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Sample records for chloroplast trnl uaa

  1. Chloroplast DNA analysis of Tunisian cork oak populations (Quercus suber L.): sequence variations and molecular evolution of the trnL (UAA)-trnF (GAA) region.

    Science.gov (United States)

    Abdessamad, A; Baraket, G; Sakka, H; Ammari, Y; Ksontini, M; Hannachi, A Salhi

    2016-10-24

    Sequences of the trnL-trnF spacer and combined trnL-trnF region in chloroplast DNA of cork oak (Quercus suber L.) were analyzed to detect polymorphisms and to elucidate molecular evolution and demographic history. The aligned sequences varied in length and nucleotide composition. The overall ratio of transition/transversion (ti/tv) of 0.724 for the intergenic spacer and 0.258 for the pooled sequences were estimated, and indicated that transversions are more frequent than transitions. The molecular evolution and demographic history of Q. suber were investigated. Neutrality tests (Tajima's D and Fu and Li) ruled out the null hypothesis of a strictly neutral model, and Fu's Fs and Ramos-Onsins and Rozas' R2 confirmed the recent expansion of cork oak trees, validating its persistency in North Africa since the last glaciation during the Quaternary. The observed uni-modal mismatch distribution and the Harpending's raggedness index confirmed the demographic history model for cork oak. A phylogenetic dendrogram showed that the distribution of Q. suber trees occurs independently of geographical origin, the relief of the population site, and the bioclimatic stages. The molecular history and cytoplasmic diversity suggest that in situ and ex situ conservation strategies can be recommended for preserving landscape value and facing predictable future climatic changes.

  2. Phylogenetic inferences of Nepenthes species in Peninsular Malaysia revealed by chloroplast (trnL intron) and nuclear (ITS) DNA sequences

    OpenAIRE

    Bunawan, Hamidun; Yen, Choong Chee; Yaakop, Salmah; Noor, Normah Mohd

    2017-01-01

    Background The chloroplastic trnL intron and the nuclear internal transcribed spacer (ITS) region were sequenced for 11 Nepenthes species recorded in Peninsular Malaysia to examine their phylogenetic relationship and to evaluate the usage of trnL intron and ITS sequences for phylogenetic reconstruction of this genus. Results Phylogeny reconstruction was carried out using neighbor-joining, maximum parsimony and Bayesian analyses. All the trees revealed two major clusters, a lowland group consi...

  3. Phylogenetic inferences of Nepenthes species in Peninsular Malaysia revealed by chloroplast (trnL intron) and nuclear (ITS) DNA sequences.

    Science.gov (United States)

    Bunawan, Hamidun; Yen, Choong Chee; Yaakop, Salmah; Noor, Normah Mohd

    2017-01-26

    The chloroplastic trnL intron and the nuclear internal transcribed spacer (ITS) region were sequenced for 11 Nepenthes species recorded in Peninsular Malaysia to examine their phylogenetic relationship and to evaluate the usage of trnL intron and ITS sequences for phylogenetic reconstruction of this genus. Phylogeny reconstruction was carried out using neighbor-joining, maximum parsimony and Bayesian analyses. All the trees revealed two major clusters, a lowland group consisting of N. ampullaria, N. mirabilis, N. gracilis and N. rafflesiana, and another containing both intermediately distributed species (N. albomarginata and N. benstonei) and four highland species (N. sanguinea, N. macfarlanei, N. ramispina and N. alba). The trnL intron and ITS sequences proved to provide phylogenetic informative characters for deriving a phylogeny of Nepenthes species in Peninsular Malaysia. To our knowledge, this is the first molecular phylogenetic study of Nepenthes species occurring along an altitudinal gradient in Peninsular Malaysia.

  4. Phylogenetic relationships in the genus Leonardoxa (Leguminosae: Caesalpinioideae) inferred from chloroplast trnL intron and trnL-trnF intergenic spacer sequences.

