coded in the nuclear genome, synthesized on cytoplasmic ribosomes, and join a group of enzymes which cannot be precisely localized, since they may be active anywhere from their site of synthesis in the cytoplasm to their place of deposition in the chloroplast. PMID:16659534

  13. Chloroplast DNA evolution and phylogenetic relationships in Lycopersicon

    PubMed Central

    Palmer, Jeffrey D.; Zamir, Daniel

    1982-01-01

    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. Images PMID:16593219

  14. Small effect of fragmentation on the genetic diversity of Dalbergia monticola, an endangered tree species of the eastern forest of Madagascar, detected by chloroplast and nuclear microsatellites

    PubMed Central

    Andrianoelina, O.; Favreau, B.; Ramamonjisoa, L.; Bouvet, J.-M.

    2009-01-01

    Background and Aims The oriental forest ecosystem in Madagascar has been seriously impacted by fragmentation. The pattern of genetic diversity was analysed on a tree species, Dalbergia monticola, which plays an important economic role in Madagascar and is one of the many endangered tree species in the eastern forest. Methods Leaves from 546 individuals belonging to 18 small populations affected by different levels of fragmentation were genotyped using eight nuclear (nuc) and three chloroplast (cp) microsatellite markers. Key Results For nuclear microsatellites, allelic richness (R) and heterozygosity (He,nuc) differed between types of forest: R = 7·36 and R = 9·55, He,nuc = 0·64 and He,nuc = 0·80 in fragmented and non-fragmented forest, respectively, but the differences were not significant. Only the mean number of alleles (Na,nuc) and the fixation index FIS differed significantly: Na,nuc = 9·41 and Na,nuc = 13·18, FIS = 0·06 and FIS = 0·15 in fragmented and non-fragmented forests, respectively. For chloroplast microsatellites, estimated genetic diversity was higher in non-fragmented forest, but the difference was not significant. No recent bottleneck effect was detected for either population. Overall differentiation was low for nuclear microsatellites (FST,nuc = 0·08) and moderate for chloroplast microsatellites (FST,cp = 0·49). A clear relationship was observed between genetic and geographic distance (r = 0·42 P < 0·01 and r = 0·42 P = 0·03 for nuclear and chloroplast microsatellites, respectively), suggesting a pattern of isolation by distance. Analysis of population structure using the neighbor-joining method or Bayesian models separated southern populations from central and northern populations with nuclear microsatellites, and grouped the population according to regions with chloroplast microsatellites, but did not separate the fragmented populations. Conclusions Residual diversity and genetic structure of populations of D. monticola in

  15. Photoreduction of α-Ketoglutarate to Glutamate by Vicia faba Chloroplasts 1

    PubMed Central

    Givan, Curtis V.; Givan, Alice L.; Leech, Rachel M.

    1970-01-01

    Intact chloroplasts isolated from leaves of Vicia faba L. var. the Sutton show a decline in the endogenous level of α-ketoglutarate upon illumination. α-Ketoglutarate supplied to the chloroplasts is similarly utilized in this light-dependent reaction, and its consumption is paralleled by a concomitant increase in the level of glutamate. There is no photostimulation of glutamate synthesis in chloroplasts broken by osmotic shock, but it can be somewhat restored by addition of ferredoxin and NADP. These results suggest that in the isolated chloroplast the synthesis of glutamate from α-ketoglutarate is regulated by the availability of reduced pyridine nucleotide generated by photosynthetic electron transport. This conclusion is supported by the finding of an apparent competition between the photoreduction of phosphoglycerate to triose phosphate and the photoutilization of α-ketoglutarate. PMID:16657357

  16. Activity of phosphatidylcholine-transfer protein from spinach (Spinacia oleracea) leaves with mitochondria and chloroplasts.

    PubMed

    Julienne, M; Vergnolle, C; Kader, J C

    1981-09-01

    A low-molecular-weight protein catalysing the transfer of phosphatidylcholine from liposomes to mitochondria and chloroplasts has been isolated from spinach (Spinacia oleracea) by chromatography on Sephadex G-75. PMID:7325986

  17. The complete chloroplast genome of Eleutherococcus gracilistylus (W.W.Sm.) S.Y.Hu (Araliaceae).

    PubMed

    Kim, Kyunghee; Lee, Junki; Lee, Sang-Choon; Kim, Nam-Hoon; Jang, Woojong; Kim, Soonok; Sung, Sangmin; Lee, Jungho; Yang, Tae-Jin

    2016-09-01

    Eleutherococcus gracilistylus is a plant species that is close to E. senticosus, a famous medicinal plant called Siberian ginseng. The complete chloroplast genome sequence of the E. gracilistylus was determined by de novo assembly using whole genome next generation sequences. The chloroplast genome of E. gracilistylus was 156 770 bp long and showed distinct four partite structures such as a large single copy region of 86 729 bp, a small single copy region of 18 175 bp, and a pair of inverted repeat regions of 25 933 bp. The overall GC contents of the genome sequence were 36.8%. The chloroplast genome of E. gracilistylus contains 79 protein-coding sequences, 30 tRNA genes, and four rRNA genes. The phylogenetic analysis with the reported chloroplast genomes confirmed close taxonomical relationship of E. gracilistylus with E. senticosus. PMID:26358682

  18. In Vivo Quantification of Peroxisome Tethering to Chloroplasts in Tobacco Epidermal Cells Using Optical Tweezers.

    PubMed

    Gao, Hongbo; Metz, Jeremy; Teanby, Nick A; Ward, Andy D; Botchway, Stanley W; Coles, Benjamin; Pollard, Mark R; Sparkes, Imogen

    2016-01-01

    Peroxisomes are highly motile organelles that display a range of motions within a short time frame. In static snapshots, they can be juxtaposed to chloroplasts, which has led to the hypothesis that they are physically interacting. Here, using optical tweezers, we tested the dynamic physical interaction in vivo. Using near-infrared optical tweezers combined with TIRF microscopy, we were able to trap peroxisomes and approximate the forces involved in chloroplast association in vivo in tobacco (Nicotiana tabacum) and observed weaker tethering to additional unknown structures within the cell. We show that chloroplasts and peroxisomes are physically tethered through peroxules, a poorly described structure in plant cells. We suggest that peroxules have a novel role in maintaining peroxisome-organelle interactions in the dynamic environment. This could be important for fatty acid mobilization and photorespiration through the interaction with oil bodies and chloroplasts, highlighting a fundamentally important role for organelle interactions for essential biochemistry and physiological processes. PMID:26518344

  19. Comparative analyses of chloroplast genome data representing nine green algae in Sphaeropleales (Chlorophyceae, Chlorophyta).

    PubMed

    Fučíková, Karolina; Lewis, Louise A; Lewis, Paul O

    2016-06-01

    The chloroplast genomes of green algae are highly variable in their architecture. In this article we summarize gene content across newly obtained and published chloroplast genomes in Chlorophyceae, including new data from nine of species in Sphaeropleales (Chlorophyceae, Chlorophyta). We present genome architecture information, including genome synteny analysis across two groups of species. Also, we provide a phylogenetic tree obtained from analysis of gene order data for species in Chlorophyceae with fully sequenced chloroplast genomes. Further analyses and interpretation of the data can be found in "Chloroplast phylogenomic data from the green algal order Sphaeropleales (Chlorophyceae, Chlorophyta) reveal complex patterns of sequence evolution" (Fučíková et al., In review) [1]. PMID:27054159

  20. Sequence analysis and protein import studies of an outer chloroplast envelope polypeptide.

    PubMed Central

    Salomon, M; Fischer, K; Flügge, U I; Soll, J

    1990-01-01

    A chloroplast outer envelope membrane protein was cloned and sequenced and from the sequence it was possible to deduce a polypeptide of 6.7 kDa. It has only one membrane-spanning region; the C terminus extends into the cytosol, whereas the N terminus is exposed to the space between the two envelope membranes. The protein was synthesized in an in vitro transcription-translation system to study its routing into isolated chloroplasts. The import studies revealed that the 6.7-kDa protein followed a different and heretofore undescribed translocation pathway in the respect that (i) it does not have a cleavable transit sequence, (ii) it does not require ATP hydrolysis for import, and (iii) protease-sensitive components that are responsible for recognition of precursor proteins destined for the inside of the chloroplasts are not involved in routing the 6.7-kDa polypeptide to the outer chloroplast envelope. Images PMID:2377616

  1. Regulation of chloroplast number and DNA synthesis in higher plants. Final report

    SciTech Connect

    Mullet, J.E.

    1995-11-10

    The long term objective of this research is to understand the process of chloroplast development and its coordination with leaf development in higher plants. This is important because the photosynthetic capacity of plants is directly related to leaf and chloroplast development. This research focuses on obtaining a detailed description of leaf development and the early steps in chloroplast development including activation of plastid DNA synthesis, changes in plastid DNA copy number, activation of chloroplast transcription and increases in plastid number per cell. The grant will also begin analysis of specific biochemical mechanisms by isolation of the plastid DNA polymerase, and identification of genetic mutants which are altered in their accumulation of plastid DNA and plastid number per cell.

  2. Sensing and signaling of oxidative stress in chloroplasts by inactivation of the SAL1 phosphoadenosine phosphatase.

    PubMed

    Chan, Kai Xun; Mabbitt, Peter D; Phua, Su Yin; Mueller, Jonathan W; Nisar, Nazia; Gigolashvili, Tamara; Stroeher, Elke; Grassl, Julia; Arlt, Wiebke; Estavillo, Gonzalo M; Jackson, Colin J; Pogson, Barry J

    2016-08-01

    Intracellular signaling during oxidative stress is complex, with organelle-to-nucleus retrograde communication pathways ill-defined or incomplete. Here we identify the 3'-phosphoadenosine 5'-phosphate (PAP) phosphatase SAL1 as a previously unidentified and conserved oxidative stress sensor in plant chloroplasts. Arabidopsis thaliana SAL1 (AtSAL1) senses changes in photosynthetic redox poise, hydrogen peroxide, and superoxide concentrations in chloroplasts via redox regulatory mechanisms. AtSAL1 phosphatase activity is suppressed by dimerization, intramolecular disulfide formation, and glutathionylation, allowing accumulation of its substrate, PAP, a chloroplast stress retrograde signal that regulates expression of plastid redox associated nuclear genes (PRANGs). This redox regulation of SAL1 for activation of chloroplast signaling is conserved in the plant kingdom, and the plant protein has evolved enhanced redox sensitivity compared with its yeast ortholog. Our results indicate that in addition to sulfur metabolism, SAL1 orthologs have evolved secondary functions in oxidative stress sensing in the plant kingdom. PMID:27432987

  3. PCR BASED MOLECULAR MARKERS THAT DISTINGUISH MAIZE CHLOROPLAST AND MITOCHONDRIAL GENOMES FROM TRIPSACUM

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Progeny from interspecific and intergeneric cross hybridizations result in novel nuclear-cytoplasmic genetic interactions. In contrast to the usually biparentally inherited nuclear genome, inheritance of mitochondrial and chloroplast genomes will vary depending on the mechanisms regulating transmis...

  4. Regulation of chloroplast number and DNA synthesis in higher plants. Final report

    SciTech Connect

    Mullet, J.E.

    1995-11-10

    The long term objective of this research is to understand the process of chloroplast development and its coordination with leaf development in higher plants. This is important because the photosynthetic capacity of plants is directly related to leaf and chloroplast development. This research focuses on obtaining a detailing description of leaf development and the early steps in chloroplast development including activation of plastid DNA synthesis, changes in plastid DNA copy number, activation of chloroplast transcription and increases in plastid number per cell. The grant will also begin analysis of specific biochemical mechanisms by isolation of the plastid DNA polymerase, and identification of genetic mutants which are altered in their accumulation of plastid DNA and plastid number per cell.

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

    PubMed

    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. PMID:26969728

  6. The complete chloroplast genome sequence of an important medicinal plant Cynanchum wilfordii (Maxim.) Hemsl. (Apocynaceae).

    PubMed

    Park, Hyun-Seung; Kim, Kyu-Yeob; Kim, Kyunghee; Lee, Sang-Choon; Lee, Junki; Seong, Rack Seon; Shim, Young Hun; Sung, Sang Hyun; Yang, Tae-Jin

    2016-09-01

    Cynanchum wilfordii (Maxim.) Hemsl. is a traditional medicinal herb belonging to the Asclepiadoideae subfamily, whose dried roots have been used as traditional medicine in Asia. The complete chloroplast genome of C. wilfordii was generated by de novo assembly using the small amount of whole genome sequencing data. The chloroplast genome of C. wilfordii was 161 241 bp long, composed of large single copy region (91 995 bp), small single copy region (19 930 bp) and a pair of inverted repeat regions (24 658 bp). The overall GC contents of the chloroplast genome was 37.8%. A total of 114 genes were annotated, which included 80 protein-coding genes, 30 tRNA genes and 4 rRNA genes. Phylogenetic analysis with the reported chloroplast genomes revealed that C. wilfordii is most closely related to Asclepias nivea (Caribbean milkweed) and Asclepias syriaca (common milkweed) within the Asclepiadoideae subfamily. PMID:26358391

  7. Frataxin Is Localized to Both the Chloroplast and Mitochondrion and Is Involved in Chloroplast Fe-S Protein Function in Arabidopsis

    PubMed Central

    Turowski, Valeria R.; Aknin, Cindy; Maliandi, Maria V.; Buchensky, Celeste; Leaden, Laura; Peralta, Diego A.; Busi, Maria V.; Araya, Alejandro; Gomez-Casati, Diego F.

    2015-01-01

    Frataxin plays a key role in eukaryotic cellular iron metabolism, particularly in mitochondrial heme and iron-sulfur (Fe-S) cluster biosynthesis. However, its precise role has yet to be elucidated. In this work, we studied the subcellular localization of Arabidopsis frataxin, AtFH, using confocal microscopy, and found a novel dual localization for this protein. We demonstrate that plant frataxin is targeted to both the mitochondria and the chloroplast, where it may play a role in Fe-S cluster metabolism as suggested by functional studies on nitrite reductase (NIR) and ferredoxin (Fd), two Fe-S containing chloroplast proteins, in AtFH deficient plants. Our results indicate that frataxin deficiency alters the normal functioning of chloroplasts by affecting the levels of Fe, chlorophyll, and the photosynthetic electron transport chain in this organelle. PMID:26517126

  8. Uncovering the Protein Lysine and Arginine Methylation Network in Arabidopsis Chloroplasts

    PubMed Central

    Mininno, Morgane; Brugière, Sabine; Gilgen, Annabelle; Ma, Sheng; Mazzoleni, Meryl; Gigarel, Océane; Martin-Laffon, Jacqueline; Ferro, Myriam; Ravanel, Stéphane

    2014-01-01

    Post-translational modification of proteins by the addition of methyl groups to the side chains of Lys and Arg residues is proposed to play important roles in many cellular processes. In plants, identification of non-histone methylproteins at a cellular or subcellular scale is still missing. To gain insights into the extent of this modification in chloroplasts we used a bioinformatics approach to identify protein methyltransferases targeted to plastids and set up a workflow to specifically identify Lys and Arg methylated proteins from proteomic data used to produce the Arabidopsis chloroplast proteome. With this approach we could identify 31 high-confidence Lys and Arg methylation sites from 23 chloroplastic proteins, of which only two were previously known to be methylated. These methylproteins are split between the stroma, thylakoids and envelope sub-compartments. They belong to essential metabolic processes, including photosynthesis, and to the chloroplast biogenesis and maintenance machinery (translation, protein import, division). Also, the in silico identification of nine protein methyltransferases that are known or predicted to be targeted to plastids provided a foundation to build the enzymes/substrates relationships that govern methylation in chloroplasts. Thereby, using in vitro methylation assays with chloroplast stroma as a source of methyltransferases we confirmed the methylation sites of two targets, plastid ribosomal protein L11 and the β-subunit of ATP synthase. Furthermore, a biochemical screening of recombinant chloroplastic protein Lys methyltransferases allowed us to identify the enzymes involved in the modification of these substrates. The present study provides a useful resource to build the methyltransferases/methylproteins network and to elucidate the role of protein methylation in chloroplast biology. PMID:24748391

  9. Uncovering the protein lysine and arginine methylation network in Arabidopsis chloroplasts.

    PubMed

    Alban, Claude; Tardif, Marianne; Mininno, Morgane; Brugière, Sabine; Gilgen, Annabelle; Ma, Sheng; Mazzoleni, Meryl; Gigarel, Océane; Martin-Laffon, Jacqueline; Ferro, Myriam; Ravanel, Stéphane

    2014-01-01

    Post-translational modification of proteins by the addition of methyl groups to the side chains of Lys and Arg residues is proposed to play important roles in many cellular processes. In plants, identification of non-histone methylproteins at a cellular or subcellular scale is still missing. To gain insights into the extent of this modification in chloroplasts we used a bioinformatics approach to identify protein methyltransferases targeted to plastids and set up a workflow to specifically identify Lys and Arg methylated proteins from proteomic data used to produce the Arabidopsis chloroplast proteome. With this approach we could identify 31 high-confidence Lys and Arg methylation sites from 23 chloroplastic proteins, of which only two were previously known to be methylated. These methylproteins are split between the stroma, thylakoids and envelope sub-compartments. They belong to essential metabolic processes, including photosynthesis, and to the chloroplast biogenesis and maintenance machinery (translation, protein import, division). Also, the in silico identification of nine protein methyltransferases that are known or predicted to be targeted to plastids provided a foundation to build the enzymes/substrates relationships that govern methylation in chloroplasts. Thereby, using in vitro methylation assays with chloroplast stroma as a source of methyltransferases we confirmed the methylation sites of two targets, plastid ribosomal protein L11 and the β-subunit of ATP synthase. Furthermore, a biochemical screening of recombinant chloroplastic protein Lys methyltransferases allowed us to identify the enzymes involved in the modification of these substrates. The present study provides a useful resource to build the methyltransferases/methylproteins network and to elucidate the role of protein methylation in chloroplast biology. PMID:24748391

  10. THE OCCURRENCE AND SIGNIFICANCE OF ENDOSYMBIOTIC CHLOROPLASTS IN THE DIGESTIVE GLANDS OF HERBIVOROUS OPISTHOBRANCHS(1).