    Science.gov (United States)

    Brouat, Carine; Gielly, Ludovic; McKey, Doyle

    2001-01-01

    The African genus LEONARDOXA: (Leguminosae: Caesalpinioideae) comprises two Congolean species and a group of four mostly allopatric subspecies principally located in Cameroon and clustered together in the L. africana complex. LEONARDOXA: provides a good opportunity to investigate the evolutionary history of ant-plant mutualisms, as it exhibits various grades of ant-plant interactions from diffuse to obligate and symbiotic associations. We present in this paper the first molecular phylogenetic study of this genus. We sequenced both the chloroplast DNA trnL intron (677 aligned base pairs [bp]) and trnL-trnF intergene spacer (598 aligned bp). Inferred phylogenetic relationships suggested first that the genus is paraphyletic. The L. africana complex is clearly separated from the two Congolean species, and the integrity of the genus is thus in question. In the L. africana complex, our data showed a lack of congruence between clades suggested by morphological and chloroplast characters. This, and the low level of molecular divergence found between subspecies, suggests gene flow and introgressive events in the L. africana complex.

  5. Evolution of the tRNALeu (UAA) Intron and Congruence of Genetic Markers in Lichen-Symbiotic Nostoc.

    Science.gov (United States)

    Kaasalainen, Ulla; Olsson, Sanna; Rikkinen, Jouko

    2015-01-01

    The group I intron interrupting the tRNALeu UAA gene (trnL) is present in most cyanobacterial genomes as well as in the plastids of many eukaryotic algae and all green plants. In lichen symbiotic Nostoc, the P6b stem-loop of trnL intron always involves one of two different repeat motifs, either Class I or Class II, both with unresolved evolutionary histories. Here we attempt to resolve the complex evolution of the two different trnL P6b region types. Our analysis indicates that the Class II repeat motif most likely appeared first and that independent and unidirectional shifts to the Class I motif have since taken place repeatedly. In addition, we compare our results with those obtained with other genetic markers and find strong evidence of recombination in the 16S rRNA gene, a marker widely used in phylogenetic studies on Bacteria. The congruence of the different genetic markers is successfully evaluated with the recently published software Saguaro, which has not previously been utilized in comparable studies.

  6. Evolution of the tRNALeu (UAA Intron and Congruence of Genetic Markers in Lichen-Symbiotic Nostoc.

    Directory of Open Access Journals (Sweden)

    Ulla Kaasalainen

    Full Text Available The group I intron interrupting the tRNALeu UAA gene (trnL is present in most cyanobacterial genomes as well as in the plastids of many eukaryotic algae and all green plants. In lichen symbiotic Nostoc, the P6b stem-loop of trnL intron always involves one of two different repeat motifs, either Class I or Class II, both with unresolved evolutionary histories. Here we attempt to resolve the complex evolution of the two different trnL P6b region types. Our analysis indicates that the Class II repeat motif most likely appeared first and that independent and unidirectional shifts to the Class I motif have since taken place repeatedly. In addition, we compare our results with those obtained with other genetic markers and find strong evidence of recombination in the 16S rRNA gene, a marker widely used in phylogenetic studies on Bacteria. The congruence of the different genetic markers is successfully evaluated with the recently published software Saguaro, which has not previously been utilized in comparable studies.

  7. A CRM domain protein functions dually in group I and group II intron splicing in land plant chloroplasts.

    Science.gov (United States)

    Asakura, Yukari; Barkan, Alice

    2007-12-01

    The CRM domain is a recently recognized RNA binding domain found in three group II intron splicing factors in chloroplasts, in a bacterial protein that associates with ribosome precursors, and in a family of uncharacterized proteins in plants. To elucidate the functional repertoire of proteins with CRM domains, we studied CFM2 (for CRM Family Member 2), which harbors four CRM domains. RNA coimmunoprecipitation assays showed that CFM2 in maize (Zea mays) chloroplasts is associated with the group I intron in pre-trnL-UAA and group II introns in the ndhA and ycf3 pre-mRNAs. T-DNA insertions in the Arabidopsis thaliana ortholog condition a defective-seed phenotype (strong allele) or chlorophyll-deficient seedlings with impaired splicing of the trnL group I intron and the ndhA, ycf3-int1, and clpP-int2 group II introns (weak alleles). CFM2 and two previously described CRM proteins are bound simultaneously to the ndhA and ycf3-int1 introns and act in a nonredundant fashion to promote their splicing. With these findings, CRM domain proteins are implicated in the activities of three classes of catalytic RNA: group I introns, group II introns, and 23S rRNA.