    PubMed

    Taylor, D L

    1967-12-01

    Intact algal chloroplasts have been found in the digestive glands of 5 species of Opisthobranchia belonging to the order Saccoglossa. Preliminary studies on 3 of these confirm their endosymbiotic nature. It is suggested that the occurrence of these endosymbiotic organelles may be widespread among related species of Saccoglossa. Their independent functional existence supports the view that chloroplasts possess a system of nonchromosomal inheritance. PMID:27065036

  11. The Conserved Endoribonuclease YbeY Is Required for Chloroplast Ribosomal RNA Processing in Arabidopsis1

    PubMed Central

    Liu, Jinwen; Zhou, Wenbin; Liu, Guifeng; Yang, Chuanping; Sun, Yi; Wu, Wenjuan; Cao, Shenquan; Wang, Chong; Hai, Guanghui; Wang, Zhifeng; Bock, Ralph; Huang, Jirong

    2015-01-01

    Maturation of chloroplast ribosomal RNAs (rRNAs) comprises several endoribonucleolytic and exoribonucleolytic processing steps. However, little is known about the specific enzymes involved and the cleavage steps they catalyze. Here, we report the functional characterization of the single Arabidopsis (Arabidopsis thaliana) gene encoding a putative YbeY endoribonuclease. AtYbeY null mutants are seedling lethal, indicating that AtYbeY function is essential for plant growth. Knockdown plants display slow growth and show pale-green leaves. Physiological and ultrastructural analyses of atybeY mutants revealed impaired photosynthesis and defective chloroplast development. Fluorescent microcopy analysis showed that, when fused with the green fluorescence protein, AtYbeY is localized in chloroplasts. Immunoblot and RNA gel-blot assays revealed that the levels of chloroplast-encoded subunits of photosynthetic complexes are reduced in atybeY mutants, but the corresponding transcripts accumulate normally. In addition, atybeY mutants display defective maturation of both the 5′ and 3′ ends of 16S, 23S, and 4.5S rRNAs as well as decreased accumulation of mature transcripts from the transfer RNA genes contained in the chloroplast rRNA operon. Consequently, mutant plants show a severe deficiency in ribosome biogenesis, which, in turn, results in impaired plastid translational activity. Furthermore, biochemical assays show that recombinant AtYbeY is able to cleave chloroplast rRNAs as well as messenger RNAs and transfer RNAs in vitro. Taken together, our findings indicate that AtYbeY is a chloroplast-localized endoribonuclease that is required for chloroplast rRNA processing and thus for normal growth and development. PMID:25810095

  12. Time Gating of Chloroplast Autofluorescence Allows Clearer Fluorescence Imaging In Planta

    PubMed Central

    Kodama, Yutaka

    2016-01-01

    Chloroplast, an organelle facilitating photosynthesis, exhibits strong autofluorescence, which is an undesired background signal that restricts imaging experiments with exogenous fluorophore in plants. In this study, the autofluorescence was characterized in planta under confocal laser microscopy, and it was found that the time-gated imaging technique completely eliminates the autofluorescence. As a demonstration of the technique, a clearer signal of fluorescent protein-tagged phototropin, a blue-light photoreceptor localized at the chloroplast periphery, was visualized in planta. PMID:27027881

  13. Time Gating of Chloroplast Autofluorescence Allows Clearer Fluorescence Imaging In Planta.

    PubMed

    Kodama, Yutaka

    2016-01-01

    Chloroplast, an organelle facilitating photosynthesis, exhibits strong autofluorescence, which is an undesired background signal that restricts imaging experiments with exogenous fluorophore in plants. In this study, the autofluorescence was characterized in planta under confocal laser microscopy, and it was found that the time-gated imaging technique completely eliminates the autofluorescence. As a demonstration of the technique, a clearer signal of fluorescent protein-tagged phototropin, a blue-light photoreceptor localized at the chloroplast periphery, was visualized in planta. PMID:27027881

  14. Localization of the Enzymes of Quinolizidine Alkaloid Biosynthesis in Leaf Chloroplasts of Lupinus polyphyllus1

    PubMed Central

    Wink, Michael; Hartmann, Thomas

    1982-01-01

    Studies with purified chloroplasts of Lupinus polyphyllus LINDL. leaflets indicate that the first two enzymes of quinolizidine alkaloid biosynthesis, lysine decarboxylase and 17-oxosparteine synthase, are localized in the chloroplast stroma. Thus, both enzymes share the same subcellular compartment as the biosynthetic pathway of lysine, the precursor of quinolizidine alkaloids. The activity of diaminopimelate decarboxylase, the final enzyme in lysine biosynthesis, is about two to three orders of magnitude higher than that of the enzymes of alkaloid formation. PMID:16662483

  15. Diversity in Biosynthetic Pathways of Galactolipids in the Light of Endosymbiotic Origin of Chloroplasts

    PubMed Central

    Sato, Naoki; Awai, Koichiro

    2016-01-01

    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. PMID:26904079

  16. A Zinc Finger Motif-Containing Protein Is Essential for Chloroplast RNA Editing

    PubMed Central

    Sun, Tao; Shi, Xiaowen; Friso, Giulia; Van Wijk, Klaas; Bentolila, Stephane; Hanson, Maureen R.

    2015-01-01

    C-to-U editing of transcripts in plant organelles is carried out by small (<400 kD) protein complexes called editosomes. Recognition of the proper C target for editing is mediated by pentatricopeptide repeat (PPR) containing proteins that recognize cis-elements. Members of two additional gene families, the RIP/MORF and ORRM families, have each been found to be required for editing of particular sets of Cs in mitochondria and/or chloroplasts. By co-immunoprecipitation of the chloroplast editing factor ORRM1, followed by mass spectrometry, we have now identified a member of the RanBP2 type zinc fingers (pFAM00641) protein family that is required for editing of 14 sites in chloroplasts and affects editing efficiency of another 16 chloroplast C targets. In yeast two-hybrid assays, OZ1 (Organelle Zinc finger 1) interacts with PPR site recognition factors whose cognate sites are affected when OZ1 is mutated. No interaction of OZ1 with the chloroplast editing factors RIP2 and RIP9 was detected; however, OZ1 interacts with ORRM1, which binds to RIP proteins, allowing us to build a model for the chloroplast RNA editosome. The RNA editosomes that act upon most chloroplast C targets are likely to contain a PPR protein recognition factor, either RIP2 or RIP9, ORRM1, and OZ1. The organelle zinc finger editing factor family (OZ) contains 4 members in Arabidopsis, three that are predicted to be targeted to chloroplasts and one to mitochondria. With the identification of OZ1, there are now 4 nuclear-encoded protein families known to be essential for plant organelle RNA editing. PMID:25768119

  17. Rust fungal effectors mimic host transit peptides to translocate into chloroplasts.

    PubMed

    Petre, Benjamin; Lorrain, Cécile; Saunders, Diane G O; Win, Joe; Sklenar, Jan; Duplessis, Sébastien; Kamoun, Sophien

    2016-04-01

    Parasite effector proteins target various host cell compartments to alter host processes and promote infection. How effectors cross membrane-rich interfaces to reach these compartments is a major question in effector biology. Growing evidence suggests that effectors use molecular mimicry to subvert host cell machinery for protein sorting. We recently identified chloroplast-targeted protein 1 (CTP1), a candidate effector from the poplar leaf rust fungus Melampsora larici-populina that carries a predicted transit peptide and accumulates in chloroplasts and mitochondria. Here, we show that the CTP1 transit peptide is necessary and sufficient for accumulation in the stroma of chloroplasts. CTP1 is part of a Melampsora-specific family of polymorphic secreted proteins. Two members of that family, CTP2 and CTP3, also translocate in chloroplasts in an N-terminal signal-dependent manner. CTP1, CTP2 and CTP3 are cleaved when they accumulate in chloroplasts, while they remain intact when they do not translocate into chloroplasts. Our findings reveal that fungi have evolved effector proteins that mimic plant-specific sorting signals to traffic within plant cells. PMID:26426202

  18. Tic40 is important for reinsertion of proteins from the chloroplast stroma into the inner membrane

    PubMed Central

    Chiu, Chi-Chou; Li, Hsou-min

    2008-01-01

    Chloroplast inner-membrane proteins Tic40 and Tic110 are first imported from the cytosol into the chloroplast stroma, and subsequently reinserted from the stroma into the inner membrane. However, the mechanism of reinsertion remains unclear. Here we show that Tic40 itself is involved in this reinsertion process. When precursors of either Tic40 or a Tic110 C-terminal truncate, tpTic110-Tic110N, were imported into chloroplasts isolated from a tic40-null mutant, soluble Tic40 and Tic110N intermediates accumulated in the stroma of tic40-mutant chloroplasts, due to a slower rate of reinsertion. We further show that a larger quantity of soluble Tic21 intermediates also accumulated in the stroma of tic40-mutant chloroplasts. In contrast, inner-membrane insertion of the triose-phosphate/phosphate translocator was not affected by the tic40 mutation. Our data suggest that multiple pathways exist for the insertion of chloroplast inner-membrane proteins. PMID:18657235

  19. Incongruence between Nuclear and Chloroplast DNA Phylogenies in Pedicularis Section Cyathophora (Orobanchaceae)

    PubMed Central

    Yu, Wen-Bin; Huang, Pan-Hui; Li, De-Zhu; Wang, Hong

    2013-01-01

    Pedicularis section Cyathophora is a monophyletic group characterized by perfoliate leaf and/or bract bases at each node. This section comprises four series, corresponding to four general corolla types of Pedicularis, i.e. toothless, toothed, beaked and long-tubed corollas. In this study, we aim to reconstruct a comprehensive phylogeny of section Cyathophora, and compare phylogenetic incongruence between nuclear and chloroplast datasets. Sixty-seven accessions belonging to section Cyathophora and 9 species for other Pedicularis were sampled, and one nuclear gene (nrITS) and four chloroplast genes (matK, rbcL, trnH-psbA and trnL-F) were sequenced. Phylogenetic analyses show that the topologies and networks inferred from nrITS and the concatenated chloroplast datasets were incongruent, and the nrITS phylogenies and network agreed with the morphology-based taxonomy to some degree. The chloroplast genome of two Sichuan samples of P. cyathophylloides (E4 and E5) may show introgression from an ancestor of P. cyathophylla. Neither the nrITS dataset nor the concatenated chloroplast dataset were able to adequately resolve relationships among species in the series Reges; this is most likely due to incomplete lineage sorting and/or introgression/hybridization. The nrITS phylogeny indicates the beakless (toothed and toothless) and beaked galeas may have evolved independently within section Cyathophora, and the chloroplast phylogeny reveals that the long corolla tube with beaked galea is derived from the short one. PMID:24069353

  20. Chloroplast Activity and 3'phosphadenosine 5'phosphate Signaling Regulate Programmed Cell Death in Arabidopsis.

    PubMed

    Bruggeman, Quentin; Mazubert, Christelle; Prunier, Florence; Lugan, Raphaël; Chan, Kai Xun; Phua, Su Yin; Pogson, Barry James; Krieger-Liszkay, Anja; Delarue, Marianne; Benhamed, Moussa; Bergounioux, Catherine; Raynaud, Cécile

    2016-03-01

    Programmed cell death (PCD) is a crucial process both for plant development and responses to biotic and abiotic stress. There is accumulating evidence that chloroplasts may play a central role during plant PCD as for mitochondria in animal cells, but it is still unclear whether they participate in PCD onset, execution, or both. To tackle this question, we have analyzed the contribution of chloroplast function to the cell death phenotype of the myoinositol phosphate synthase1 (mips1) mutant that forms spontaneous lesions in a light-dependent manner. We show that photosynthetically active chloroplasts are required for PCD to occur in mips1, but this process is independent of the redox state of the chloroplast. Systematic genetic analyses with retrograde signaling mutants reveal that 3'-phosphoadenosine 5'-phosphate, a chloroplast retrograde signal that modulates nuclear gene expression in response to stress, can inhibit cell death and compromises plant innate immunity via inhibition of the RNA-processing 5'-3' exoribonucleases. Our results provide evidence for the role of chloroplast-derived signal and RNA metabolism in the control of cell death and biotic stress response. PMID:26747283

  1. The cleavable pre-sequence of an imported chloroplast protein directs attached polypeptides into yeast mitochondria

    PubMed Central

    Hurt, Eduard C.; Soltanifar, Nouchine; Goldschmidt-Clermont, Michel; Rochaix, Jean-David; Schatz, Gottfried

    1986-01-01

    The cleavable pre-sequences of imported chloroplast and mitochondrial proteins have several features in common. This structural similarity prompted us to test whether a chloroplast pre-sequence (`transit peptide') can also be decoded by the mitochondrial import machinery. In the green alga, Chlamydomonas reinhardtii, the small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) (a chloroplast protein) is nuclear-encoded and synthesized in the cytosol with a transient pre-sequence of 45 residues. The 31 amino-terminal residues of this chloroplast pre-sequence were fused to mouse dihydrofolate reductase (a cytosolic protein) and to yeast cytochrome oxidase subunit IV (an imported mitochondrial protein) from which the authentic pre-sequence had been removed. The chloroplast pre-sequence transported both attached proteins into the yeast mitochondrial matrix or inner membrane, although it functioned less efficiently than an authentic mitochondrial pre-sequence. We conclude that mitochondrial and chloroplast pre-sequences perform their function by a similar mechanism. ImagesFig. 3.Fig. 4.Fig. 5.Fig. 6. PMID:16453686

  2. Rapid Mass Movement of Chloroplasts during Segment Formation of the Calcifying Siphonalean Green Alga, Halimeda macroloba

    PubMed Central

    Larkum, Anthony W. D.; Salih, Anya; Kühl, Michael

    2011-01-01

    Background The calcifying siphonalean green alga, Halimeda macroloba is abundant on coral reefs and is important in the production of calcium carbonate sediments. The process by which new green segments are formed over-night is revealed here for the first time. Methodology/Principal Findings Growth of new segments was visualised by epifluorescence and confocal microscopy and by pulse amplitude modulation (PAM) fluorimetry. Apical colourless proto-segments were initiated on day 1, and formed a loose network of non-calcified, non-septate filaments, containing no chloroplasts. Rapid greening was initiated at dusk by i) the mass movement of chloroplasts into these filaments from the parent segment and ii) the growth of new filaments containing chloroplasts. Greening was usually complete in 3–5 h and certainly before dawn on day 2 when the first signs of calcification were apparent. Mass chloroplast movement took place at a rate of ∼0.65 µm/s. Photosynthetic yield and rate remained low for a period of 1 to several hours, indicating that the chloroplasts were made de novo. Use of the inhibitors colchicine and cytochalasin d indicated that the movement process is dependent on both microtubules and microfilaments. Significance This unusual process involves the mass movement of chloroplasts at a high rate into new segments during the night and rapid calcification on the following day and may be an adaptation to minimise the impact of herbivorous activity. PMID:21750703

  3. Ribonuclease J is required for chloroplast and embryo development in Arabidopsis

    PubMed Central

    Chen, Hongyu; Zou, Wenxuan; Zhao, Jie

    2015-01-01

    Chloroplasts perform many essential metabolic functions and their proper development is critically important in embryogenesis. However, little is known about how chloroplasts function in embryogenesis and more relevant components need to be characterized. In this study, we show that Arabidopsis Ribonuclease J (RNase J) is required for chloroplast and embryo development. Mutation of AtRNJ led to albino ovules containing aborted embryos; the morphological development of rnj embryos was disturbed after the globular stage. Observation of ultrastructures indicated that these aborted embryos may result from impaired chloroplast development. Furthermore, by analyzing the molecular markers of cell fate decisions (STM, FIL, ML1, SCR, and WOX5) in rnj embryos, we found that this impairment of chloroplast development may lead to aberrant embryo patterning along the apical-basal axis, indicating that AtRNJ is important in initiating and maintaining the organization of shoot apical meristems (SAMs), cotyledons, and hypocotyls. Moreover, the transport and response of auxin in rnj embryos was found to be disrupted, suggesting that AtRNJ may be involved in auxin-mediated pathways during embryogenesis. Therefore, we speculate that RNJ plays a vital role in embryo morphogenesis and apical-basal pattern formation by regulating chloroplast development. PMID:25871650

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

    PubMed

    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. PMID:23001039

  5. Gustav Senn (1875-1945): the pioneer of chloroplast movement research.

    PubMed

    Kataoka, Hironao

    2015-01-01

    Gustav Senn analyzed for the first time light-induced movement and arrangement of chloroplasts. Using many plant species he performed physiological analyses of chloroplast migration in response to external stimuli, with emphasis on light. He determined light paths within a cell by measuring refractive indices and optical thickness of cellular compartments and confirmed that chloroplasts migrate towards the region where the light intensity is optimum. After 6 to 7 years' concentrated study, Senn published the famous monograph "Die Gestalts- und Lageveränderung der Pflanzen- Chromatophoren" (The Changes in Shape and Position of Plant Chloroplasts) in 1908. This book has stimulated many plant physiologists and photobiologists, because Senn not only thoroughly classified and defined various types of light-induced chloroplast migration but also already described possible interaction of different photoreceptor systems in Mougeotia more than 50 years before the discovery of phytochrome. This book also contains still useful experimental hints and overlooked findings on the interaction between light and other factors, such as temperature, water content, and nourishment. After publishing this book, Senn retreated from the study of chloroplasts and became a researcher of the Greek philosopher, Theophrastus. In this review, I introduce his biographical background and then summarize some of his key research accomplishment. PMID:25406543

  6. Inferring Phylogenetic Relationships of Indian Citron (Citrus medica L.) based on rbcL and matK Sequences of Chloroplast DNA.

    PubMed

    Uchoi, Ajit; Malik, Surendra Kumar; Choudhary, Ravish; Kumar, Susheel; Rohini, M R; Pal, Digvender; Ercisli, Sezai; Chaudhury, Rekha

    2016-06-01

    Phylogenetic relationships of Indian Citron (Citrus medica L.) with other important Citrus species have been inferred through sequence analyses of rbcL and matK gene region of chloroplast DNA. The study was based on 23 accessions of Citrus genotypes representing 15 taxa of Indian Citrus, collected from wild, semi-wild, and domesticated stocks. The phylogeny was inferred using the maximum parsimony (MP) and neighbor-joining (NJ) methods. Both MP and NJ trees separated all the 23 accessions of Citrus into five distinct clusters. The chloroplast DNA (cpDNA) analysis based on rbcL and matK sequence data carried out in Indian taxa of Citrus was useful in differentiating all the true species and species/varieties of probable hybrid origin in distinct clusters or groups. Sequence analysis based on rbcL and matK gene provided unambiguous identification and disposition of true species like C. maxima, C. medica, C. reticulata, and related hybrids/cultivars. The separation of C. maxima, C. medica, and C. reticulata in distinct clusters or sub-clusters supports their distinctiveness as the basic species of edible Citrus. However, the cpDNA sequence analysis of rbcL and matK gene could not find any clear cut differentiation between subgenera Citrus and Papeda as proposed in Swingle's system of classification. PMID:26956119

  7. Protein-Induced Modulation of Chloroplast Membrane Morphology

    PubMed Central

    Machettira, Anu B.; Groß, Lucia E.; Tillmann, Bodo; Weis, Benjamin L.; Englich, Gisela; Sommer, Maik S.; Königer, Martina; Schleiff, Enrico

    2012-01-01

    Organelles are surrounded by membranes with a distinct lipid and protein composition. While it is well established that lipids affect protein functioning and vice versa, it has been only recently suggested that elevated membrane protein concentrations may affect the shape and organization of membranes. We therefore analyzed the effects of high chloroplast envelope protein concentrations on membrane structures using an in vivo approach with protoplasts. Transient expression of outer envelope proteins or protein domains such as CHUP1-TM–GFP, outer envelope protein of 7 kDa–GFP, or outer envelope protein of 24 kDa–GFP at high levels led to the formation of punctate, circular, and tubular membrane protrusions. Expression of inner membrane proteins such as translocase of inner chloroplast membrane 20, isoform II (Tic20-II)–GFP led to membrane protrusions including invaginations. Using increasing amounts of DNA for transfection, we could show that the frequency, size, and intensity of these protrusions increased with protein concentration. The membrane deformations were absent after cycloheximide treatment. Co-expression of CHUP1-TM–Cherry and Tic20-II–GFP led to membrane protrusions of various shapes and sizes including some stromule-like structures, for which several functions have been proposed. Interestingly, some structures seemed to contain both proteins, while others seem to contain one protein exclusively, indicating that outer and inner envelope dynamics might be regulated independently. While it was more difficult to investigate the effects of high expression levels of membrane proteins on mitochondrial membrane shapes using confocal imaging, it was striking that the expression of the outer membrane protein Tom20 led to more elongate mitochondria. We discuss that the effect of protein concentrations on membrane structure is possibly caused by an imbalance in the lipid to protein ratio and may be involved in a signaling pathway regulating membrane

  8. Habitat Loss other than Fragmentation per se Decreased Nuclear and Chloroplast Genetic Diversity in a Monoecious Tree

    PubMed Central

    Shen, Dong-Wei; Chen, Xiao-Yong

    2012-01-01

    Generally, effect of fragmentation per se on biodiversity has not been separated from the effect of habitat loss. In this paper, using nDNA and cpDNA SSRs, we studied genetic diversity of Castanopsis sclerophylla (Lindl. & Paxton) Schotty populations and decoupled the effects of habitat loss and fragmentation per se. We selected seven nuclear and six cpDNA microsatellite loci and genotyped 460 individuals from mainland and island populations, which were located in the impoundment created in 1959. Number of alleles per locus of populations in larger habitats was significantly higher than that in smaller habitats. There was a significant relationship between the number of alleles per locus and habitat size. Based on this relationship, the predicted genetic diversity of an imaginary population of size equaling the total area of the islands was lower than that of the global population on the islands. Re-sampling demonstrated that low genetic diversity of populations in small habitats was caused by unevenness in sample size. Fisher's α index was similar among habitat types. These results indicate that the decreased nuclear and chloroplast genetic diversity of populations in smaller habitats was mainly caused by habitat loss. For nuclear and chloroplast microsatellite loci, values of FST were 0.066 and 0.893, respectively, and the calculated pollen/seed dispersal ratio was 162.2. When separated into pre-and post-fragmentation cohorts, pollen/seed ratios were 121.2 and 189.5, respectively. Our results suggest that habitat loss explains the early decrease in genetic diversity, while fragmentation per se may play a major role in inbreeding and differentiation among fragmented populations and later loss of genetic diversity. PMID:22723951

  9. Habitat loss other than fragmentation per se decreased nuclear and chloroplast genetic diversity in a monoecious tree.

    PubMed

    Zhang, Xin; Shi, Miao-Miao; Shen, Dong-Wei; Chen, Xiao-Yong

    2012-01-01

    Generally, effect of fragmentation per se on biodiversity has not been separated from the effect of habitat loss. In this paper, using nDNA and cpDNA SSRs, we studied genetic diversity of Castanopsis sclerophylla (Lindl. & Paxton) Schotty populations and decoupled the effects of habitat loss and fragmentation per se. We selected seven nuclear and six cpDNA microsatellite loci and genotyped 460 individuals from mainland and island populations, which were located in the impoundment created in 1959. Number of alleles per locus of populations in larger habitats was significantly higher than that in smaller habitats. There was a significant relationship between the number of alleles per locus and habitat size. Based on this relationship, the predicted genetic diversity of an imaginary population of size equaling the total area of the islands was lower than that of the global population on the islands. Re-sampling demonstrated that low genetic diversity of populations in small habitats was caused by unevenness in sample size. Fisher's α index was similar among habitat types. These results indicate that the decreased nuclear and chloroplast genetic diversity of populations in smaller habitats was mainly caused by habitat loss. For nuclear and chloroplast microsatellite loci, values of F(ST) were 0.066 and 0.893, respectively, and the calculated pollen/seed dispersal ratio was 162.2. When separated into pre-and post-fragmentation cohorts, pollen/seed ratios were 121.2 and 189.5, respectively. Our results suggest that habitat loss explains the early decrease in genetic diversity, while fragmentation per se may play a major role in inbreeding and differentiation among fragmented populations and later loss of genetic diversity. PMID:22723951

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

    PubMed

    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. PMID:25096976

  11. The novel protein DELAYED PALE-GREENING1 is required for early chloroplast biogenesis in Arabidopsis thaliana.

    PubMed

    Liu, Dong; Li, Weichun; Cheng, Jianfeng

    2016-01-01

    Chloroplast biogenesis is one of the most important subjects in plant biology. In this study, an Arabidopsis early chloroplast biogenesis mutant with a delayed pale-greening phenotype (dpg1) was isolated from a T-DNA insertion mutant collection. Both cotyledons and true leaves of dpg1 mutants were initially albino but gradually became pale green as the plant matured. Transmission electron microscopic observations revealed that the mutant displayed a delayed proplastid-to-chloroplast transition. Sequence and transcription analyses showed that AtDPG1 encodes a putatively chloroplast-localized protein containing three predicted transmembrane helices and that its expression depends on both light and developmental status. GUS staining for AtDPG1::GUS transgenic lines showed that this gene was widely expressed throughout the plant and that higher expression levels were predominantly found in green tissues during the early stages of Arabidopsis seedling development. Furthermore, quantitative real-time RT-PCR analyses revealed that a number of chloroplast- and nuclear-encoded genes involved in chlorophyll biosynthesis, photosynthesis and chloroplast development were substantially down-regulated in the dpg1 mutant. These data indicate that AtDPG1 plays an essential role in early chloroplast biogenesis, and its absence triggers chloroplast-to-nucleus retrograde signalling, which ultimately down-regulates the expression of nuclear genes encoding chloroplast-localized proteins. PMID:27160321

  12. Genetic interactions reveal that specific defects of chloroplast translation are associated with the suppression of var2-mediated leaf variegation.

    PubMed

    Liu, Xiayan; Zheng, Mengdi; Wang, Rui; Wang, Ruijuan; An, Lijun; Rodermel, Steve R; Yu, Fei

    2013-10-01

    Arabidopsis thaliana L. yellow variegated (var2) mutant is defective in a chloroplast FtsH family metalloprotease, AtFtsH2/VAR2, and displays an intriguing green and white leaf variegation. This unique var2-mediated leaf variegation offers a simple yet powerful tool for dissecting the genetic regulation of chloroplast development. Here, we report the isolation and characterization of a new var2 suppressor gene, SUPPRESSOR OF VARIEGATION8 (SVR8), which encodes a putative chloroplast ribosomal large subunit protein, L24. Mutations in SVR8 suppress var2 leaf variegation at ambient temperature and partially suppress the cold-induced chlorosis phenotype of var2. Loss of SVR8 causes unique chloroplast rRNA processing defects, particularly the 23S-4.5S dicistronic precursor. The recovery of the major abnormal processing site in svr8 23S-4.5S precursor indicate that it does not lie in the same position where SVR8/L24 binds on the ribosome. Surprisingly, we found that the loss of a chloroplast ribosomal small subunit protein, S21, results in aberrant chloroplast rRNA processing but not suppression of var2 variegation. These findings suggest that the disruption of specific aspects of chloroplast translation, rather than a general impairment in chloroplast translation, suppress var2 variegation and the existence of complex genetic interactions in chloroplast development. PMID:23721655

  13. The novel protein DELAYED PALE-GREENING1 is required for early chloroplast biogenesis in Arabidopsis thaliana

    PubMed Central

    Liu, Dong; Li, Weichun; Cheng, Jianfeng

    2016-01-01

    Chloroplast biogenesis is one of the most important subjects in plant biology. In this study, an Arabidopsis early chloroplast biogenesis mutant with a delayed pale-greening phenotype (dpg1) was isolated from a T-DNA insertion mutant collection. Both cotyledons and true leaves of dpg1 mutants were initially albino but gradually became pale green as the plant matured. Transmission electron microscopic observations revealed that the mutant displayed a delayed proplastid-to-chloroplast transition. Sequence and transcription analyses showed that AtDPG1 encodes a putatively chloroplast-localized protein containing three predicted transmembrane helices and that its expression depends on both light and developmental status. GUS staining for AtDPG1::GUS transgenic lines showed that this gene was widely expressed throughout the plant and that higher expression levels were predominantly found in green tissues during the early stages of Arabidopsis seedling development. Furthermore, quantitative real-time RT-PCR analyses revealed that a number of chloroplast- and nuclear-encoded genes involved in chlorophyll biosynthesis, photosynthesis and chloroplast development were substantially down-regulated in the dpg1 mutant. These data indicate that AtDPG1 plays an essential role in early chloroplast biogenesis, and its absence triggers chloroplast-to-nucleus retrograde signalling, which ultimately down-regulates the expression of nuclear genes encoding chloroplast-localized proteins. PMID:27160321

  14. An Ancient Bacterial Signaling Pathway Regulates Chloroplast Function to Influence Growth and Development in Arabidopsis[OPEN

    PubMed Central

    Sugliani, Matteo; Ke, Hang; Bouveret, Emmanuelle; Robaglia, Christophe; Caffarri, Stefano

    2016-01-01

    The chloroplast originated from the endosymbiosis of an ancient photosynthetic bacterium by a eukaryotic cell. Remarkably, the chloroplast has retained elements of a bacterial stress response pathway that is mediated by the signaling nucleotides guanosine penta- and tetraphosphate (ppGpp). However, an understanding of the mechanism and outcomes of ppGpp signaling in the photosynthetic eukaryotes has remained elusive. Using the model plant Arabidopsis thaliana, we show that ppGpp is a potent regulator of chloroplast gene expression in vivo that directly reduces the quantity of chloroplast transcripts and chloroplast-encoded proteins. We then go on to demonstrate that the antagonistic functions of different plant RelA SpoT homologs together modulate ppGpp levels to regulate chloroplast function and show that they are required for optimal plant growth, chloroplast volume, and chloroplast breakdown during dark-induced and developmental senescence. Therefore, our results show that ppGpp signaling is not only linked to stress responses in plants but is also an important mediator of cooperation between the chloroplast and the nucleocytoplasmic compartment during plant growth and development. PMID:26908759

  15. An Essential Component in Chloroplast Development and Maintenance at Moderate High Temperature in Higher Plants: Chloroplast-targeted FtsH11 Proteases

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Among the 12 predicted FtsH proteases in Arabidopsis, AtFtsH11 is the only metalloprotease targeting to both chloroplast and mitochondria and the only one essential for Arabidopsis plant to survive at moderate heat stress at all developmental stages. Under optimal conditions, atftsh11 mutants were...

  16. BIOGENESIS FACTOR REQUIRED FOR ATP SYNTHASE 3 Facilitates Assembly of the Chloroplast ATP Synthase Complex.

    PubMed

    Zhang, Lin; Duan, Zhikun; Zhang, Jiao; Peng, Lianwei

    2016-06-01

    Thylakoid membrane-localized chloroplast ATP synthases use the proton motive force generated by photosynthetic electron transport to produce ATP from ADP. Although it is well known that the chloroplast ATP synthase is composed of more than 20 proteins with α3β3γ1ε1δ1I1II1III14IV1 stoichiometry, its biogenesis process is currently unclear. To unravel the molecular mechanisms underlying the biogenesis of chloroplast ATP synthase, we performed extensive screening for isolating ATP synthase mutants in Arabidopsis (Arabidopsis thaliana). In the recently identified bfa3 (biogenesis factors required for ATP synthase 3) mutant, the levels of chloroplast ATP synthase subunits were reduced to approximately 25% of wild-type levels. In vivo labeling analysis showed that assembly of the CF1 component of chloroplast ATP synthase was less efficient in bfa3 than in the wild type, indicating that BFA3 is required for CF1 assembly. BFA3 encodes a chloroplast stromal protein that is conserved in higher plants, green algae, and a few species of other eukaryotic algae, and specifically interacts with the CF1β subunit. The BFA3 binding site was mapped to a region in the catalytic site of CF1β. Several residues highly conserved in eukaryotic CF1β are crucial for the BFA3-CF1β interaction, suggesting a coevolutionary relationship between BFA3 and CF1β. BFA3 appears to function as a molecular chaperone that transiently associates with unassembled CF1β at its catalytic site and facilitates subsequent association with CF1α during assembly of the CF1 subcomplex of chloroplast ATP synthase. PMID:27208269

  17. Complete Chloroplast Genome Sequences of Mongolia Medicine Artemisia frigida and Phylogenetic Relationships with Other Plants

    PubMed Central

    Liu, Yue; Huo, Naxin; Dong, Lingli; Wang, Yi; Zhang, Shuixian; Young, Hugh A.; Feng, Xiaoxiao; Gu, Yong Qiang

    2013-01-01

    Background Artemisia frigida Willd. is an important Mongolian traditional medicinal plant with pharmacological functions of stanch and detumescence. However, there is little sequence and genomic information available for Artemisia frigida, which makes phylogenetic identification, evolutionary studies, and genetic improvement of its value very difficult. We report the complete chloroplast genome sequence of Artemisia frigida based on 454 pyrosequencing. Methodology/Principal Findings The complete chloroplast genome of Artemisia frigida is 151,076 bp including a large single copy (LSC) region of 82,740 bp, a small single copy (SSC) region of 18,394 bp and a pair of inverted repeats (IRs) of 24,971 bp. The genome contains 114 unique genes and 18 duplicated genes. The chloroplast genome of Artemisia frigida contains a small 3.4 kb inversion within a large 23 kb inversion in the LSC region, a unique feature in Asteraceae. The gene order in the SSC region of Artemisia frigida is inverted compared with the other 6 Asteraceae species with the chloroplast genomes sequenced. This inversion is likely caused by an intramolecular recombination event only occurred in Artemisia frigida. The existence of rich SSR loci in the Artemisia frigida chloroplast genome provides a rare opportunity to study population genetics of this Mongolian medicinal plant. Phylogenetic analysis demonstrates a sister relationship between Artemisia frigida and four other species in Asteraceae, including Ageratina adenophora, Helianthus annuus, Guizotia abyssinica and Lactuca sativa, based on 61 protein-coding sequences. Furthermore, Artemisia frigida was placed in the tribe Anthemideae in the subfamily Asteroideae (Asteraceae) based on ndhF and trnL-F sequence comparisons. Conclusion The chloroplast genome sequence of Artemisia frigida was assembled and analyzed in this study, representing the first plastid genome sequenced in the Anthemideae tribe. This complete chloroplast genome sequence will be

  18. Structural characterizations of the chloroplast translocon protein Tic110

    PubMed Central

    Tsai, Jia-Yin; Chu, Chiung-Chih; Yeh, Yi-Hung; Chen, Lih-Jen; Li, Hsou-min; Hsiao, Chwan-Deng

    2013-01-01

    Tic110 is a major component of the chloroplast protein import translocon. Two functions with mutually exclusive structures have been proposed for Tic110: a protein-conducting channel with six transmembrane domains and a scaffold with two N-terminal transmembrane domains followed by a large soluble domain for binding transit peptides and other stromal translocon components. To investigate the structure of Tic110, Tic110 from Cyanidioschyzon merolae (CmTic110) was characterized. We constructed three fragments, CmTic110A, CmTic110B and CmTic110C, with increasing N-terminal truncations, to perform small-angle X-ray scattering (SAXS) and X-ray crystallography analyses and Dali structural comparison. Here we report the molecular envelope of CmTic110B and CmTic110C determined by SAXS, and the crystal structure of CmTic110C at 4.2 Å. Our data indicate that the C-terminal half of CmTic110 possesses a rod-shaped helix-repeat structure that is too flattened and elongated to be a channel. The structure is most similar to the HEAT-repeat motif that functions as scaffolds for protein–protein interactions. PMID:23711301

  19. Wood identification with PCR targeting noncoding chloroplast DNA.

    PubMed

    Tang, Xiaoshu; Zhao, Guangjie; Ping, Liyan

    2011-12-01

    Wood identification is extremely important in the modern forest industry. It also has significant applications in forensics, as well as in archeology and ecological research. In this study, five universal primer pairs amplifying chloroplast noncoding sequences of 300-1,200 bp were designed. Sequencing these amplicons in combination can lead to reliable identification of logs and wood products to cultivar, ecotype, or even the falling population. These primer pairs work on both gymnosperms and angiosperm trees. They also are potentially applicable to accurately identify shrubs and herbaceous species. In addition, a wood DNA purification method is proposed in which N-phenacylthiazolium bromide (PTB) is used to increase the quality and quantity of extracted DNA. This method was first validated using air-dried timber disks from three different tree species that were felled 4 years ago. The sapwood and outer heartwood provided the best locations for DNA extraction. The method was also successfully applied to extract DNA from the recalcitrant processed white oak wood, randomly selected staves of wine barrels. The single nucleotide polymorphism detected on the oak DNA sequences showed correlation to their geographical origins. PMID:22038094

  20. Identification of the Elusive Pyruvate Reductase of Chlamydomonas reinhardtii Chloroplasts.

    PubMed

    Burgess, Steven J; Taha, Hussein; Yeoman, Justin A; Iamshanova, Oksana; Chan, Kher Xing; Boehm, Marko; Behrends, Volker; Bundy, Jacob G; Bialek, Wojciech; Murray, James W; Nixon, Peter J

    2016-01-01

    Under anoxic conditions the green alga Chlamydomonas reinhardtii activates various fermentation pathways leading to the creation of formate, acetate, ethanol and small amounts of other metabolites including d-lactate and hydrogen. Progress has been made in identifying the enzymes involved in these pathways and their subcellular locations; however, the identity of the enzyme involved in reducing pyruvate to d-lactate has remained unclear. Based on sequence comparisons, enzyme activity measurements, X-ray crystallography, biochemical fractionation and analysis of knock-down mutants, we conclude that pyruvate reduction in the chloroplast is catalyzed by a tetrameric NAD(+)-dependent d-lactate dehydrogenase encoded by Cre07.g324550. Its expression during aerobic growth supports a possible function as a 'lactate valve' for the export of lactate to the mitochondrion for oxidation by cytochrome-dependent d-lactate dehydrogenases and by glycolate dehydrogenase. We also present a revised spatial model of fermentation based on our immunochemical detection of the likely pyruvate decarboxylase, PDC3, in the cytoplasm. PMID:26574578

  1. Chloroplast Genetics of Chlamydomonas. III. Closing the Circle

    PubMed Central

    Singer, Burt; Sager, Ruth; Ramanis, Zenta

    1976-01-01

    A novel mapping procedure is presented for organelle genes or any other genetic system exhibiting a measurable frequency of exchanges occurring at a constant rate over a measurable time interval. For a set of markers in a multiply-marked cross, the exchange rates measure relative map distances from a centromere-like attachment point. With this method, we present mapping data and a linear map of genes in the chlcroplast genome of Chlamydomonas. The data are plotted as log (percent remaining heterozygotes) against time and map distances are taken as proportional to slope. A statistical method which is an adaptation of jackknife methodology to a regression problem was developed to estimate slope values. A single line is fitted to pooled data for each marker from several crosses, and then lines are re-fit to a series of pooled data sets in each of which the observations from a single cross have been omitted. From these data sets a final summary slope is computed as well as a statement of its variability. The relative positions of new markers present in single crosses can then be estimated utilizing data from many crosses. The method does not distinguish between one-armed and two-armed linear or circular maps. However, evaluation of this map in conjunction with cosegregation frequency data (Sager and Ramanis 1976b) provides unambiguous evidence of the genetic circularity of the Chlamydomonas chloroplast genome. PMID:17248718

  2. A Proteomic Analysis of Maize Chloroplast Biogenesis1

    PubMed Central

    Lonosky, Patricia M.; Zhang, Xiaosi; Honavar, Vasant G.; Dobbs, Drena L.; Fu, Aigen; Rodermel, Steve R.