  8. A trnL-F based phylogeny for species of Pelargonium (Geraniaceaea) with small chromosomes

    NARCIS (Netherlands)

    Bakker, F.T.; Culham, A.; Daugherty, L.C.; Gibby, M.

    1999-01-01

    Phylogenetic analysis was performed of 921 positions of trnL (UAA) 5 exon ¿ trnF (GAA) exon chloroplast DNA regions from 68 representatives ofPelargonium sectt.Campylia, Cortusina, Glaucophyllum, Hoarea, Isopetalum, Ligularia, Otidia, Pelargonium, Peristera, Polyactium, andReniformia, together with

  9. Phylogenetic relationships in Taxodiaceae and Cupressaceae sensu stricto based on matK gene, chlL gene, trnL-trnF IGS region, and trnL intron sequences.

    Science.gov (United States)

    Kusumi, J; Tsumura, Y; Yoshimaru, H; Tachida, H

    2000-10-01

    Nucleotide sequences from four chloroplast genes, the matK, chlL, intergenic spacer (IGS) region between trnL and trnF, and an intron of trnL, were determined from all species of Taxodiaceae and five species of Cupressaceae sensu stricto (s.s.). Phylogenetic trees were constructed using the maximum parsimony and the neighbor-joining methods with Cunninghamia as an outgroup. These analyses provided greater resolution of relationships among genera and higher bootstrap supports for clades compared to previous analyses. Results indicate that Taiwania diverged first, and then Athrotaxis diverged from the remaining genera. Metasequoia, Sequoia, and Sequoiadendron form a clade. Taxodium and Glyptostrobus form a clade, which is the sister to Cryptomeria. Cupressaceae s.s. are derived from within Taxodiaceae, being the most closely related to the Cryptomeria/Taxodium/Glyptostrobus clade. These relationships are consistent with previous morphological groupings and the analyses of molecular data. In addition, we found acceleration of evolutionary rates in Cupressaceae s.s. Possible causes for the acceleration are discussed.

  10. Phylogenetic relationships among vietnamese cocoa accessions using a non-coding region of the chloroplast dna

    International Nuclear Information System (INIS)

    Ha, L.T.V.; Dung, T.N.; Phuoc, P.H.D.

    2017-01-01

    Cocoa cultivation has increased in tropical areas around the world, including Vietnam, due to the high demand of cocoa beans for chocolate production. The genetic diversity of cocoa genotypes is recognized to be complex, however, their phylogenetic relationships need to be clarified. The present study aimed to classify the cocoa genotypes, that are imported and cultivated in Vietnam, based on a chloroplast DNA region. Sixty-three Vietnamese Cocoa accessions were collected from different regions in Southern Vietnam. Their phylogenetic relationships were identified using the universal primers c-B49317 and d-A49855 from the chloroplast DNA region. The sequences were situated in the trnL intron genes which are identify the closest terrestrial plant species of the chloroplast genome. DNA sequences were determined and subjected to an analysis of the phylogenetic relationship using the maximum evolution method. The genetic analysis showed clustering of 63 cocoa accessions in three groups: the domestically cultivated Trinitario group, the Indigenous cultivars, and the cultivations from Peru. The analyzed sequencing data also illustrated that the TD accessions and CT accessions were related genetically closed. Based on those results the genetic relation between PA and NA accessions was established as the hybrid origins of the TD and CT accessions. Some foreign accessions, including UIT, SCA and IMC accessions were confirmed of their genetic relationship. The present study is the first report of phylogenetic relationships of Vietnamese cocoa collections. The cocoa program in Vietnam has been in development for thirty years. (author)

  11. Chloroplast Redox Poise

    DEFF Research Database (Denmark)

    Steccanella, Verdiana

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

  12. Dichroism in spinach chloroplasts

    NARCIS (Netherlands)

    Thomas, J.B.; Lierop, J.H. van; Ham, M. ten

    1967-01-01

    In spinach chloroplasts oriented at steel-water interfaces parallel to the light beam a distinct dichroism is measured at about 680 nm. This dichroism is minimal upon addition of sucrose up to a final concentration of 0.18 M to the medium, the dichroic ratio amounting to 1.02. It is concluded that

  13. The use of the hypervariable P8 region of trnL(UAA intron for identification of orchid species: Evidence from restriction site polymorphism analysis.