    2004-01-01

    Proteomics studies to explore global patterns of protein expression in plant and green algal systems have proliferated within the past few years. Although most of these studies have involved mapping of the proteomes of various organs, tissues, cells, or organelles, comparative proteomics experiments have also led to the identification of proteins that change in abundance in various developmental or physiological contexts. Despite the growing use of proteomics in plant studies, questions of reproducibility have not generally been addressed, nor have quantitative methods been widely used, for example, to identify protein expression classes. In this report, we use the de-etiolation (“greening”) of maize (Zea mays) chloroplasts as a model system to explore these questions, and we outline a reproducible protocol to identify changes in the plastid proteome that occur during the greening process using techniques of two-dimensional gel electrophoresis and mass spectrometry. We also evaluate hierarchical and nonhierarchical statistical methods to analyze the patterns of expression of 526 “high-quality,” unique spots on the two-dimensional gels. We conclude that Adaptive Resonance Theory 2—a nonhierarchical, neural clustering technique that has not been previously applied to gene expression data—is a powerful technique for discriminating protein expression classes during greening. Our experiments provide a foundation for the use of proteomics in the design of experiments to address fundamental questions in plant physiology and molecular biology. PMID:14966246

  3. Two complete chloroplast genome sequences of Cannabis sativa varieties.

    PubMed

    Oh, Hyehyun; Seo, Boyoung; Lee, Seunghwan; Ahn, Dong-Ha; Jo, Euna; Park, Jin-Kyoung; Min, Gi-Sik

    2016-07-01

    In this study, we determined the complete chloroplast (cp) genomes from two varieties of Cannabis sativa. The genome sizes were 153,848 bp (the Korean non-drug variety, Cheungsam) and 153,854 bp (the African variety, Yoruba Nigeria). The genome structures were identical with 131 individual genes [86 protein-coding genes (PCGs), eight rRNA, and 37 tRNA genes]. Further, except for the presence of an intron in the rps3 genes of two C. sativa varieties, the cp genomes of C. sativa had conservative features similar to that of all known species in the order Rosales. To verify the position of C. sativa within the order Rosales, we conducted phylogenetic analysis by using concatenated sequences of all PCGs from 17 complete cp genomes. The resulting tree strongly supported monophyly of Rosales. Further, the family Cannabaceae, represented by C. sativa, showed close relationship with the family Moraceae. The phylogenetic relationship outlined in our study is well congruent with those previously shown for the order Rosales. PMID:26104156

  4. Mechanism of proton permeation through chloroplast lipid membranes.

    PubMed Central

    Fuks, B; Homblé, F

    1996-01-01

    Electrical measurements were carried out to investigate the contribution of chloroplast lipids to the passive proton permeability of both the thylakoid and inner-envelope membranes. Permeability coefficient and conductance to protons were measured for solvent-free bilayers made from monogalactosyldiglyceride:digalactosyldiglycerid: sulfoquinovosyldiglyceride:phosphatidylglycerol (2:1:0.5:0.5, w/w) in the presence of a pH gradient of 7.4/8.1. The permeability coefficient for protons in glycolipids was 5.5 +/- 1.1 x 10(-4) cm s-1 (n = 14). To determine whether this high H+ permeability could be explained by the presence of lipid contaminants such as weak acids, we investigated the effects of (a) bovine serum albumin, which can remove some amphiphilic molecules such as free fatty acids, (b) 6-ketocholestanol, which increases the membrane dipole potential, (c) oleic acid, and (d) chlorodecane, which increases the dielectric constant of the lipid bilayer. Our results show that free fatty acids are inefficient protonophores, as compared with carbonylcyanide-m-chlorphenythydrazone, and that the hypothesis of a weak acid mechanism is not valid with glycolipid bilayers. In the presence of deuterium oxide the H+ conductane was reduced significantly, indicating that proton transport through the glycolipid matrix could occur directly by a hydrogen bond process. The passive transport of H+ through the glycolipid matrix is discussed with regard to the activity of the thylakoid ATP synthase and the inner-envelope H(+)-ATPase. PMID:8883387

  5. Artemisinin Inhibits Chloroplast Electron Transport Activity: Mode of Action

    PubMed Central

    Bharati, Adyasha; Kar, Monaranjan; Sabat, Surendra Chandra

    2012-01-01

    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 QB; 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. PMID:22719995

  6. Plastome Mutations and Recombination Events in Barley Chloroplast Mutator Seedlings.

    PubMed

    Landau, Alejandra; Lencina, Franco; Pacheco, María G; Prina, Alberto R

    2016-05-01

    The barley chloroplast mutator (cpm) is an allele of a nuclear gene that when homozygous induces several types of cytoplasmically inherited chlorophyll deficiencies. In this work, a plastome Targeting Induced Local Lesions in Genomes (TILLING) strategy based on mismatch digestion was used on families that carried the cpm genotype through many generations. Extensive scanning of 33 plastome genes and a few intergenic regions was conducted. Numerous polymorphisms were detected on both genic and intergenic regions. The detected polymorphisms can be accounted for by at least 61 independent mutational events. The vast majority of the polymorphisms originated in substitutions and small indels (insertions/deletions) in microsatellites. The rpl23 and the rps16 genes were the most polymorphic. Interestingly, the variation observed in the rpl23 gene consisted of several combinations of 5 different one nucleotide polymorphisms. Besides, 4 large indels that have direct repeats at both ends were also observed, which appear to be originated from recombinational events. The cpm mutation spectrum suggests that the CPM gene product is probably involved in plastome mismatch repair. The numerous subtle molecular changes that were localized in a wide range of plastome sites show the cpm as a valuable source of plastome variability for plant research and/or plant breeding. Moreover, the cpm mutant appears to be an interesting experimental material for investigating the mechanisms responsible for maintaining the stability of plant organelle DNA. PMID:26774059

  7. Identification of the Elusive Pyruvate Reductase of Chlamydomonas reinhardtii Chloroplasts

    PubMed Central

    Burgess, Steven J.; Taha, Hussein; Yeoman, Justin A.; Iamshanova, Oksana; Chan, Kher Xing; Boehm, Marko; Behrends, Volker; Bundy, Jacob G.; Bialek, Wojciech; Murray, James W.; Nixon, Peter J.

    2016-01-01

    Under anoxic conditions the green alga Chlamydomonas reinhardtii activates various fermentation pathways leading to the creation of formate, acetate, ethanol and small amounts of other metabolites including d-lactate and hydrogen. Progress has been made in identifying the enzymes involved in these pathways and their subcellular locations; however, the identity of the enzyme involved in reducing pyruvate to d-lactate has remained unclear. Based on sequence comparisons, enzyme activity measurements, X-ray crystallography, biochemical fractionation and analysis of knock-down mutants, we conclude that pyruvate reduction in the chloroplast is catalyzed by a tetrameric NAD+-dependent d-lactate dehydrogenase encoded by Cre07.g324550. Its expression during aerobic growth supports a possible function as a ‘lactate valve’ for the export of lactate to the mitochondrion for oxidation by cytochrome-dependent d-lactate dehydrogenases and by glycolate dehydrogenase. We also present a revised spatial model of fermentation based on our immunochemical detection of the likely pyruvate decarboxylase, PDC3, in the cytoplasm. PMID:26574578

  8. Radiation inactivation analysis of chloroplast CF0-CF1 ATPase

    SciTech Connect

    Wang, M.Y.; Chien, L.F.; Pan, R.L.

    1988-06-25

    Radiation inactivation technique was employed to measure the functional size of adenosine triphosphatase of spinach chloroplasts. The functional size for acid-base-induced ATP synthesis was 450 +/- 24 kilodaltons; for phenazine methosulfate-mediated ATP synthesis, 613 +/- 33 kilodaltons; and for methanol-activated ATP hydrolysis, 280 +/- 14 kilodaltons. The difference (170 +/- 57 kilodaltons) between 450 +/- 24 and 280 +/- 14 kilodaltons is explained to be the molecular mass of proton channel (coupling factor 0) across the thylakoid membrane. Our data suggest that the stoichiometry of subunits I, II, and III of coupling factor 0 is 1:2:15. Ca2+- and Mg2+-ATPase activated by methanol, heat, and trypsin digestion have a similar functional size. However, anions such as SO/sub 3/(2-) and CO/sub 3/(2-) increased the molecular mass for both ATPase's (except trypsin-activated Mg2+-ATPase) by 12-30%. Soluble coupling factor 1 has a larger target size than that of membrane-bound. This is interpreted as the cold effect during irradiation.

  9. Plastoglobules: a new address for targeting recombinant proteins in the chloroplast

    PubMed Central

    Vidi, Pierre-Alexandre; Kessler, Felix; Bréhélin, Claire

    2007-01-01

    Background The potential of transgenic plants for cost-effective production of pharmaceutical molecules is now becoming apparent. Plants have the advantage over established fermentation systems (bacterial, yeast or animal cell cultures) to circumvent the risk of pathogen contamination, to be amenable to large scaling up and to necessitate only established farming procedures. Chloroplasts have proven a useful cellular compartment for protein accumulation owing to their large size and number, as well as the possibility for organellar transformation. They therefore represent the targeting destination of choice for recombinant proteins in leaf crops such as tobacco. Extraction and purification of recombinant proteins from leaf material contribute to a large extent to the production costs. Developing new strategies facilitating these processes is therefore necessary. Results Here, we evaluated plastoglobule lipoprotein particles as a new subchloroplastic destination for recombinant proteins. The yellow fluorescent protein as a trackable cargo was targeted to plastoglobules when fused to plastoglobulin 34 (PGL34) as the carrier. Similar to adipocyte differentiation related protein (ADRP) in animal cells, most of the protein sequence of PGL34 was necessary for targeting to lipid bodies. The recombinant protein was efficiently enriched in plastoglobules isolated by simple flotation centrifugation. The viability of plants overproducing the recombinant protein was not affected, indicating that plastoglobule targeting did not significantly impair photosynthesis or sugar metabolism. Conclusion Our data identify plastoglobules as a new targeting destination for recombinant protein in leaf crops. The wide-spread presence of plastoglobules and plastoglobulins in crop species promises applications comparable to those of transgenic oilbody-oleosin technology in molecular farming. PMID:17214877

  10. Blue light regulates the accumulation of two psbD-psbC transcripts in barley chloroplasts.

    PubMed Central

    Gamble, P E; Mullet, J E

    1989-01-01

    Synthesis of D2, a Photosystem II reaction center protein encoded by psbD, is differentially maintained during light-induced chloroplast maturation. The continued synthesis of D2 is paralleled by selective light-induced accumulation of two psbD-psbC transcripts which share a common 5' terminus. In the present study, we examine the nature of the photoreceptor and the fluence requirement for psbD-psbC transcript induction. The light-induced change in psbD-psbC RNA population can be detected between 1 and 2 h after 4.5 day old dark-grown barley seedlings are transferred to the light. Light-induced transcript accumulation occurs normally in the chlorophyll-deficient barley mutant, xan-f10, indicating that light-activated chlorophyll formation and photosynthesis are not required for RNA induction. High fluence blue light fully induces psbD-psbC transcript accumulation; low or high fluence red or far-red light do not. However, psbD-psbC transcript accumulation elicited by blue light pulses can be partially attenuated if far-red light is given immediately following the blue light treatment. Thus, although blue light is needed to initiate transcript accumulation, phytochrome modulates the amplitude of the response. Pretreatment of dark-grown plants with cycloheximide blocks light-induced psbD-psbC transcript accumulation. This could implicate a blue-light responsive nuclear gene in the light-induced accumulation of the two psbD-psbC transcripts. Images PMID:2479534

  11. Phylogeography of East Asian Lespedeza buergeri (Fabaceae) based on chloroplast and nuclear ribosomal DNA sequence variations.

    PubMed

    Jin, Dong-Pil; Lee, Jung-Hyun; Xu, Bo; Choi, Byoung-Hee

    2016-09-01

    The dynamic changes in land configuration during the Quaternary that were accompanied by climatic oscillations have significantly influenced the current distribution and genetic structure of warm-temperate forests in East Asia. Although recent surveys have been conducted, the historical migration of forest species via land bridges and, especially, the origins of Korean populations remains conjectural. Here, we reveal the genetic structure of Lespedeza buergeri, a warm-temperate shrub that is disjunctively distributed around the East China Sea (ECS) at China, Korea, and Japan. Two non-coding regions (rpl32-trnL, psbA-trnH) of chloroplast DNA (cpDNA) and the internal transcribed spacer of nuclear ribosomal DNA (nrITS) were analyzed for 188 individuals from 16 populations, which covered almost all of its distribution. The nrITS data demonstrated a genetic structure that followed geographic boundaries. This examination utilized AMOVA, comparisons of genetic differentiation based on haplotype frequency/genetic mutations among haplotypes, and Mantel tests. However, the cpDNA data showed contrasting genetic pattern, implying that this difference was due to a slower mutation rate in cpDNA than in nrITS. These results indicated frequent migration by this species via an ECS land bridge during the early Pleistocene that then tapered gradually toward the late Pleistocene. A genetic isolation between western and eastern Japan coincided with broad consensus that was suggested by the presence of other warm-temperate plants in that country. For Korean populations, high genetic diversity indicated the existence of refugia during the Last Glacial Maximum on the Korean Peninsula. However, their closeness with western Japanese populations at the level of haplotype clade implied that gene flow from western Japanese refugia was possible until post-glacial processing occurred through the Korea/Tsushima Strait land bridge. PMID:27206725

  12. Chloroplast genome sequencing analysis of Heterosigma akashiwo CCMP452 (West Atlantic) and NIES293 (West Pacific) strains

    PubMed Central

    Cattolico, Rose Ann; Jacobs, Michael A; Zhou, Yang; Chang, Jean; Duplessis, Melinda; Lybrand, Terry; McKay, John; Ong, Han Chuan; Sims, Elizabeth; Rocap, Gabrielle

    2008-01-01

    Background Heterokont algae form a monophyletic group within the stramenopile branch of the tree of life. These organisms display wide morphological diversity, ranging from minute unicells to massive, bladed forms. Surprisingly, chloroplast genome sequences are available only for diatoms, representing two (Coscinodiscophyceae and Bacillariophyceae) of approximately 18 classes of algae that comprise this taxonomic cluster. A universal challenge to chloroplast genome sequencing studies is the retrieval of highly purified DNA in quantities sufficient for analytical processing. To circumvent this problem, we have developed a simplified method for sequencing chloroplast genomes, using fosmids selected from a total cellular DNA library. The technique has been used to sequence chloroplast DNA of two Heterosigma akashiwo strains. This raphidophyte has served as a model system for studies of stramenopile chloroplast biogenesis and evolution. Results H. akashiwo strain CCMP452 (West Atlantic) chloroplast DNA is 160,149 bp in size with a 21,822-bp inverted repeat, whereas NIES293 (West Pacific) chloroplast DNA is 159,370 bp in size and has an inverted repeat of 21,665 bp. The fosmid cloning technique reveals that both strains contain an isomeric chloroplast DNA population resulting from an inversion of their single copy domains. Both strains contain multiple small inverted and tandem repeats, non-randomly distributed within the genomes. Although both CCMP452 and NIES293 chloroplast DNAs contains 197 genes, multiple nucleotide polymorphisms are present in both coding and intergenic regions. Several protein-coding genes contain large, in-frame inserts relative to orthologous genes in other plastids. These inserts are maintained in mRNA products. Two genes of interest in H. akashiwo, not previously reported in any chloroplast genome, include tyrC, a tyrosine recombinase, which we hypothesize may be a result of a lateral gene transfer event, and an unidentified 456 amino acid

  13. The YlmG protein has a conserved function related to the distribution of nucleoids in chloroplasts and cyanobacteria

    PubMed Central

    2010-01-01

    Background Reminiscent of their free-living cyanobacterial ancestor, chloroplasts proliferate by division coupled with the partition of nucleoids (DNA-protein complexes). Division of the chloroplast envelope membrane is performed by constriction of the ring structures at the division site. During division, nucleoids also change their shape and are distributed essentially equally to the daughter chloroplasts. Although several components of the envelope division machinery have been identified and characterized, little is known about the molecular components/mechanisms underlying the change of the nucleoid structure. Results In order to identify new factors that are involved in the chloroplast division, we isolated Arabidopsis thaliana chloroplast division mutants from a pool of random cDNA-overexpressed lines. We found that the overexpression of a previously uncharacterized gene (AtYLMG1-1) of cyanobacterial origin results in the formation of an irregular network of chloroplast nucleoids, along with a defect in chloroplast division. In contrast, knockdown of AtYLMG1-1 resulted in a concentration of the nucleoids into a few large structures, but did not affect chloroplast division. Immunofluorescence microscopy showed that AtYLMG1-1 localizes in small puncta on thylakoid membranes, to which a subset of nucleoids colocalize. In addition, in the cyanobacterium Synechococcus elongates, overexpression and deletion of ylmG also displayed defects in nucleoid structure and cell division. Conclusions These results suggest that the proper distribution of nucleoids requires the YlmG protein, and the mechanism is conserved between cyanobacteria and chloroplasts. Given that ylmG exists in a cell division gene cluster downstream of ftsZ in gram-positive bacteria and that ylmG overexpression impaired the chloroplast division, the nucleoid partitioning by YlmG might be related to chloroplast and cyanobacterial division processes. PMID:20359373

  14. Identification of Nuclear Genes Encoding Chloroplast-Localized Proteins Required for Embryo Development in Arabidopsis1[W][OA

    PubMed Central

    Bryant, Nicole; Lloyd, Johnny; Sweeney, Colleen; Myouga, Fumiyoshi; Meinke, David

    2011-01-01

    We describe here the diversity of chloroplast proteins required for embryo development in Arabidopsis (Arabidopsis thaliana). Interfering with certain chloroplast functions has long been known to result in embryo lethality. What has not been reported before is a comprehensive screen for embryo-defective (emb) mutants altered in chloroplast proteins. From a collection of transposon and T-DNA insertion lines at the RIKEN chloroplast function database (http://rarge.psc.riken.jp/chloroplast/) that initially appeared to lack homozygotes and segregate for defective seeds, we identified 23 additional examples of EMB genes that likely encode chloroplast-localized proteins. Fourteen gene identities were confirmed with allelism tests involving duplicate mutant alleles. We then queried journal publications and the SeedGenes database (www.seedgenes.org) to establish a comprehensive dataset of 381 nuclear genes encoding chloroplast proteins of Arabidopsis associated with embryo-defective (119 genes), plant pigment (121 genes), gametophyte (three genes), and alternate (138 genes) phenotypes. Loci were ranked based on the level of certainty that the gene responsible for the phenotype had been identified and the protein product localized to chloroplasts. Embryo development is frequently arrested when amino acid, vitamin, or nucleotide biosynthesis is disrupted but proceeds when photosynthesis is compromised and when levels of chlorophyll, carotenoids, or terpenoids are reduced. Chloroplast translation is also required for embryo development, with genes encoding chloroplast ribosomal and pentatricopeptide repeat proteins well represented among EMB datasets. The chloroplast accD locus, which is necessary for fatty acid biosynthesis, is essential in Arabidopsis but not in Brassica napus or maize (Zea mays), where duplicated nuclear genes compensate for its absence or loss of function. PMID:21139083

  15. Expression of the Native Cholera Toxin B Subunit Gene and Assembly as Functional Oligomers in Transgenic Tobacco Chloroplasts

    PubMed Central

    Daniell, Henry; Lee, Seung-Bum; Panchal, Tanvi; Wiebe, Peter O.

    2012-01-01

    The B subunits of enterotoxigenic Escherichia coli (LTB) and cholera toxin of Vibrio cholerae (CTB) are candidate vaccine antigens. Integration of an unmodified CTB-coding sequence into chloroplast genomes (up to 10,000 copies per cell), resulted in the accumulation of up to 4.1% of total soluble tobacco leaf protein as functional oligomers (410-fold higher expression levels than that of the unmodified LTB gene expressed via the nuclear genome). However, expresssion levels reported are an underestimation of actual accumulation of CTB in transgenic chloroplasts, due to aggregation of the oligomeric forms in unboiled samples similar to the aggregation observed for purified bacterial antigen. PCR and Southern blot analyses confirmed stable integration of the CTB gene into the chloroplast genome. Western blot analysis showed that the chloroplast-synthesized CTB assembled into oligomers and were antigenically identical with purified native CTB. Also, binding assays confirmed that chloroplast- synthesized CTB binds to the intestinal membrane GM1-ganglioside receptor, indicating correct folding and disulfide bond formation of CTB pentamers within transgenic chloroplasts. In contrast to stunted nuclear transgenic plants, chloroplast transgenic plants were morphologically indistinguishable from untransformed plants, when CTB was constitutively expressed in chloroplasts. Introduced genes were inherited stably in subsequent generations, as confirmed by PCR and Southern blot analyses. Increased production of an efficient transmucosal carrier molecule and delivery system, like CTB, in transgenic chloroplasts makes plant-based oral vaccines and fusion proteins with CTB needing oral administration commercially feasible. Successful expression of foreign genes in transgenic chromoplasts and availability of marker-free chloroplast transformation techniques augurs well for development of vaccines in edible parts of transgenic plants. Furthermore, since the quaternary structure of

  16. The effect of UV-B radiation on chloroplast translation in Pisum sativum

    SciTech Connect

    Raab, M.M.; Jagendorf, A.T. )

    1990-05-01

    UV-B radiation has previously been reported to reduce growth, flowering, and net photosynthesis. The present study examines the effect of UV-B radiation on isolated chloroplast of 7-10 day old pea seedlings. Amount of ({sup 3}H)-Leu incorporated into isolated chloroplasts was measured in the presence or absence of UV-B exposure. Preliminary experiments show a 30% inhibition of protein synthesis in isolated chloroplasts after only 20 mins of UV-B exposure (6.9 J/m{sup 2}/30 min). Percent inhibition of chloroplast translation is directly correlated with UV-B exposure over a 60 min time span. Preliminary studies also show no change in both cold and radiolabeled protein profiles as expressed on 1-D PAGE and autofluorography. Comparative studies on the sensitivity of e{sup {minus}} flow vs protein synthesis following UV-B exposure are underway. Further work on the role of oxygen free radicals and the specific site of action of UV-B damage to the translation machinery of chloroplasts will be discussed.