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    Rajkumar Kishor

    Full Text Available The P8 stem-loop region of the trnL intron, which is known to be hypervariable in size with multiple repeat motifs and created difficulties in alignment, is always excluded in phylogenetic as well as barcode analyses. This region was investigated for species discrimination in 98 taxa of orchids belonging to the tribe Vandeae using in silico mapping of restriction site polymorphism. The length of the P8 regions varied from 200 nucleotides in Aerides rosea to 669 nucleotides in Dendrophylax sallei. Forty two taxa had unique lengths, while as many as eight shared a common length of 521 nucleotides. Of the 35 restriction endonucleases producing digestions in the P8 regions, three, viz., AgsI, ApoI and TspDTI turned out to have recognition sites across all the 98 taxa being studied. When their restriction data were combined, 92 taxa could be discriminated leaving three taxon pairs. However, Acampe papillosa and Aeranthes arachnites despite having similar restriction sites differed in their P8 lengths. This is the first report on thorough investigation of the P8 region of trnL intron for search of species specific restriction sites and hence its use as a potential plant DNA barcode.

  14. Length and nucleotide sequence polymorphism at the trnL and trnF non-coding regions of chloroplast genomes among Saccharum and Erianthus species

    Science.gov (United States)

    The aneupolyploidy genome of sugarcane (Saccharum hybrids spp.) and lack of a classical genetic linkage map make genetics research most difficult for sugarcane. Whole genome sequencing and genetic characterization of sugarcane and related taxa are far behind other crops. In this study, universal PCR...

  15. A Comparison of the First Two Sequenced Chloroplast Genomes in Asteraceae: Lettuce and Sunflower

    Energy Technology Data Exchange (ETDEWEB)

    Timme, Ruth E.; Kuehl, Jennifer V.; Boore, Jeffrey L.; Jansen, Robert K.

    2006-01-20

    Asteraceae is the second largest family of plants, with over 20,000 species. For the past few decades, numerous phylogenetic studies have contributed to our understanding of the evolutionary relationships within this family, including comparisons of the fast evolving chloroplast gene, ndhF, rbcL, as well as non-coding DNA from the trnL intron plus the trnLtrnF intergenic spacer, matK, and, with lesser resolution, psbA-trnH. This culminated in a study by Panero and Funk in 2002 that used over 13,000 bp per taxon for the largest taxonomic revision of Asteraceae in over a hundred years. Still, some uncertainties remain, and it would be very useful to have more information on the relative rates of sequence evolution among various genes and on genome structure as a potential set of phylogenetic characters to help guide future phylogenetic structures. By way of contributing to this, we report the first two complete chloroplast genome sequences from members of the Asteraceae, those of Helianthus annuus and Lactuca sativa. These plants belong to two distantly related subfamilies, Asteroideae and Cichorioideae, respectively. In addition to these, there is only one other published chloroplast genome sequence for any plant within the larger group called Eusterids II, that of Panax ginseng (Araliaceae, 156,318 bps, AY582139). Early chloroplast genome mapping studies demonstrated that H. annuus and L. sativa share a 22 kb inversion relative to members of the subfamily Barnadesioideae. By comparison to outgroups, this inversion was shown to be derived, indicating that the Asteroideae and Cichorioideae are more closely related than either is to the Barnadesioideae. Later sequencing study found that taxa that share this 22 kb inversion also contain within this region a second, smaller, 3.3 kb inversion. These sequences also enable an analysis of patterns of shared repeats in the genomes at fine level and of RNA editing by comparison to available EST sequences. In addition, since

  16. Taxonomic Identification of Mediterranean Pines and Their Hybrids Based on the High Resolution Melting (HRM) and trnL Approaches: From Cytoplasmic Inheritance to Timber Tracing

    Science.gov (United States)

    Ganopoulos, Ioannis; Aravanopoulos, Filippos; Madesis, Panagiotis; Pasentsis, Konstantinos; Bosmali, Irene; Ouzounis, Christos; Tsaftaris, Athanasios

    2013-01-01

    Fast and accurate detection of plant species and their hybrids using molecular tools will facilitate the assessment and monitoring of local biodiversity in an era of climate and environmental change. Herein, we evaluate the utility of the plastid trnL marker for species identification applied to Mediterranean pines (Pinus spp.). Our results indicate that trnL is a very sensitive marker for delimiting species biodiversity. Furthermore, High Resolution Melting (HRM) analysis was exploited as a molecular fingerprint for fast and accurate discrimination of Pinus spp. DNA sequence variants. The trnL approach and the HRM analyses were extended to wood samples of two species (Pinus nigra and Pinus sylvestris) with excellent results, congruent to those obtained using leaf tissue. Both analyses demonstrate that hybrids from the P. brutia (maternal parent) × P. halepensis (paternal parent) cross, exhibit the P. halepensis profile, confirming paternal plastid inheritance in Group Halepensis pines. Our study indicates that a single one-step reaction method and DNA marker are sufficient for the identification of Mediterranean pines, their hybrids and the origin of pine wood. Furthermore, our results underline the potential for certain DNA regions to be used as novel biological information markers combined with existing morphological characters and suggest a relatively reliable and open taxonomic system that can link DNA variation to phenotype-based species or hybrid assignment status and direct taxa identification from recalcitrant tissues such as wood samples. PMID:23577179

  17. Protein import into chloroplasts requires a chloroplast ATPase

    International Nuclear Information System (INIS)

    Pain, D.; Blobel, G.

    1987-01-01

    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 [ 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 + , K + , Na + , 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

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

    Directory of Open Access Journals (Sweden)

    Prakitchai Chotewutmontri

    2016-07-01

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

  19. Towards a synthetic chloroplast.

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    Christina M Agapakis

    2011-04-01

    Full Text Available The evolution of eukaryotic cells is widely agreed to have proceeded through a series of endosymbiotic events between larger cells and proteobacteria or cyanobacteria, leading to the formation of mitochondria or chloroplasts, respectively. Engineered endosymbiotic relationships between different species of cells are a valuable tool for synthetic biology, where engineered pathways based on two species could take advantage of the unique abilities of each mutualistic partner.We explored the possibility of using the photosynthetic bacterium Synechococcus elongatus PCC 7942 as a platform for studying evolutionary dynamics and for designing two-species synthetic biological systems. We observed that the cyanobacteria were relatively harmless to eukaryotic host cells compared to Escherichia coli when injected into the embryos of zebrafish, Danio rerio, or taken up by mammalian macrophages. In addition, when engineered with invasin from Yersinia pestis and listeriolysin O from Listeria monocytogenes, S. elongatus was able to invade cultured mammalian cells and divide inside macrophages.Our results show that it is possible to engineer photosynthetic bacteria to invade the cytoplasm of mammalian cells for further engineering and applications in synthetic biology. Engineered invasive but non-pathogenic or immunogenic photosynthetic bacteria have great potential as synthetic biological devices.

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

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    Miao An

    2015-06-01

    Full Text Available 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.

  1. Protein import into chloroplasts requires a chloroplast ATPase

    Energy Technology Data Exchange (ETDEWEB)

    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.

  2. Assessment of the food habits of the Moroccan dorcas gazelle in M'Sabih Talaa, west central Morocco, using the trnL approach.