  17. Native architecture of the Chlamydomonas chloroplast revealed by in situ cryo-electron tomography

    PubMed Central

    Engel, Benjamin D; Schaffer, Miroslava; Kuhn Cuellar, Luis; Villa, Elizabeth; Plitzko, Jürgen M; Baumeister, Wolfgang

    2015-01-01

    Chloroplast function is orchestrated by the organelle's intricate architecture. By combining cryo-focused ion beam milling of vitreous Chlamydomonas cells with cryo-electron tomography, we acquired three-dimensional structures of the chloroplast in its native state within the cell. Chloroplast envelope inner membrane invaginations were frequently found in close association with thylakoid tips, and the tips of multiple thylakoid stacks converged at dynamic sites on the chloroplast envelope, implicating lipid transport in thylakoid biogenesis. Subtomogram averaging and nearest neighbor analysis revealed that RuBisCO complexes were hexagonally packed within the pyrenoid, with ∼15 nm between their centers. Thylakoid stacks and the pyrenoid were connected by cylindrical pyrenoid tubules, physically bridging the sites of light-dependent photosynthesis and light-independent carbon fixation. Multiple parallel minitubules were bundled within each pyrenoid tubule, possibly serving as conduits for the targeted one-dimensional diffusion of small molecules such as ATP and sugars between the chloroplast stroma and the pyrenoid matrix. DOI: http://dx.doi.org/10.7554/eLife.04889.001 PMID:25584625

  18. Complete Chloroplast Genome Sequence of Omani Lime (Citrus aurantiifolia) and Comparative Analysis within the Rosids

    PubMed Central

    Su, Huei-Jiun; Hogenhout, Saskia A.; Al-Sadi, Abdullah M.; Kuo, Chih-Horng

    2014-01-01

    The genus Citrus contains many economically important fruits that are grown worldwide for their high nutritional and medicinal value. Due to frequent hybridizations among species and cultivars, the exact number of natural species and the taxonomic relationships within this genus are unclear. To compare the differences between the Citrus chloroplast genomes and to develop useful genetic markers, we used a reference-assisted approach to assemble the complete chloroplast genome of Omani lime (C. aurantiifolia). The complete C. aurantiifolia chloroplast genome is 159,893 bp in length; the organization and gene content are similar to most of the rosids lineages characterized to date. Through comparison with the sweet orange (C. sinensis) chloroplast genome, we identified three intergenic regions and 94 simple sequence repeats (SSRs) that are potentially informative markers with resolution for interspecific relationships. These markers can be utilized to better understand the origin of cultivated Citrus. A comparison among 72 species belonging to 10 families of representative rosids lineages also provides new insights into their chloroplast genome evolution. PMID:25398081

  19. Sub-plastidial localization of two different phage-type RNA polymerases in spinach chloroplasts

    PubMed Central

    Azevedo, Jacinthe; Courtois, Florence; Lerbs-Mache, Silva

    2006-01-01

    Plant plastids contain a circular genome of ∼150 kb organized into ∼35 transcription units. The plastid genome is organized into nucleoids and attached to plastid membranes. This relatively small genome is transcribed by at least two different RNA polymerases, one being of the prokaryotic type and plastid-encoded (PEP), the other one being of the phage-type and nucleus-encoded (NEP). The presumed localization of a second phage-type RNA polymerase in plastids is still questionable. There is strong evidence for a sequential action of NEP and PEP enzymes during plant development attributing a prevailing role of NEP during early plant and plastid development, although NEP is present in mature chloroplasts. In the present paper, we have analysed two different NEP enzymes from spinach with respect to subcellular and intra-plastidial localization in mature chloroplasts with the help of specific antibodies. Results show the presence of the two different NEP enzymes in mature chloroplasts. Both enzymes are entirely membrane bound but, unlike previously thought, this membrane binding is not mediated via DNA. This finding indicates that NEP enzymes are not found as elongating transcription complexes on the template DNA in mature chloroplasts and raises the question of their function in mature chloroplasts. PMID:16421271

  20. Discrete Redox Signaling Pathways Regulate Photosynthetic Light-Harvesting and Chloroplast Gene Transcription

    PubMed Central

    Allen, John F.; Santabarbara, Stefano; Allen, Carol A.; Puthiyaveetil, Sujith

    2011-01-01

    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. PMID:22039472

  1. The tetratricopeptide repeat-containing protein slow green1 is required for chloroplast development in Arabidopsis

    PubMed Central

    Hu, Zhihong; Xu, Fan; Hou, Suiwen

    2014-01-01

    A new gene, SG1, was identified in a slow-greening mutant (sg1) isolated from an ethylmethanesulphonate-mutagenized population of Arabidopsis thaliana. The newly formed leaves of sg1 were initially albino, but gradually became pale green. After 3 weeks, the leaves of the mutant were as green as those of the wild-type plants. Transmission electron microscopic observations revealed that the mutant displayed delayed proplastid to chloroplast transition. The results of map-based cloning showed that SG1 encodes a chloroplast-localized tetratricopeptide repeat-containing protein. Quantitative real-time reverse transcription–PCR data demonstrated the presence of SG1 gene expression in all tissues, particularly young green tissues. The sg1 mutation disrupted the expression levels of several genes associated with chloroplast development, photosynthesis, and chlorophyll biosynthesis. The results of genetic analysis indicated that gun1 and gun4 partially restored the expression patterns of the previously detected chloroplast-associated genes, thereby ameliorating the slow-greening phenotype of sg1. Taken together, the results suggest that the newly identified protein, SG1, is required for chloroplast development in Arabidopsis. PMID:24420572

  2. Chloroplast translocations in Lemna trisulca L. induced by continuous irradiation and by light pulses : Kinetic analysis.

    PubMed

    Zurzycki, J; Walczak, T; Gabryś, H; Kajfosz, J

    1983-05-01

    The analytical model describing the steady state position of chloroplasts in dependence of fluence rate as well as the chloroplast response to single strong light pulses has been proposed. The model is based on the following assumptions: 1. Irradiation of the cell generates the state X in the cell membrane region, proportional to the local fluence rate. After switching on the light, the value of X increases exponentially with the time constant of about 3 min. The dark decay of X is also exponential with the same time constant. The level of X controls all kinds of chloroplast arrangements. 2. The state X generates two further states: Y 1 and Y 2, the first of them representing attraction forces for chloroplasts and the second representing repulsion forces. Empirical equations have been found for both Y states. The fluence rate response curve can be described with the use of functions Y 1 and Y 2. 3. The kinetic analysis requires the introduction of two additional functions Z in order to account for delays and time dispersion of the chloroplast movement in response to driving and resistance factors. The computer program for the proposed model was developed and the results of calculations were compared with experimental data (fluence rate response curve and pulse effects) with satisfactory agreement. Initially no attempt was made to ascribe any physical meaning to the postulated states. Some suggestions in this respect are mentioned in the discussion. PMID:24264414

  3. SPAD chlorophyll meter reading can be pronouncedly affected by chloroplast movement.

    PubMed

    Nauš, Jan; Prokopová, Jitka; Rebíček, Jiří; Spundová, Martina

    2010-09-01

    Non-destructive assessment of chlorophyll content has recently been widely done by chlorophyll meters based on measurement of leaf transmittance (e.g. the SPAD-502 chlorophyll meter measures the leaf transmittance at 650 and 940 nm). However, the leaf transmittance depends not only on the content of chlorophylls but also on their distribution in leaves. The chlorophyll distribution within leaves is co-determined by chloroplast arrangement in cells that depends on light conditions. When tobacco leaves were exposed to a strong blue light (about 340 μmol of photons m⁻² s⁻¹), a very pronounced increase in the leaf transmittance was observed as chloroplasts migrated from face position (along cell walls perpendicular to the incident light) to side position (along cell walls parallel to the incoming light) and the SPAD reading decreased markedly. This effect was more pronounced in the leaves of young tobacco plants compared with old ones; the difference between SPAD values in face and side position reached even about 35%. It is shown how the chloroplast movement changes a relationship between the SPAD readings and real chlorophyll content. For an elimination of the chloroplast movement effect, it can be recommended to measure the SPAD values in leaves with a defined chloroplasts arrangement. PMID:20661644

  4. CHLOROPLAST GENETIC TOOL FOR THE GREEN MICROALGAE HAEMATOCOCCUS PLUVIALIS (CHLOROPHYCEAE, VOLVOCALES)(1).

    PubMed

    Gutiérrez, Carla L; Gimpel, Javier; Escobar, Carolina; Marshall, Sergio H; Henríquez, Vitalia

    2012-08-01

    At present, there is strong commercial demand for recombinant proteins, such as antigens, antibodies, biopharmaceuticals, and industrial enzymes, which cannot be fulfilled by existing procedures. Thus, an intensive search for alternative models that may provide efficiency, safety, and quality control is being undertaken by a number of laboratories around the world. The chloroplast of the eukaryotic microalgae Haematococcus pluvialis Flotow has arisen as a candidate for a novel expression platform for recombinant protein production. However, there are important drawbacks that need to be resolved before it can become such a system. The most significant of these are chloroplast genome characterizations, and the development of chloroplast transformation vectors based upon specific endogenous promoters and on homologous targeting regions. In this study, we report the identification and characterization of endogenous chloroplast sequences for use as genetic tools for the construction of H. pluvialis specific expression vectors to efficiently transform the chloroplast of this microalga via microprojectile bombardment. As a consequence, H. pluvialis shows promise as a platform for expressing recombinant proteins for biotechnological applications, for instance, the development of oral vaccines for aquaculture. PMID:27009007

  5. Overoxidation of chloroplast 2-Cys peroxiredoxins: balancing toxic and signaling activities of hydrogen peroxide

    PubMed Central

    Puerto-Galán, Leonor; Pérez-Ruiz, Juan M.; Ferrández, Julia; Cano, Beatriz; Naranjo, Belén; Nájera, Victoria A.; González, Maricruz; Lindahl, Anna M.; Cejudo, Francisco J.

    2013-01-01

    Photosynthesis, the primary source of biomass and oxygen into the biosphere, involves the transport of electrons in the presence of oxygen and, therefore, chloroplasts constitute an important source of reactive oxygen species, including hydrogen peroxide. If accumulated at high level, hydrogen peroxide may exert a toxic effect; however, it is as well an important second messenger. In order to balance the toxic and signaling activities of hydrogen peroxide its level has to be tightly controlled. To this end, chloroplasts are equipped with different antioxidant systems such as 2-Cys peroxiredoxins (2-Cys Prxs), thiol-based peroxidases able to reduce hydrogen and organic peroxides. At high peroxide concentrations the peroxidase function of 2-Cys Prxs may become inactivated through a process of overoxidation. This inactivation has been proposed to explain the signaling function of hydrogen peroxide in eukaryotes, whereas in prokaryotes, the 2-Cys Prxs of which were considered to be insensitive to overoxidation, the signaling activity of hydrogen peroxide is less relevant. Here we discuss the current knowledge about the mechanisms controlling 2-Cys Prx overoxidation in chloroplasts, organelles with an important signaling function in plants. Given the prokaryotic origin of chloroplasts, we discuss the occurrence of 2-Cys Prx overoxidation in cyanobacteria with the aim of identifying similarities between chloroplasts and their ancestors regarding their response to hydrogen peroxide. PMID:23967002

  6. Effects of Pb2+ on energy distribution and photochemical activity of spinach chloroplast.

    PubMed

    Wu, Xiao; Hong, Fashui; Liu, Chao; Su, Mingyu; Zheng, Lei; Gao, Fengqing; Yang, Fan

    2008-03-01

    Lead (Pb(2+)) is a well-known highly toxic element. The mechanisms of the Pb(2+) toxicity are not well understood for photosynthesis. In this paper, we reported the effect of Pb(2+) on light absorption, distribution and conversion of spinach chloroplast by spectroscopy, and photochemical reaction activities. Several effects of Pb(2+) were observed: (1) the absorption peak intensity of chloroplast obviously decreased in red and blue region and produced optical flattering; (2) fluorescence quantum yield nearby 680 nm of chloroplast greatly declined; (3) the excitation band nearby 440 nm of chloroplast significantly descended; (4) Pb(2+) treatments reduced of the rate of whole chain electron transport, photochemical activities of PSII DCPIP photoreduction and oxygen evolution, but the photoreduction activities of PSI were little changed. Together, the studies of the experiments showed that Pb(2+) decreased absorption of light on spinach chloroplast and inhibited excitation energy to be absorbed by LHCII and transferred to PSII, then reduced the conversion from light energy to electron energy, and decelerated electron transport, water photolysis and oxygen evolution. PMID:17602861

  7. Chloroplast lipid transfer processes in Chlamydomonas reinhardtii involving a TRIGALACTOSYLDIACYLGLYCEROL 2 (TGD2) orthologue.

    PubMed

    Warakanont, Jaruswan; Tsai, Chia-Hong; Michel, Elena J S; Murphy, George R; Hsueh, Peter Y; Roston, Rebecca L; Sears, Barbara B; Benning, Christoph

    2015-12-01

    In plants, lipids of the photosynthetic membrane are synthesized by parallel pathways associated with the endoplasmic reticulum (ER) and the chloroplast envelope membranes. Lipids derived from the two pathways are distinguished by their acyl-constituents. Following this plant paradigm, the prevalent acyl composition of chloroplast lipids suggests that Chlamydomonas reinhardtii (Chlamydomonas) does not use the ER pathway; however, the Chlamydomonas genome encodes presumed plant orthologues of a chloroplast lipid transporter consisting of TGD (TRIGALACTOSYLDIACYLGLYCEROL) proteins that are required for ER-to-chloroplast lipid trafficking in plants. To resolve this conundrum, we identified a mutant of Chlamydomonas deleted in the TGD2 gene and characterized the respective protein, CrTGD2. Notably, the viability of the mutant was reduced, showing the importance of CrTGD2. Galactoglycerolipid metabolism was altered in the tgd2 mutant with monogalactosyldiacylglycerol (MGDG) synthase activity being strongly stimulated. We hypothesize this to be a result of phosphatidic acid accumulation in the chloroplast outer envelope membrane, the location of MGDG synthase in Chlamydomonas. Concomitantly, increased conversion of MGDG into triacylglycerol (TAG) was observed. This TAG accumulated in lipid droplets in the tgd2 mutant under normal growth conditions. Labeling kinetics indicate that Chlamydomonas can import lipid precursors from the ER, a process that is impaired in the tgd2 mutant. PMID:26496373

  8. The Complete Chloroplast Genome Sequences of the Medicinal Plant Pogostemon cablin.

    PubMed

    He, Yang; Xiao, Hongtao; Deng, Cao; Xiong, Liang; Yang, Jian; Peng, Cheng

    2016-01-01

    Pogostemon cablin, the natural source of patchouli alcohol, is an important herb in the Lamiaceae family. Here, we present the entire chloroplast genome of P. cablin. This genome, with 38.24% GC content, is 152,460 bp in length. The genome presents a typical quadripartite structure with two inverted repeats (each 25,417 bp in length), separated by one small and one large single-copy region (17,652 and 83,974 bp in length, respectively). The chloroplast genome encodes 127 genes, of which 107 genes are single-copy, including 79 protein-coding genes, four rRNA genes, and 24 tRNA genes. The genome structure, GC content, and codon usage of this chloroplast genome are similar to those of other species in the family, except that it encodes less protein-coding genes and tRNA genes. Phylogenetic analysis reveals that P. cablin diverged from the Scutellarioideae clade about 29.45 million years ago (Mya). Furthermore, most of the simple sequence repeats (SSRs) are short polyadenine or polythymine repeats that contribute to high AT content in the chloroplast genome. Complete sequences and annotation of P. cablin chloroplast genome will facilitate phylogenic, population and genetic engineering research investigations involving this particular species. PMID:27275817

  9. S1 domain-containing STF modulates plastid transcription and chloroplast biogenesis in Nicotiana benthamiana.

    PubMed

    Jeon, Young; Jung, Hyun Ju; Kang, Hunseung; Park, Youn-Il; Lee, Soon Hee; Pai, Hyun-Sook

    2012-01-01

    • In this study, we examined the biochemical and physiological functions of Nicotiana benthamiana S1 domain-containing Transcription-Stimulating Factor (STF) using virus-induced gene silencing (VIGS), cosuppression, and overexpression strategies. • STF : green fluorescent protein (GFP) fusion protein colocalized with sulfite reductase (SiR), a chloroplast nucleoid-associated protein also present in the stroma. Full-length STF and its S1 domain preferentially bound to RNA, probably in a sequence-nonspecific manner. • STF silencing by VIGS or cosuppression resulted in severe leaf yellowing caused by disrupted chloroplast development. STF deficiency significantly perturbed plastid-encoded multimeric RNA polymerase (PEP)-dependent transcript accumulation. Chloroplast transcription run-on assays revealed that the transcription rate of PEP-dependent plastid genes was reduced in the STF-silenced leaves. Conversely, the exogenously added recombinant STF protein increased the transcription rate, suggesting a direct role of STF in plastid transcription. Etiolated seedlings of STF cosuppression lines showed defects in the light-triggered transition from etioplasts to chloroplasts, accompanied by reduced light-induced expression of plastid-encoded genes. • These results suggest that STF plays a critical role as an auxiliary factor of the PEP transcription complex in the regulation of plastid transcription and chloroplast biogenesis in higher plants. PMID:22050604

  10. Increased resistance to oxidative stress in transgenic plants by targeting mannitol biosynthesis to chloroplasts

    SciTech Connect

    Shen, B.; Jensen, R.G.; Bohnert, H.J.

    1997-04-01

    To investigate the potential role of a polyol, mannitol, in oxidative stress protection, a bacterial mannitol-1-phosphate dehydrogenase gene was targeted to chloroplasts by the addition of an aminoterminal transit peptide. Transgenic tobacco (Nicotiana tabacum) lines accumulate mannitol at concentrations ranging from 2.5 to 7 {mu}mol/g fresh weight. Line BS1-31 accumulated approximately 100 mm mannitol in chloroplasts and was identical to the wild type in phenotype and photosynthetic performance. The presence of mannitol in chloroplasts resulted in an increased resistance to methyl viologen (MV)-induced oxidative stress, documented by the increased retention of chlorophyll in transgenic leaf tissue following MV treatment. In the presence of MV, isolated mesophyll cells of BS1-31 exhibited higher CO{sub 2} fixation than the wild type. When the hydroxyl radical probe dimethyl sulfoxide was introduced into cells, the initial formation rate of methane sulfinic acid was significantly lower in cells containing mannitol in the chloroplast compartment than in wild-type cells, indicating an increased hydroxyl radical-scavenging capacity in BS1-31 tobacco. We suggest that the chloroplast location of mannitol can supplement endogenous radical-scavenging mechanisms and reduce oxidative damage of cells by hydroxyl radicals. 43 refs., 5 figs., 1 tab.

  11. A Small Family of Chloroplast Atypical Thioredoxins1[C][W][OA

    PubMed Central

    Dangoor, Inbal; Peled-Zehavi, Hadas; Levitan, Alexander; Pasand, Ohad; Danon, Avihai

    2009-01-01

    The reduction and the formation of regulatory disulfide bonds serve as a key signaling element in chloroplasts. Members of the thioredoxin (Trx) superfamily of oxidoreductases play a major role in these processes. We have characterized a small family of plant-specific Trxs in Arabidopsis (Arabidopsis thaliana) that are rich in cysteine and histidine residues and are typified by a variable noncanonical redox active site. We found that the redox midpoint potential of three selected family members is significantly less reducing than that of the classic Trxs. Assays of subcellular localization demonstrated that all proteins are localized to the chloroplast. Selected members showed high activity, contingent on a dithiol electron donor, toward the chloroplast 2-cysteine peroxiredoxin A and poor activity toward the chloroplast NADP-malate dehydrogenase. The expression profile of the family members suggests that they have distinct roles. The intermediate redox midpoint potential value of the atypical Trxs might imply adaptability to function in modulating the redox state of chloroplast proteins with regulatory disulfides. PMID:19109414

  12. Effects of Pb 2+ on energy distribution and photochemical activity of spinach chloroplast

    NASA Astrophysics Data System (ADS)

    Wu, Xiao; Hong, Fashui; Liu, Chao; Su, Mingyu; Zheng, Lei; Gao, Fengqing; Yang, Fan

    2008-03-01

    Lead (Pb 2+) is a well-known highly toxic element. The mechanisms of the Pb 2+ toxicity are not well understood for photosynthesis. In this paper, we reported the effect of Pb 2+ on light absorption, distribution and conversion of spinach chloroplast by spectroscopy, and photochemical reaction activities. Several effects of Pb 2+ were observed: (1) the absorption peak intensity of chloroplast obviously decreased in red and blue region and produced optical flattering; (2) fluorescence quantum yield nearby 680 nm of chloroplast greatly declined; (3) the excitation band nearby 440 nm of chloroplast significantly descended; (4) Pb 2+ treatments reduced of the rate of whole chain electron transport, photochemical activities of PSII DCPIP photoreduction and oxygen evolution, but the photoreduction activities of PSI were little changed. Together, the studies of the experiments showed that Pb 2+ decreased absorption of light on spinach chloroplast and inhibited excitation energy to be absorbed by LHCII and transferred to PSII, then reduced the conversion from light energy to electron energy, and decelerated electron transport, water photolysis and oxygen evolution.

  13. The Complete Chloroplast Genome Sequences of Three Veroniceae Species (Plantaginaceae): Comparative Analysis and Highly Divergent Regions

    PubMed Central

    Choi, Kyoung Su; Chung, Myong Gi; Park, SeonJoo

    2016-01-01

    Previous studies of Veronica and related genera were weakly supported by molecular and paraphyletic taxa. Here, we report the complete chloroplast genome sequence of Veronica nakaiana and the related species Veronica persica and Veronicastrum sibiricum. The chloroplast genome length of V. nakaiana, V. persica, and V. sibiricum ranged from 150,198 bp to 152,930 bp. A total of 112 genes comprising 79 protein coding genes, 29 tRNA genes, and 4 rRNA genes were observed in three chloroplast genomes. The total number of SSRs was 48, 51, and 53 in V. nakaiana, V. persica, and V. sibiricum, respectively. Two SSRs (10 bp of AT and 12 bp of AATA) were observed in the same regions (rpoC2 and ndhD) in three chloroplast genomes. A comparison of coding genes and non-coding regions between V. nakaiana and V. persica revealed divergent sites, with the greatest variation occurring petD-rpoA region. The complete chloroplast genome sequence information regarding the three Veroniceae will be helpful for elucidating Veroniceae phylogenetic relationships. PMID:27047524

  14. Thiol modulation of the chloroplast ATP synthase is dependent on the energization of thylakoid membranes.

    PubMed

    Konno, Hiroki; Nakane, Takeshi; Yoshida, Masasuke; Ueoka-Nakanishi, Hanayo; Hara, Satoshi; Hisabori, Toru

    2012-04-01

    Thiol modulation of the chloroplast ATP synthase γ subunit has been recognized as an important regulatory system for the activation of ATP hydrolysis activity, although the physiological significance of this regulation system remains poorly characterized. Since the membrane potential required by this enzyme to initiate ATP synthesis for the reduced enzyme is lower than that needed for the oxidized form, reduction of this enzyme was interpreted as effective regulation for efficient photophosphorylation. However, no concrete evidence has been obtained to date relating to the timing and mode of chloroplast ATP synthase reduction and oxidation in green plants. In this study, thorough analysis of the redox state of regulatory cysteines of the chloroplast ATP synthase γ subunit in intact chloroplasts and leaves shows that thiol modulation of this enzyme is pivotal in prohibiting futile ATP hydrolysis activity in the dark. However, the physiological importance of efficient ATP synthesis driven by the reduced enzyme in the light could not be demonstrated. In addition, we investigated the significance of the electrochemical proton gradient in reducing the γ subunit by the reduced form of thioredoxin in chloroplasts, providing strong insights into the molecular mechanisms underlying the formation and reduction of the disulfide bond on the γ subunit in vivo. PMID:22362842

  15. Post transcriptional regulation of chloroplast gene expression by nuclear encoded gene products

    SciTech Connect

    Kuchka, M.R.

    1992-01-01

    Many individual chloroplast genes require the products of a collection of nuclear genes for their successful expression. These nuclear gene products apparently work with great specificity, each committed to the expression of a single chloroplast gene. We have chosen as a model nuclear mutants of Chlamydomonas affected in different stages in the expression of the chloroplast encoded Photosystem II polypeptide, D2. We have made the progress in understanding how nuclear gene products affect the translation of the D2 encoding MRNA. Two nuclear genes are required for this process which have been mapped genetically. In contrast to other examples of nuclear control of translation in the chloroplast, these nuclear gene products appear to be required either for specific stages in translation elongation or for the post-translational stabilization of the nascent D2 protein. Pseudoreversion analysis has led us to a locus which may be directly involved in D2 expression. We have made considerable progress in pursuing the molecular basis of psbd MRNA stabilization. psbD 5' UTR specific transcripts have been synthesized in vitro and used in gel mobility shift assays. UV-crosslinking studies are underway to identify the transacting factors which bind to these sequences. The continued examination of these mutants will help us to understand how nuclear gene products work in this specific case of chloroplast gene expression, and will elucidate how two distinct genomes can interact generally.

  16. Enhanced green fluorescent protein (egfp) gene expression in Tetraselmis subcordiformis chloroplast with endogenous regulators.

    PubMed

    Cui, Yulin; Zhao, Jialin; Hou, Shichang; Qin, Song

    2016-05-01

    On the basis of fundamental genetic transformation technologies, the goal of this study was to optimize Tetraselmis subcordiformis chloroplast transformation through the use of endogenous regulators. The genes rrn16S, rbcL, psbA, and psbC are commonly highly expressed in chloroplasts, and the regulators of these genes are often used in chloroplast transformation. For lack of a known chloroplast genome sequence, the genome-walking method was used here to obtain full sequences of T. subcordiformis endogenous regulators. The resulting regulators, including three promoters, two terminators, and a ribosome combination sequence, were inserted into the previously constructed plasmid pPSC-R, with the egfp gene included as a reporter gene, and five chloroplast expression vectors prepared. These vectors were successfully transformed into T. subcordiformis by particle bombardment and the efficiency of each vector tested by assessing EGFP fluorescence via microscopy. The results showed that these vectors exhibited higher efficiency than the former vector pPSC-G carrying exogenous regulators, and the vector pRFA with Prrn, psbA-5'RE, and TpsbA showed the highest efficiency. This research provides a set of effective endogenous regulators for T. subcordiformis and will facilitate future fundamental studies of this alga. PMID:27038953

  17. A Cyan Fluorescent Reporter Expressed from the Chloroplast Genome of Marchantia polymorpha.

    PubMed

    Boehm, Christian R; Ueda, Minoru; Nishimura, Yoshiki; Shikanai, Toshiharu; Haseloff, Jim

    2016-02-01

    Recently, the liverwort Marchantia polymorpha has received increasing attention as a basal plant model for multicellular studies. Its ease of handling, well-characterized plastome and proven protocols for biolistic plastid transformation qualify M. polymorpha as an attractive platform to study the evolution of chloroplasts during the transition from water to land. In addition, chloroplasts of M. polymorpha provide a convenient test-bed for the characterization of genetic elements involved in plastid gene expression due to the absence of mechanisms for RNA editing. While reporter genes have proven valuable to the qualitative and quantitative study of gene expression in chloroplasts, expression of green fluorescent protein (GFP) in chloroplasts of M. polymorpha has proven problematic. We report the design of a codon-optimized gfp varian, mturq2cp, which allowed successful expression of a cyan fluorescent protein under control of the tobacco psbA promoter from the chloroplast genome of M. polymorpha. We demonstrate the utility of mturq2cp in (i) early screening for transplastomic events following biolistic transformation of M. polymorpha spores; (ii) visualization of stromules as elements of plastid structure in Marchantia; and (iii) quantitative microscopy for the analysis of promoter activity. PMID:26634291

  18. A simple, low-cost method for chloroplast transformation of the green alga Chlamydomonas reinhardtii.

    PubMed

    Economou, Chloe; Wannathong, Thanyanan; Szaub, Joanna; Purton, Saul

    2014-01-01

    The availability of routine techniques for the genetic manipulation of the chloroplast genome of Chlamydomonas reinhardtii has allowed a plethora of reverse-genetic studies of chloroplast biology using this alga as a model organism. These studies range from fundamental investigations of chloroplast gene function and regulation to sophisticated metabolic engineering programs and to the development of the algal chloroplast as a platform for producing high-value recombinant proteins. The established method for delivering transforming DNA into the Chlamydomonas chloroplast involves microparticle bombardment, with the selection of transformant lines most commonly involving the use of antibiotic resistance markers. In this chapter we describe a simpler and cheaper delivery method in which cell/DNA suspensions are agitated with glass beads: a method that is more commonly used for nuclear transformation of Chlamydomonas. Furthermore, we highlight the use of an expression vector (pASapI) that employs an endogenous gene as a selectable marker, thereby avoiding the contentious issue of antibiotic resistance determinants in transgenic lines. PMID:24599870

  19. A Cyan Fluorescent Reporter Expressed from the Chloroplast Genome of Marchantia polymorpha

    PubMed Central

    Boehm, Christian R.; Ueda, Minoru; Nishimura, Yoshiki; Shikanai, Toshiharu; Haseloff, Jim

    2016-01-01

    Recently, the liverwort Marchantia polymorpha has received increasing attention as a basal plant model for multicellular studies. Its ease of handling, well-characterized plastome and proven protocols for biolistic plastid transformation qualify M. polymorpha as an attractive platform to study the evolution of chloroplasts during the transition from water to land. In addition, chloroplasts of M. polymorpha provide a convenient test-bed for the characterization of genetic elements involved in plastid gene expression due to the absence of mechanisms for RNA editing. While reporter genes have proven valuable to the qualitative and quantitative study of gene expression in chloroplasts, expression of green fluorescent protein (GFP) in chloroplasts of M. polymorpha has proven problematic. We report the design of a codon-optimized gfp varian, mturq2cp, which allowed successful expression of a cyan fluorescent protein under control of the tobacco psbA promoter from the chloroplast genome of M. polymorpha. We demonstrate the utility of mturq2cp in (i) early screening for transplastomic events following biolistic transformation of M. polymorpha spores; (ii) visualization of stromules as elements of plastid structure in Marchantia; and (iii) quantitative microscopy for the analysis of promoter activity. PMID:26634291

  20. Purification of intact chloroplasts from marine plant Posidonia oceanica suitable for organelle proteomics.

    PubMed

    Piro, Amalia; Serra, Ilia Anna; Spadafora, Antonia; Cardilio, Monica; Bianco, Linda; Perrotta, Gaetano; Santos, Rui; Mazzuca, Silvia

    2015-12-01

    Posidonia oceanica is a marine angiosperm, or seagrass, adapted to grow to the underwater life from shallow waters to 50 m depth. This raises questions of how their photosynthesis adapted to the attenuation of light through the water column and leads to the assumption that biochemistry and metabolism of the chloroplast are the basis of adaptive capacity. In the present study, we described a protocol that was adapted from those optimized for terrestrial plants, to extract chloroplasts from as minimal tissue as possible. We obtained the best balance between tissue amount/intact chloroplasts yield using one leaf from one plant. After isopynic separations, the chloroplasts purity and integrity were evaluated by biochemical assay and using a proteomic approach. Chloroplast proteins were extracted from highly purified organelles and resolved by 1DE SDS-PAGE. Proteins were sequenced by nLC-ESI-IT-MS/MS of 1DE gel bands and identified against NCBInr green plant databases, Dr. Zompo database for seagrasses in a local customized dataset. The curated localization of proteins in sub-plastidial compartments (i.e. envelope, stroma and thylakoids) was retrieved in the AT_CHLORO database. This purification protocol and the validation of compartment markers may serve as basis for sub-cellular proteomics in P. oceanica and other seagrasses. PMID:26444578

  1. The Complete Chloroplast Genome Sequences of the Medicinal Plant Pogostemon cablin

    PubMed Central

    He, Yang; Xiao, Hongtao; Deng, Cao; Xiong, Liang; Yang, Jian; Peng, Cheng

    2016-01-01

    Pogostemon cablin, the natural source of patchouli alcohol, is an important herb in the Lamiaceae family. Here, we present the entire chloroplast genome of P. cablin. This genome, with 38.24% GC content, is 152,460 bp in length. The genome presents a typical quadripartite structure with two inverted repeats (each 25,417 bp in length), separated by one small and one large single-copy region (17,652 and 83,974 bp in length, respectively). The chloroplast genome encodes 127 genes, of which 107 genes are single-copy, including 79 protein-coding genes, four rRNA genes, and 24 tRNA genes. The genome structure, GC content, and codon usage of this chloroplast genome are similar to those of other species in the family, except that it encodes less protein-coding genes and tRNA genes. Phylogenetic analysis reveals that P. cablin diverged from the Scutellarioideae clade about 29.45 million years ago (Mya). Furthermore, most of the simple sequence repeats (SSRs) are short polyadenine or polythymine repeats that contribute to high AT content in the chloroplast genome. Complete sequences and annotation of P. cablin chloroplast genome will facilitate phylogenic, population and genetic engineering research investigations involving this particular species. PMID:27275817

  2. Transformation of Chloroplast Ribosomal RNA Genes in Chlamydomonas: Molecular and Genetic Characterization of Integration Events

    PubMed Central

    Newman, S. M.; Boynton, J. E.; Gillham, N. W.; Randolph-Anderson, B. L.; Johnson, A. M.; Harris, E. H.

    1990-01-01

    Transformation of chloroplast ribosomal RNA (rRNA) genes in Chlamydomonas has been achieved by the biolistic process using cloned chloroplast DNA fragments carrying mutations that confer antibiotic resistance. The sites of exchange employed during the integration of the donor DNA into the recipient genome have been localized using a combination of antibiotic resistance mutations in the 16S and 23S rRNA genes and restriction fragment length polymorphisms that flank these genes. Complete or nearly complete replacement of a region of the chloroplast genome in the recipient cell by the corresponding sequence from the donor plasmid was the most common integration event. Exchange events between the homologous donor and recipient sequences occurred preferentially near the vector:insert junctions. Insertion of the donor rRNA genes and flanking sequences into one inverted repeat of the recipient genome was followed by intramolecular copy correction so that both copies of the inverted repeat acquired identical sequences. Increased frequencies of rRNA gene transformants were achieved by reducing the copy number of the chloroplast genome in the recipient cells and by decreasing the heterology between donor and recipient DNA sequences flanking the selectable markers. In addition to producing bona fide chloroplast rRNA transformants, the biolistic process induced mutants resistant to low levels of streptomycin, typical of nuclear mutations in Chlamydomonas. PMID:1981764

  3. Arabidopsis Chloroplast RNA Binding Proteins CP31A and CP29A Associate with Large Transcript Pools and Confer Cold Stress Tolerance by Influencing Multiple Chloroplast RNA Processing Steps[W

    PubMed Central

    Kupsch, Christiane; Ruwe, Hannes; Gusewski, Sandra; Tillich, Michael; Small, Ian; Schmitz-Linneweber, Christian

    2012-01-01

    Chloroplast RNA metabolism is mediated by a multitude of nuclear encoded factors, many of which are highly specific for individual RNA processing events. In addition, a family of chloroplast ribonucleoproteins (cpRNPs) has been suspected to regulate larger sets of chloroplast transcripts. This together with their propensity for posttranslational modifications in response to external cues suggested a potential role of cpRNPs in the signal-dependent coregulation of chloroplast genes. We show here on a transcriptome-wide scale that the Arabidopsis thaliana cpRNPs CP31A and CP29A (for 31 kD and 29 kD chloroplast protein, respectively), associate with large, overlapping sets of chloroplast transcripts. We demonstrate that both proteins are essential for resistance of chloroplast development to cold stress. They are required to guarantee transcript stability of numerous mRNAs at low temperatures and under these conditions also support specific processing steps. Fine mapping of cpRNP–RNA interactions in vivo suggests multiple points of contact between these proteins and their RNA ligands. For CP31A, we demonstrate an essential function in stabilizing sense and antisense transcripts that span the border of the small single copy region and the inverted repeat of the chloroplast genome. CP31A associates with the common 3′-terminus of these RNAs and protects them against 3′-exonucleolytic activity. PMID:23110894

  4. Towards resolving Lamiales relationships: insights from rapidly evolving chloroplast sequences

    PubMed Central

    2010-01-01

    Background In the large angiosperm order Lamiales, a diverse array of highly specialized life strategies such as carnivory, parasitism, epiphytism, and desiccation tolerance occur, and some lineages possess drastically accelerated DNA substitutional rates or miniaturized genomes. However, understanding the evolution of these phenomena in the order, and clarifying borders of and relationships among lamialean families, has been hindered by largely unresolved trees in the past. Results Our analysis of the rapidly evolving trnK/matK, trnL-F and rps16 chloroplast regions enabled us to infer more precise phylogenetic hypotheses for the Lamiales. Relationships among the nine first-branching families in the Lamiales tree are now resolved with very strong support. Subsequent to Plocospermataceae, a clade consisting of Carlemanniaceae plus Oleaceae branches, followed by Tetrachondraceae and a newly inferred clade composed of Gesneriaceae plus Calceolariaceae, which is also supported by morphological characters. Plantaginaceae (incl. Gratioleae) and Scrophulariaceae are well separated in the backbone grade; Lamiaceae and Verbenaceae appear in distant clades, while the recently described Linderniaceae are confirmed to be monophyletic and in an isolated position. Conclusions Confidence about deep nodes of the Lamiales tree is an important step towards understanding the evolutionary diversification of a major clade of flowering plants. The degree of resolution obtained here now provides a first opportunity to discuss the evolution of morphological and biochemical traits in Lamiales. The multiple independent evolution of the carnivorous syndrome, once in Lentibulariaceae and a second time in Byblidaceae, is strongly supported by all analyses and topological tests. The evolution of selected morphological characters such as flower symmetry is discussed. The addition of further sequence data from introns and spacers holds promise to eventually obtain a fully resolved plastid tree of

  5. Partial characterization of cyclic electron transport in intact chloroplasts

    SciTech Connect

    Crowther, D.; Hind, G.

    1980-10-15

    Turnover of the cyclic electron transfer chain around photosystem I in intact chloroplasts was induced by addition of sodium dithionite after poisoning with 3-(3,4-dichlorophenyl)-1,1-dimethylurea. A substantial permeability barrier to dithionite allowed redox poising to a level sufficiently negative to active, but not overreduce, the cycle. Spectral changes could thus be studied without interference from photosystem II reactions. Illumination by repetitive single-turnover flashes showed the participation in the cycle of cytochromes f and b/sub 563/ with an apparent 1:1 stoichiometry. The rise of the flash-induced electrochromic bandshift showed a fast phase with rise time < 10 ..mu..s and a slow phase with rise time variable in the millisecond range. The slow phase had an amplitude equal to that of the fast phase and occurred only when electron transfer between cytochromes b/sub 563/ and f was uninhibited. A kinetic correlation was observed between the rise of the slow phase and the rereduction of cytochrome f, whereas cytochrome b/sub 563/ reoxidation was slower than both. Redox titrations of the appearance of the slow rise in the P518 response showed that it was only observed on repetitive flashes when a component of midpoint potential approx. = -55mV(pH 8.1), n = 2, was reduced before the flash. A comparison is drawn between this protonmotive electron transfer cycle and that of the purple nonsulfur bacterium Rhodopseudomonas capsulata; possible arrangements of electron carriers in the photosystem I cycle are discussed, and a modified Q cycle is proposed to account for the properties observed.