    Directory of Open Access Journals (Sweden)

    Moulay Abdeljalil Ait Baamrane

    Full Text Available Food habits of the Moroccan dorcas gazelle, Gazella dorcas massaesyla, previously investigated in the 1980s using microhistological fecal analysis, in the M'Sabih Talaa Reserve, west central Morocco, were re-evaluated over three seasons (spring, summer and autumn 2009 using the trnL approach to determine the diet composition and its seasonal variation from fecal samples. Taxonomic identification was carried out using the identification originating from the database built from EMBL and the list of plant species within the reserve. The total taxonomic richness in the reserve was 130 instead of 171 species in the 1980s. The diet composition revealed to be much more diversified (71 plant taxa belonging to 57 genus and 29 families than it was 22 years ago (29 identified taxa. Thirty-four taxa were newly identified in the diet while 13 reported in 1986-87 were not found. Moroccan dorcas gazelle showed a high preference to Acacia gummifera, Anagallis arvensis, Glebionis coronaria, Cladanthus arabicus, Diplotaxis tenuisiliqua, Erodium salzmannii, Limonium thouini, Lotus arenarius and Zizyphus lotus. Seasonal variations occurred in both number (40-41 taxa in spring-summer and 49 taxa in autumn vs. respectively 23-22 and 26 in 1986-1987 and taxonomic type of eaten plant taxa. This dietary diversification could be attributed either to the difference in methods of analysis, trnL approach having a higher taxonomic resolution, or a potential change in nutritional quality of plants over time.

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

    International Nuclear Information System (INIS)

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

    1986-01-01

    In addition to chlorophyll-protein complexes, other proteins were labeled when isolated developing pea chloroplasts were incubated with [ 14 C]-5-aminolevulinic acid [ 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 2 O 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 [ 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

  4. Protein methylation in pea chloroplasts

    International Nuclear Information System (INIS)

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

    1990-01-01

    The methylation of chloroplast proteins has been investigated by incubating intact pea (Pisum sativum) chloroplasts with [ 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 [ 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 [ 3 H]methyl group

  5. On the structure of the spinach chloroplast

    NARCIS (Netherlands)

    Thomas, J.B.; Bustraan, M.; Paris, C.H.

    1952-01-01

    The structure of spinach chloroplasts was investigated with the aid of the electron microscope. It has been established that: 1. 1. the outer membrane of the chloroplasts is composed of both proteins and lipoids. 2. 2. the stroma is also built up by these components. 3. 3. within the

  6. The role of chloroplasts in plant pathology.

    Science.gov (United States)

    Sowden, Robert G; Watson, Samuel J; Jarvis, Paul

    2018-04-13

    Plants have evolved complex tolerance systems to survive abiotic and biotic stresses. Central to these programmes is a sophisticated conversation of signals between the chloroplast and the nucleus. In this review, we examine the antagonism between abiotic stress tolerance (AST) and immunity: we propose that to generate immunogenic signals, plants must disable AST systems, in particular those that manage reactive oxygen species (ROS), while the pathogen seeks to reactivate or enhance those systems to achieve virulence. By boosting host systems of AST, pathogens trick the plant into suppressing chloroplast immunogenic signals and steer the host into making an inappropriate immune response. Pathogens disrupt chloroplast function, both transcriptionally-by secreting effectors that alter host gene expression by interacting with defence-related kinase cascades, with transcription factors, or with promoters themselves-and post-transcriptionally, by delivering effectors that enter the chloroplast or alter the localization of host proteins to change chloroplast activities. These mechanisms reconfigure the chloroplast proteome and chloroplast-originating immunogenic signals in order to promote infection. © 2017 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

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

    OpenAIRE

    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, tobacc...

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

    Science.gov (United States)

    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.

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

    Science.gov (United States)

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

    2016-02-23

    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.

  10. Fatty acid synthesis by spinach chloroplasts, 2

    International Nuclear Information System (INIS)