  6. Protomer Roles in Chloroplast Chaperonin Assembly and Function.

    PubMed

    Bai, Cuicui; Guo, Peng; Zhao, Qian; Lv, Zongyang; Zhang, Shijia; Gao, Fei; Gao, Liyan; Wang, Yingchun; Tian, Zhixi; Wang, Jifeng; Yang, Fuquan; Liu, Cuimin

    2015-10-01

    The individual roles of three chloroplast CPN60 protomers (CPN60α, CPN60β1, and CPN60β2) and whether and how they are assembled into functional chaperonin complexes are investigated in Chlamydomonas reinhardtii. Protein complexes containing all three potential subunits were identified in Chlamydomonas, and their co-expression in Escherichia coli yielded a homogeneous population of oligomers containing all three subunits (CPN60αβ1β2), with a molecular weight consistent with a tetradecameric structure. While homo-oligomers of CPN60β could form, they were dramatically reduced when CPN60α was present and homo-oligomers of CPN60β2 were readily changed into hetero-oligomers in the presence of ATP and other protomers. ATP hydrolysis caused CPN60 oligomers to disassemble and drove the purified protomers to reconstitute oligomers in vitro, suggesting that the dynamic nature of CPN60 oligomers is dependent on ATP. Only hetero-oligomeric CPN60αβ1β2, containing CPN60α, CPN60β1, and CPN60β2 subunits in a 5:6:3 ratio, cooperated functionally with GroES. The combination of CPN60α and CPN60β subunits, but not the individual subunits alone, complemented GroEL function in E. coli with subunit recognition specificity. Down-regulation of the CPN60α subunit in Chlamydomonas resulted in a slow growth defect and an inability to grow autotrophically, indicating the essential role of CPN60α in vivo. PMID:26057234

  7. Multimodal lightsheet, structured illumination and Airyscan superresolution microscopy of chloroplast size and its impact on light propagation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Altering chloroplast size changes the way light propagates through a leaf by altering light reflectance and transmission as well as absorption by chlorophyll. Thus changing chloroplast size can used to manipulate leaf optical properties to optimize photosynthetic efficiency with the ultimate goal of...

  8. Chloroplast-targeting protein expression in the oleaginous diatom Fistulifera solaris JPCC DA0580 toward metabolic engineering.

    PubMed

    Sunaga, Yoshihiko; Maeda, Yoshiaki; Yabuuchi, Takashi; Muto, Masaki; Yoshino, Tomoko; Tanaka, Tsuyoshi

    2015-01-01

    The chloroplast plays critical roles in lipid metabolism of microalgae, thus it is recognized as an attractive target of metabolic engineering to enhance biofuel production. It has been well known that recombinant protein expression in microalgal chloroplasts needs specific signal sequence which governs the transition manner of nuclear-encoded polypeptides within the subcellular compartments. However certain microalgae, including diatoms, have complex membrane systems surrounding the chloroplast, and thus chloroplast-targeting protein expression with the signal sequence has rarely been demonstrated except for a few model non-oleaginous diatoms. In this study, we performed recombinant green fluorescence protein (GFP) expression and transportation into the chloroplast of the oleaginous marine diatom, Fistulifera solaris JPCC DA0580. The signal sequence of ATP synthetase gamma subunit, which was predicted to localize in the chloroplast according to a bioinformatics analysis pipeline, was employed as a key factor of this technique. As a result, specific localization of GFP in the chloroplast was observed. It would be useful to engineer the lipid synthesis pathways existing in the chloroplast. Furthermore, intensive gathering of GFP in the rod-like structure was also detected, which has not been observed in model diatom studies. As comparing with electron microscopic observation, the structure was estimated to be a pyrenoid. PMID:25043335

  9. Specific and Efficient Targeting of Cyanobacterial Bicarbonate Transporters to the Inner Envelope Membrane of Chloroplasts in Arabidopsis

    PubMed Central

    Uehara, Susumu; Adachi, Fumi; Ito-Inaba, Yasuko; Inaba, Takehito

    2016-01-01

    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. PMID:26870048

  10. The Arabidopsis YELLOW STRIPE LIKE4 and 6 Transporters Control Iron Release from the Chloroplast[C][W

    PubMed Central

    Divol, Fanchon; Couch, Daniel; Conéjéro, Geneviève; Roschzttardtz, Hannetz; Mari, Stéphane; Curie, Catherine

    2013-01-01

    In most plant cell types, the chloroplast represents the largest sink for iron, which is both essential for chloroplast metabolism and prone to cause oxidative damage. Here, we show that to buffer the potentially harmful effects of iron, besides ferritins for storage, the chloroplast is equipped with specific iron transporters that respond to iron toxicity by removing iron from the chloroplast. We describe two transporters of the YELLOW STRIPE1-LIKE family from Arabidopsis thaliana, YSL4 and YSL6, which are likely to fulfill this function. Knocking out both YSL4 and YSL6 greatly reduces the plant’s ability to cope with excess iron. Biochemical and immunolocalization analyses showed that YSL6 resides in the chloroplast envelope. Elemental analysis and histochemical staining indicate that iron is trapped in the chloroplasts of the ysl4 ysl6 double mutants, which also accumulate ferritins. Also, vacuolar iron remobilization and NRAMP3/4 expression are inhibited. Furthermore, ubiquitous expression of YSL4 or YSL6 dramatically reduces plant tolerance to iron deficiency and decreases chloroplastic iron content. These data demonstrate a fundamental role for YSL4 and YSL6 in managing chloroplastic iron. YSL4 and YSL6 expression patterns support their physiological role in detoxifying iron during plastid dedifferentiation occurring in embryogenesis and senescence. PMID:23512854

  11. Sequencing Cucumber (Cucumis Sativus L.) Chloroplast Genomes Identifies Differences Between Chilling-Tolerant and-Susceptible Cucumber Lines

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Complete sequencing of cucumber chloroplast (cp)DNA was facilitated by the development of 414 consensus chloroplast sequencing primers (CCSPs) from conserved cpDNA sequences of Arabidopsis (Arabidopsis thaliana L.), spinach (Spinacia oleracea L.), and tobacco (Nicotiana tabacum L.) cpDNAs, using deg...

  12. GENETIC POLYMORPHISM IN GYMNODINIUM GALATHEANUM CHLOROPLAST DNA SEQUENCES AND DEVELOPMENT OF A MOLECULAR DETECTION ASSAY. (R827084)

    EPA Science Inventory

    Nuclear and chloroplast-encoded small subunit ribosomal DNA sequences were obtained
    from several strains of the toxic dinoflagellate Gymnodinium galatheanum. Phylogenetic analyses and
    comparison of sequences indicate that the chloroplast sequences show a higher degree of se...

  13. REDUCTION OF SPECIES IN THE WILD POTATO SOLANUM SECTION PETOTA SERIES LONGIPEDICELLATA: AFLP, RAPD AND CHLOROPLAST SSR DATA.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Species boundaries were assessed with three molecular markers (AFLPs, RAPDs, chloroplast microsatellites, also known as chloroplast single sequence repeats [cpSSRs]) for all six species of wild potatoes (Solanum L. section Petota Dumort.) assigned to ser. Longipedicellata: S. fendleri, S. hjertingii...

  14. Hybridization and Introgression in a Sympatric Pair of Oak Species (Fagaceae) Studied with Morphology and Chloroplast and Nuclear SSR Markers

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Chloroplast and nuclear SSR markers and morphology were used to study hybridization and introgression between Quercus douglasii and Q. lobata, two sympatric oak species native to California. Chloroplast SSR markers provided strong evidence of gene exchange via ongoing introgression between these sp...

  15. Induction events and short-term regulation of electron transport in chloroplasts: an overview.

    PubMed

    Tikhonov, Alexander N

    2015-08-01

    Regulation of photosynthetic electron transport at different levels of structural and functional organization of photosynthetic apparatus provides efficient performance of oxygenic photosynthesis in plants. This review begins with a brief overview of the chloroplast electron transport chain. Then two noninvasive biophysical methods (measurements of slow induction of chlorophyll a fluorescence and EPR signals of oxidized P700 centers) are exemplified to illustrate the possibility of monitoring induction events in chloroplasts in vivo and in situ. Induction events in chloroplasts are considered and briefly discussed in the context of short-term mechanisms of the following regulatory processes: (i) pH-dependent control of the intersystem electron transport; (ii) the light-induced activation of the Calvin-Benson cycle; (iii) optimization of electron transport due to fitting alternative pathways of electron flow and partitioning light energy between photosystems I and II; and (iv) the light-induced remodeling of photosynthetic apparatus and thylakoid membranes. PMID:25680580

  16. Changing kinetic properties of glucose-6-phosphate dehydrogenase from pea chloroplasts during photosynthetic induction

    SciTech Connect

    Yuan, X.; Anderson, L.E.

    1987-04-01

    The first enzyme of the oxidative pentose phosphate pathway, glucose-6-P dehydrogenase (EC 1.1.1.49), is inactivated when pea chloroplasts are irradiated. They have examined the kinetics of light inactivation of glucose-6-P dehydrogenase in intact chloroplasts during photosynthetic induction and the kinetic parameters of the active (dark) and less active (light) form of the dehydrogenase. Light inactivation of the dehydrogenase is rapid and occurs before photosynthetic O/sub 2/ evolution is measureable in intact chloroplasts. Likewise dark activation is quite rapid. The major change in the kinetic parameters of glucose-6-phosphate dehydrogenase is in maximal velocity. This light inactivation probably prevents operation of a futile cycle involving glucose-6-P, NADPH and oxidative and reductive pentose phosphate pathway enzymes.

  17. PEA chloroplasts under clino-rotation: lipid peroxidation and superoxide dismutase activity

    NASA Astrophysics Data System (ADS)

    Baranenko, V. V.

    The lipid peroxidation (LP) intensity and the activity of the antioxidant enzyme superoxide dismutase (SOD) were studied in chloroplasts of pea (Pisum sativum L.) plants grown for 7 and 14 days under clino-rotation. An increase in LP levels in chloroplasts during both terms of clinorotation in comparison with stationary controls was documented. SOD activity increased in chloroplasts of plants that were clino-rotated for seven days. SOD has a significant protective effect by diminishing the availability of O2-. However, under more prolonged clino-rotation (14 days), SOD activity decreased but was still higher than in the control samples. In accordance with Selye's oxidative stress theory (Selye, 1956; modified by Leshem et al., 1998), plants that were clino-rotated for seven days are presumed to be in a stage of resistance while 14-day plants reached a stage of exhaustion.

  18. Light harvesting and chloroplast electron transport in NADP-malic enzyme type C4 plants.

    PubMed

    Nakajima Munekage, Yuri

    2016-06-01

    The structure of thylakoids in chloroplasts and the organization of the electron transport chain changed dynamically during the evolution of C4 photosynthesis, especially in the nicotinamide adenine dinucleotide phosphate (NADP)-malic enzyme type C4 species. Stacked grana membranes are strongly reduced in the bundle sheath chloroplasts of these plants, where photosystem II activity is diminished and cyclic electron transport around photosystem I mainly occurs. This change optimizes the ATP/NADPH production ratio in bundle sheath chloroplasts to drive the metabolic cycle of C4 photosynthesis. This review summarizes the current model of light harvesting and electron transport in the NADP-malic enzyme type C4 plants and discusses how it changed during the evolution of C4 photosynthesis. PMID:26999307

  19. The complete chloroplast genome sequence of Lilium hansonii Leichtlin ex D.D.T.Moore.

    PubMed

    Kim, Kyunghee; Hwang, Yoon-Jung; Lee, Sang-Choon; Yang, Tae-Jin; Lim, Ki-Byung

    2016-09-01

    Lilium hansonii is a lily species native to Korea and an important wild species for lily breeding. The chloroplast genome of L. hansonii was completed by de novo assembly using the small amount of whole genome sequencing data. The chloroplast genome of L. hansonii was 152 655 bp long and consisted of large single copy region (82 051 bp), small single copy region (17 620 bp) and a pair of inverted repeat regions (26 492 bp). A total of 115 genes were annotated, which included 81 protein-coding genes, 30 tRNA genes and 4 rRNA genes. Phylogenetic analysis with the reported chloroplast genomes revealed that L. hansonii is most closely related to L. superbum (Turk's-cap lily) and L. longiflorum (Easter lily). PMID:26404645

  20. Expression and Chloroplast Targeting of Cholesterol Oxidase in Transgenic Tobacco Plants

    PubMed Central

    Corbin, David R.; Grebenok, Robert J.; Ohnmeiss, Thomas E.; Greenplate, John T.; Purcell, John P.

    2001-01-01

    Cholesterol oxidase represents a novel type of insecticidal protein with potent activity against the cotton boll weevil (Anthonomus grandis grandis Boheman). We transformed tobacco (Nicotiana tabacum) plants with the cholesterol oxidase choM gene and expressed cytosolic and chloroplast-targeted versions of the ChoM protein. Transgenic leaf tissues expressing cholesterol oxidase exerted insecticidal activity against boll weevil larvae. Our results indicate that cholesterol oxidase can metabolize phytosterols in vivo when produced cytosolically or when targeted to chloroplasts. The transgenic plants exhibiting cytosolic expression accumulated low levels of saturated sterols known as stanols, and displayed severe developmental aberrations. In contrast, the transgenic plants expressing chloroplast-targeted cholesterol oxidase maintained a greater accumulation of stanols, and appeared phenotypically and developmentally normal. These results are discussed within the context of plant sterol distribution and metabolism. PMID:11457962

  1. Measurement of Nitric Oxide (NO) Generation Rate by Chloroplasts Employing Electron Spin Resonance (ESR).

    PubMed

    Galatro, Andrea; Puntarulo, Susana

    2016-01-01

    Chloroplasts are among the more active organelles involved in free energy transduction in plants (photophosphorylation). Nitric oxide (NO) generation by soybean (Glycine max, var ADM 4800) chloroplasts was measured as an endogenous product assessed by electron paramagnetic resonance (ESR) spin-trapping technique. ESR spectroscopy is a methodology employed to detect species with unpaired electrons (paramagnetic). This technology has been successfully applied to different plant tissues and subcellular compartments to asses both, NO content and generation. The spin trap MGD-Fe(2+) is extensively employed to efficiently detect NO. Here, we describe a simple methodology to asses NO generation rate by isolated chloroplasts in the presence of either L-Arginine or nitrite (NO2 (-)) as substrates, since these compounds are required for enzymatic activities considered as the possible sources of NO generation in plants. PMID:27094414

  2. The complete chloroplast genome sequence of the medicinal plant Rheum palmatum L. (Polygonaceae).

    PubMed

    Fan, Kai; Sun, Xiao-Jie; Huang, Min; Wang, Xu-Mei

    2016-07-01

    The complete chloroplast genome of the medicinal plant Rheum palmatum L. (Polygonaceae) has been reconstructed from the whole-genome Illumina sequencing data. The genome is 161 541 bp in length, and exhibits a typical quadripartite structure of the large (LSC, 86 518 bp) and small (SSC, 13 111 bp) single-copy regions, separated by a pair of inverted repeats (IRs, 30 956 bp each). The chloroplast genome contains 131 genes, including 84 protein-coding genes (78 PCG species), eight ribosomal RNA genes (four rRNA species) and 37 transfer RNA genes (28 tRNA species). Phylogenetic tree based on the maximum parsimony (MP) analysis of 65 chloroplast protein-coding genes for 13 taxa demonstrated a close relationship between R. palmatum and Fagopyrum esculentum subsp. ancestrale in Polygonaceae. PMID:26153751

  3. Identification of the 64 kilodalton chloroplast stromal phosphoprotein as phosphoglucomutase. [Pisum sativum

    SciTech Connect

    Salvucci, M.E.; Drake, R.R.; Broadbent, K.P.; Haley, B.E. ); Hanson, K.R.; McHale, N.A. )

    1990-05-01

    Phosphorylation of the 64 kilodalton stromal phosphoprotein by incubation of pea (Pisum sativum) chloroplast extracts with ({gamma}-{sup 32}P)ATP decreased in the presence of Glc-6-P and Glc-1,6-P{sub 2}, but was stimulated by glucose. Two-dimensional gel electrophoresis following incubation of intact chloroplasts and stromal extracts with ({gamma}-{sup 32}P)ATP, or incubation of stromal extracts and partially purified phosphoglucomutase (EC 2.7.5.1) with ({sup 32}P)Glc-1-P showed that the identical 64 kilodalton polypeptide was labeled. A 62 kilodalton polypeptide was phosphorylated by incubation of tobacco (Nicotiana sylvestris) stromal extracts with either ({gamma}-{sup 32}P)ATP or ({sup 32}P)Glc-1-P. In contrast, an analogous polypeptide was not phosphorylated in extracts from a tobacco mutant deficient in plastid phosphoglucomutase activity. The results indicate that the 64 (or 62) kilodalton chloroplast stromal phosphoprotein is phosphoglucomutase.

  4. The complete sequence of the chloroplast genome of the green microalga Lobosphaera (Parietochloris) incisa.

    PubMed

    Tourasse, Nicolas J; Barbi, Tommaso; Waterhouse, Janet C; Shtaida, Nastassia; Leu, Stefan; Boussiba, Sammy; Purton, Saul; Vallon, Olivier

    2016-05-01

    We hereby report the complete chloroplast genome sequence of the green unicellular alga Lobosphaera (Parietochloris) incisa (strain SAG 2468). The genome consists of a circular chromosome of 156,028 bp, which is 72% A-T rich and does not contain a large rRNA-encoding inverted repeat. It is predicted to encode a total of 111 genes including 78 protein-coding, three rRNA, and 30 tRNA genes. The genome sequence also carries a self-splicing group I intron and a group II intron remnant. Overall, the gene and intron content of the L. incisa chloroplast genome is highly similar to that of other species of Trebouxiophyceae. In contrast, the L. incisa chloroplast genome harbors 88 copies of various intergenic dispersed DNA repeat sequences that are all unique to L. incisa. PMID:25423517

  5. [Disturbed thylakoid-formation in chloroplasts of a xantha-mutant of Arabidopsis thaliana (L.) HEYNH].

    PubMed

    Röbbeilen, G

    1966-03-01

    A monohybrid recessive xantha-mutant of Arabidopsis thaliana was grown aseptically on mineral agar +2% sucrose up to maturity. The cotyledons and rosette leaves were non-green during all developmental stages, their yellow colour bleaching with age.Palisade cells of various leaves of mutant plants were investigated light- and electron microscopically. All of the organelles of these cells, except the plastids, had a normal appearance. Special attention was paid to the existence of "dense bodies", and their relationship to sphaerosomes and lysosomes was outlined.In the mutant plastids the typical thylakoid differentiation present in the chloroplasts of the green Arabidopsis plant is entirely lacking. Within the palisade cells the plastids are normal in number and size. Under the light microscope they appear as hyaline vesicles with usually two "primary grana". Their outline is characterized by numerous invaginations and protrusions; however, it is still uncertain to what extent this "amoeboidy" is only an artifact.The specific gene dependent block in the plastid development is apparently effective during the conversion of tubules of the prolamellar body to thylakoids. From the persistent "prolamellar bodies" in the mutant only single abnormal thylakoids originate. These often have a deformed cup- or ventricle-like shape and occasionally are piled up as magnograna. In a characteristic manner tubules and fibrils are closely associated in parallel rows with the outsides of the thylakoids. The fibrils having a mean diameter of 75 Å are connected with the tubules which are derived from the "prolamellar body". From this striking aggregation the possibility of a laminal fusion of tubules as a process of normal thylakoid growth is deduced.By pigment analysis of mutant leaves no chlorophyll but only carotenoids were detected. However, many osmiophilic globules as well as large bodies of unknown composition (proteinaceous?) appear in the plastid matrix. Therefore the question

  6. Modified Clp Protease Complex in the ClpP3 Null Mutant and Consequences for Chloroplast Development and Function in Arabidopsis1[C][W][OA

    PubMed Central

    Kim, Jitae; Olinares, Paul Dominic; Oh, Soo-hyun; Ghisaura, Stefania; Poliakov, Anton; Ponnala, Lalit; van Wijk, Klaas J.

    2013-01-01

    The plastid ClpPRT protease consists of two heptameric rings of ClpP1/ClpR1/ClpR2/ClpR3/ClpR4 (the R-ring) and ClpP3/ClpP4/ClpP5/ClpP6 (the P-ring) and peripherally associated ClpT1/ClpT2 subunits. Here, we address the contributions of ClpP3 and ClpP4 to ClpPRT core organization and function in Arabidopsis (Arabidopsis thaliana). ClpP4 is strictly required for embryogenesis, similar to ClpP5. In contrast, loss of ClpP3 (clpp3-1) leads to arrest at the hypocotyl stage; this developmental arrest can be removed by supplementation with sucrose or glucose. Heterotrophically grown clpp3-1 can be transferred to soil and generate viable seed, which is surprising, since we previously showed that CLPR2 and CLPR4 null alleles are always sterile and die on soil. Based on native gels and mass spectrometry-based quantification, we show that despite the loss of ClpP3, modified ClpPR core(s) could be formed, albeit at strongly reduced levels. A large portion of ClpPR subunits accumulated in heptameric rings, with overaccumulation of ClpP1/ClpP5/ClpP6 and ClpR3. Remarkably, the association of ClpT1 to the modified Clp core was unchanged. Large-scale quantitative proteomics assays of clpp3-1 showed a 50% loss of photosynthetic capacity and the up-regulation of plastoglobules and all chloroplast stromal chaperone systems. Specific chloroplast proteases were significantly up-regulated, whereas the major thylakoid protease (FtsH1/FtsH2/FtsH5/FtsH8) was clearly unchanged, indicating a controlled protease network response. clpp3-1 showed a systematic decrease of chloroplast-encoded proteins that are part of the photosynthetic apparatus but not of chloroplast-encoded proteins with other functions. Candidate substrates and an explanation for the differential phenotypes between the CLPP3, CLPP4, and CLPP5 null mutants are discussed. PMID:23548781

  7. A J-Like Protein Influences Fatty Acid Composition of Chloroplast Lipids in Arabidopsis

    PubMed Central

    Ajjawi, Imad; Coku, Ardian; Froehlich, John E.; Yang, Yue; Osteryoung, Katherine W.; Benning, Christoph; Last, Robert L.

    2011-01-01

    A comprehensive understanding of the lipid and fatty acid metabolic machinery is needed for optimizing production of oils and fatty acids for fuel, industrial feedstocks and nutritional improvement in plants. T-DNA mutants in the poorly annotated Arabidopsis thaliana gene At1g08640 were identified as containing moderately high levels (50–100%) of 16∶1Δ7 and 18∶1Δ9 leaf fatty acids and subtle decreases (5–30%) of 16∶3 and 18∶3 (http://www.plastid.msu.edu/). TLC separation of fatty acids in the leaf polar lipids revealed that the chloroplastic galactolipids monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG) were the main lipid types affected by this mutation. Analysis of the inferred amino acid sequence of At1g08640 predicted the presence of a transit peptide, three transmembrane domains and an N-terminal J-like domain, and the gene was named CJD1 for Chloroplast J-like Domain 1. GFP reporter experiments and in vitro chloroplast import assays demonstrated CJD1 is a chloroplast membrane protein. Screening of an Arabidopsis cDNA library by yeast-2-hybrid (Y2H) using the J-like domain of CJD1 as bait identified a plastidial inner envelope protein (Accumulation and Replication of Chloroplasts 6, ARC6) as the primary interacting partner in the Y2H assay. ARC6 plays a central role in chloroplast division and binds CJD1 via its own J-like domain along with an adjacent conserved region whose function is not fully known. These results provide a starting point for future investigations of how mutations in CJD1 affect lipid composition. PMID:22028775

  8. Slugs' last meals: molecular identification of sequestered chloroplasts from different algal origins in Sacoglossa (Opisthobranchia, Gastropoda).

    PubMed

    Händeler, Katharina; Wägele, Heike; Wahrmund, Ute; Rüdinger, Mareike; Knoop, Volker

    2010-11-01

    Some sacoglossan sea slugs have become famous for their unique capability to extract and incorporate functional chloroplasts from algal food organisms (mainly Ulvophyceae) into their gut cells. The functional incorporation of the so-called kleptoplasts allows the slugs to rely on photosynthetic products for weeks to months, enabling them to survive long periods of food shortage over most of their life-span. The algal food spectrum providing kleptoplasts as temporary, non-inherited endosymbionts appears to vary among sacoglossan slugs, but detailed knowledge is sketchy or unavailable. Accurate identification of algal donor species, which provide the chloroplasts for long-term retention is of primary importance to elucidate the biochemical mechanisms allowing long-term functionality of the captured chloroplast in the foreign animal cell environment. Whereas some sacoglossans forage on a variety of algal species, (e.g. Elysia crispata and E. viridis) others are more selective. Hence, characterizing the range of functional sacoglossan-chloroplast associations in nature is a prerequisite to understand the basis of this enigmatic endosymbiosis. Here, we present a suitable chloroplast gene (tufA) as a marker, which allows identification of the respective algal kleptoplast donor taxa by analysing DNA from whole animals. This novel approach allows identification of donor algae on genus or even species level, thus providing evidence for the taxonomic range of food organisms. We report molecular evidence that chloroplasts from different algal sources are simultaneously incorporated in some species of Elysia. NeigborNet analyses for species assignments are preferred over tree reconstruction methods because the former allow more reliable statements on species identification via barcoding, or rather visualize alternative allocations not to be seen in the latter. PMID:21565106

  9. The Complete Chloroplast Genome of the Hare’s Ear Root, Bupleurum falcatum: Its Molecular Features

    PubMed Central

    Shin, Dong-Ho; Lee, Jeong-Hoon; Kang, Sang-Ho; Ahn, Byung-Ohg; Kim, Chang-Kug

    2016-01-01

    Bupleurum falcatum, which belongs to the family Apiaceae, has long been applied for curative treatments, especially as a liver tonic, in herbal medicine. The chloroplast (cp) genome has been an ideal model to perform the evolutionary and comparative studies because of its highly conserved features and simple structure. The Apiaceae family is taxonomically close to the Araliaceae family and there have been numerous complete chloroplast genome sequences reported in the Araliaceae family, while little is known about the Apiaceae family. In this study, the complete sequence of the B. falcatum chloroplast genome was obtained. The full-length of the cp genome is 155,989 nucleotides with a 37.66% overall guanine-cytosine (GC) content and shows a quadripartite structure composed of three nomenclatural regions: a large single-copy (LSC) region, a small single-copy (SSC) region, and a pair of inverted repeat (IR) regions. The genome occupancy is 85,912-bp, 17,517-bp, and 26,280-bp for LSC, SSC, and IR, respectively. B. falcatum was shown to contain 111 unique genes (78 for protein-coding, 29 for tRNAs, and four for rRNAs, respectively) on its chloroplast genome. Genic comparison found that B. falcatum has no pseudogenes and has two gene losses, accD in the LSC and ycf15 in the IRs. A total of 55 unique tandem repeat sequences were detected in the B. falcatum cp genome. This report is the first to describe the complete chloroplast genome sequence in B. falcatum and will open up further avenues of research to understand the evolutionary panorama and the chloroplast genome conformation in related plant species. PMID:27187480

  10. Comparison of Transcriptional Changes to Chloroplast and Mitochondrial Perturbations Reveals Common and Specific Responses in Arabidopsis

    PubMed Central

    Van Aken, Olivier; Whelan, James

    2012-01-01

    Throughout the life of a plant, the biogenesis and fine-tuning of energy organelles is essential both under normal growth and stress conditions. Communication from organelle to nucleus is essential to adapt gene regulation and protein synthesis specifically to the current needs of the plant. This organelle-to-nuclear communication is termed retrograde signaling and has been studied extensively over the last decades. In this study we have used large-scale gene expression data sets relating to perturbations of chloroplast and mitochondrial function to gain further insights into plant retrograde signaling and how mitochondrial and chloroplast retrograde pathways interact and differ. Twenty seven studies were included that assess transcript profiles in response to chemical inhibition as well as genetic mutations of organellar proteins. The results show a highly significant overlap between gene expression changes triggered by chloroplast and mitochondrial perturbations. These overlapping gene expression changes appear to be common with general abiotic, biotic, and nutrient stresses. However, retrograde signaling pathways are capable of distinguishing the source of the perturbation as indicated by a statistical overrepresentation of changes in genes encoding proteins of the affected organelle. Organelle-specific overrepresented functional categories among others relate to energy metabolism and protein synthesis. Our analysis also suggests that WRKY transcription factors play a coordinating role on the interface of both organellar signaling pathways. Global comparison of the expression profiles for each experiment revealed that the recently identified chloroplast retrograde pathway using phospho-adenosine phosphate is possibly more related to mitochondrial than chloroplast perturbations. Furthermore, new marker genes have been identified that respond specifically to mitochondrial and/or chloroplast dysfunction. PMID:23269925

  11. Albino Leaf1 That Encodes the Sole Octotricopeptide Repeat Protein Is Responsible for Chloroplast Development.

    PubMed

    Zhang, Zemin; Tan, Jianjie; Shi, Zhenying; Xie, Qingjun; Xing, Yi; Liu, Changhong; Chen, Qiaoling; Zhu, Haitao; Wang, Jiang; Zhang, Jingliu; Zhang, Guiquan

    2016-06-01

    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

  12. The Complete Chloroplast Genome of the Hare's Ear Root, Bupleurum falcatum: Its Molecular Features.

    PubMed

    Shin, Dong-Ho; Lee, Jeong-Hoon; Kang, Sang-Ho; Ahn, Byung-Ohg; Kim, Chang-Kug

    2016-01-01

    Bupleurum falcatum, which belongs to the family Apiaceae, has long been applied for curative treatments, especially as a liver tonic, in herbal medicine. The chloroplast (cp) genome has been an ideal model to perform the evolutionary and comparative studies because of its highly conserved features and simple structure. The Apiaceae family is taxonomically close to the Araliaceae family and there have been numerous complete chloroplast genome sequences reported in the Araliaceae family, while little is known about the Apiaceae family. In this study, the complete sequence of the B. falcatum chloroplast genome was obtained. The full-length of the cp genome is 155,989 nucleotides with a 37.66% overall guanine-cytosine (GC) content and shows a quadripartite structure composed of three nomenclatural regions: a large single-copy (LSC) region, a small single-copy (SSC) region, and a pair of inverted repeat (IR) regions. The genome occupancy is 85,912-bp, 17,517-bp, and 26,280-bp for LSC, SSC, and IR, respectively. B. falcatum was shown to contain 111 unique genes (78 for protein-coding, 29 for tRNAs, and four for rRNAs, respectively) on its chloroplast genome. Genic comparison found that B. falcatum has no pseudogenes and has two gene losses, accD in the LSC and ycf15 in the IRs. A total of 55 unique tandem repeat sequences were detected in the B. falcatum cp genome. This report is the first to describe the complete chloroplast genome sequence in B. falcatum and will open up further avenues of research to understand the evolutionary panorama and the chloroplast genome conformation in related plant species. PMID:27187480

  13. The first complete chloroplast genome sequence of a lycophyte, Huperzia lucidula (Lycopodiaceae).

    PubMed

    Wolf, Paul G; Karol, Kenneth G; Mandoli, Dina F; Kuehl, Jennifer; Arumuganathan, K; Ellis, Mark W; Mishler, Brent D; Kelch, Dean G; Olmstead, Richard G; Boore, Jeffrey L

    2005-05-01

    We used a unique combination of techniques to sequence the first complete chloroplast genome of a lycophyte, Huperzia lucidula. This plant belongs to a significant clade hypothesized to represent the sister group to all other vascular plants. We used fluorescence-activated cell sorting (FACS) to isolate the organelles, rolling circle amplification (RCA) to amplify the genome, and shotgun sequencing to 8x depth coverage to obtain the complete chloroplast genome sequence. The genome is 154,373 bp, containing inverted repeats of 15,314 bp each, a large single-copy region of 104,088 bp, and a small single-copy region of 19,657 bp. Gene order is more similar to those of mosses, liverworts, and hornworts than to gene order for other vascular plants. For example, the Huperzia chloroplast genome possesses the bryophyte gene order for a previously characterized 30 kb inversion, thus supporting the hypothesis that lycophytes are sister to all other extant vascular plants. The lycophyte chloroplast genome data also enable a better reconstruction of the basal tracheophyte genome, which is useful for inferring relationships among bryophyte lineages. Several unique characters are observed in Huperzia, such as movement of the gene ndhF from the small single copy region into the inverted repeat. We present several analyses of evolutionary relationships among land plants by using nucleotide data, inferred amino acid sequences, and by comparing gene arrangements from chloroplast genomes. The results, while still tentative pending the large number of chloroplast genomes from other key lineages that are soon to be sequenced, are intriguing in themselves, and contribute to a growing comparative database of genomic and morphological data across the green plants. PMID:15788152

  14. Chloroplast Iron Transport Proteins – Function and Impact on Plant Physiology

    PubMed Central

    López-Millán, Ana F.; Duy, Daniela; Philippar, Katrin

    2016-01-01

    Chloroplasts originated about three billion years ago by endosymbiosis of an ancestor of today’s cyanobacteria with a mitochondria-containing host cell. During evolution chloroplasts of higher plants established as the site for photosynthesis and thus became the basis for all life dependent on oxygen and carbohydrate supply. To fulfill this task, plastid organelles are loaded with the transition metals iron, copper, and manganese, which due to their redox properties are essential for photosynthetic electron transport. In consequence, chloroplasts for example represent the iron-richest system in plant cells. However, improvement of oxygenic photosynthesis in turn required adaptation of metal transport and homeostasis since metal-catalyzed generation of reactive oxygen species (ROS) causes oxidative damage. This is most acute in chloroplasts, where radicals and transition metals are side by side and ROS-production is a usual feature of photosynthetic electron transport. Thus, on the one hand when bound by proteins, chloroplast-intrinsic metals are a prerequisite for photoautotrophic life, but on the other hand become toxic when present in their highly reactive, radical generating, free ionic forms. In consequence, transport, storage and cofactor-assembly of metal ions in plastids have to be tightly controlled and are crucial throughout plant growth and development. In the recent years, proteins for iron transport have been isolated from chloroplast envelope membranes. Here, we discuss their putative functions and impact on cellular metal homeostasis as well as photosynthetic performance and plant metabolism. We further consider the potential of proteomic analyses to identify new players in the field. PMID:27014281

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

    PubMed

    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. PMID:26031839

  16. Light modulation of glyceraldehyde-3-phosphate dehydrogenase and glucose-6-phosphate dehydrogenase by photosynthetic electron flow in pea chloroplasts

    SciTech Connect

    Akamba, L.M.; Anderson, L.E.

    1981-02-01

    Light activation of NADP-linked glyceraldehyde-3-P dehydrogenase (EC 1.2.1.13) and light inactivation of glucose-6-P dehydrogenase (EC 1.1.1.49) appear to be modulated within pea leaf chloroplasts by mediators which are reduced by photosynthetic electron flow from the photosystem I reaction center. Dichlorophenyl-1,1-dimethylurea inhibition of this modulation can be completely reversed by ascorbate plus 2,6-dichlorophenolindophenol in broken chloroplasts, but not in intact chloroplasts. Intact chloroplasts are impermeable to 2,6-dichlorophenolindophenol at pH 7.5. Studies on the effect of light in reconstituted chloroplasts with photosystem I-enriched particles in the place of whole thylakoids revealed that photosystem I participants in the light modulation of NADP-linked glyceraldehyde-3-P dehydrogenase and of glucose-6-P dehydrogenase.

  17. Endosymbiotic origin and codon bias of the nuclear gene for chloroplast glyceraldehyde-3-phosphate dehydrogenase from maize.

    PubMed

    Brinkmann, H; Martinez, P; Quigley, F; Martin, W; Cerff, R

    1987-01-01

    The nuclei of plant cells harbor genes for two types of glyceraldehyde-3-phosphate dehydrogenases (GAPDH) displaying a sequence divergence corresponding to the prokaryote/eukaryote separation. This strongly supports the endosymbiotic theory of chloroplast evolution and in particular the gene transfer hypothesis suggesting that the gene for the chloroplast enzyme, initially located in the genome of the endosymbiotic chloroplast progenitor, was transferred during the course of evolution into the nuclear genome of the endosymbiotic host. Codon usage in the gene for chloroplast GAPDH of maize is radically different from that employed by present-day chloroplasts and from that of the cytosolic (glycolytic) enzyme from the same cell. This reveals the presence of subcellular selective pressures which appear to be involved in the optimization of gene expression in the economically important graminaceous monocots. PMID:3131533

  18. Phylogeography of Camellia taliensis (Theaceae) inferred from chloroplast and nuclear DNA: insights into evolutionary history and conservation

    PubMed Central

    2012-01-01

    Background As one of the most important but seriously endangered wild relatives of the cultivated tea, Camellia taliensis harbors valuable gene resources for tea tree improvement in the future. The knowledge of genetic variation and population structure may provide insights into evolutionary history and germplasm conservation of the species. Results Here, we sampled 21 natural populations from the species' range in China and performed the phylogeography of C. taliensis by using the nuclear PAL gene fragment and chloroplast rpl32-trnL intergenic spacer. Levels of haplotype diversity and nucleotide diversity detected at rpl32-trnL (h = 0.841; π = 0.00314) were almost as high as at PAL (h = 0.836; π = 0.00417). Significant chloroplast DNA population subdivision was detected (GST = 0.988; NST = 0.989), suggesting fairly high genetic differentiation and low levels of recurrent gene flow through seeds among populations. Nested clade phylogeographic analysis of chlorotypes suggests that population genetic structure in C. taliensis has been affected by habitat fragmentation in the past. However, the detection of a moderate nrDNA population subdivision (GST = 0.222; NST = 0.301) provided the evidence of efficient pollen-mediated gene flow among populations and significant phylogeographical structure (NST > GST; P < 0.01). The analysis of PAL haplotypes indicates that phylogeographical pattern of nrDNA haplotypes might be caused by restricted gene flow with isolation by distance, which was also supported by Mantel’s test of nrDNA haplotypes (r = 0.234, P < 0.001). We found that chlorotype C1 was fixed in seven populations of Lancang River Region, implying that the Lancang River might have provided a corridor for the long-distance dispersal of the species. Conclusions We found that C. taliensis showed fairly high genetic differentiation resulting from restricted gene flow and habitat fragmentation. This phylogeographical

  19. The complete chloroplast genome sequence of Tetrastigma hemsleyanum Diels at Gilg.

    PubMed

    Li, Mengzhu; Chen, Qinyi; Yang, Bingxian; Ma, Ji; Li, Baoguo; Zhang, Lin

    2016-09-01

    The complete chloroplast genome sequence of Tetrastigma hemsleyanum Diels at Gilg, a critical Chinese medicine, is reported here. The complete chloroplast genome of Tetrastigma hemsleyanum Diels at Gilg is 159 914 bp in length with 37.55% overall GC content. A pair of IRs (inverted repeats) of 26 510 bp were separated by LSC (87 927 bp) and SSC (18 967 bp). The phylogenetic analysis of 40 taxa showed a strong sister relationship with all other rosids. However, the placement of Myrtales still needs further verification. PMID:26329851

  20. Multiple regulatory mechanisms in the chloroplast of green algae: relation to hydrogen production.

    PubMed

    Antal, Taras K; Krendeleva, Tatyana E; Tyystjärvi, Esa

    2015-09-01

    A complex regulatory network in the chloroplast of green algae provides an efficient tool for maintenance of energy and redox balance in the cell under aerobic and anaerobic conditions. In this review, we discuss the structural and functional organizations of electron transport pathways in the chloroplast, and regulation of photosynthesis in the green microalga Chlamydomonas reinhardtii. The focus is on the regulatory mechanisms induced in response to nutrient deficiency stress and anoxia and especially on the role of a hydrogenase-mediated reaction in adaptation to highly reducing conditions and ATP deficiency in the cell. PMID:25986411