    Yamada, Mitsuhiro; Nakamura, Yasunori

    1975-01-01

    By incorporation of 3 H 2 O into the fatty acid chain in the presence of unlabelled precursor, we showed that fatty acids are synthesized from PGA, PEP and pyruvate by intact spinach chloroplasts in the light. 13 C-tracer experiments confirmed that 1-C of pyruvate is decarboxylated and 2-C is incorporated into fatty acids by the chloroplasts. The patterns of fatty acids synthesized from PGA and pyruvate were the same as that from acetate. The highest rate of fatty acid synthesis was reached at the physiological concentration of PGA (3 mM) and pyruvate (1 mM). These results indicate the operation of the following path in the chloroplasts in light: PGA→PEP→pyruvate→acetylCoA→fatty acids. Since citrate and OAA were much less active and malate and glyoxylate were inert as precursors for fatty acid synthesis, PEP or pyruvate carboxylation, citrate lyase reaction and malate synthetase reaction are not involved in the formation of acetylCoA and fatty acids. Since pyruvate was much more effective as a substrate for fatty acid synthesis than lactate, acetaldehyde or acetate, direct decarboxylation path is considered to be the primary path from pyruvate to acetylCoA. The insignificant effect of chloroplast-washing on fatty acid synthesis from PGA and pyruvate indicates that the glycolytic path from PGA to pyruvate is associated with the chloroplasts. Since pyruvate was more effectively incorporated into fatty acids than acetylCoA, it is unlikely that pyruvate decarboxylation to acetylCoA is due to mitochondria contaminating the chloroplast preparation. On the basis of measurements of 3 H 2 O incorporation in the light and dark, the activity of fatty acid synthesis in spincah leaves appears to be shared by the activities in chloroplasts (87%) and other organelles (13%). (author)

  11. Phylogenomic Analysis and Dynamic Evolution of Chloroplast Genomes in Salicaceae

    Directory of Open Access Journals (Sweden)

    Yuan Huang

    2017-06-01

    Full Text Available Chloroplast genomes of plants are highly conserved in both gene order and gene content. Analysis of the whole chloroplast genome is known to provide much more informative DNA sites and thus generates high resolution for plant phylogenies. Here, we report the complete chloroplast genomes of three Salix species in family Salicaceae. Phylogeny of Salicaceae inferred from complete chloroplast genomes is generally consistent with previous studies but resolved with higher statistical support. Incongruences of phylogeny, however, are observed in genus Populus, which most likely results from homoplasy. By comparing three Salix chloroplast genomes with the published chloroplast genomes of other Salicaceae species, we demonstrate that the synteny and length of chloroplast genomes in Salicaceae are highly conserved but experienced dynamic evolution among species. We identify seven positively selected chloroplast genes in Salicaceae, which might be related to the adaptive evolution of Salicaceae species. Comparative chloroplast genome analysis within the family also indicates that some chloroplast genes are lost or became pseudogenes, infer that the chloroplast genes horizontally transferred to the nucleus genome. Based on the complete nucleus genome sequences from two Salicaceae species, we remarkably identify that the entire chloroplast genome is indeed transferred and integrated to the nucleus genome in the individual of the reference genome of P. trichocarpa at least once. This observation, along with presence of the large nuclear plastid DNA (NUPTs and NUPTs-containing multiple chloroplast genes in their original order in the chloroplast genome, favors the DNA-mediated hypothesis of organelle to nucleus DNA transfer. Overall, the phylogenomic analysis using chloroplast complete genomes clearly elucidates the phylogeny of Salicaceae. The identification of positively selected chloroplast genes and dynamic chloroplast-to-nucleus gene transfers in

  12. Nitrogen control of chloroplast development: Progress report

    International Nuclear Information System (INIS)

    Schmidt, G.W.

    1987-11-01

    A manifestation of nitrogen deficiency in vascular plants and algae is chlorosis, indicating that chloroplast biogenesis can be strongly restricted by direct or indirect effects of nitrogen assimilation products. To define the molecular basis of nitrogen responses we are using Chlamydomonas reinhardtii. Depending on the levels of ammonium, steady-state deficiency conditions are established such that the cellular levels of chlorophylls and xanthophylls are depressed. Chloroplasts in nitrogen-deficient cells contain appreciable levels of carbon assimilation enzyme and thylakoids with high electron transport activities. However, the light harvesting complexes are nearly absent and Photosystem I exhibits unusual characteristics. Studies of rates of protein synthesis by in vivo pulse-chase labeling and levels of RNAs encoded by the chloroplast and nuclear genomes have been initiated: the accumulation of transcripts for the nuclear light-harvesting apoproteins is dramatically altered qualitatively and quantitatively; there is no major effect on chloroplast RNAs but, in general, these are inefficiently utilized for protein synthesis until nitrogen is provided to the cultures. Supplying nitrogen results in an almost immediate release of chloroplast mRNAs from a translational arrest but the stimulation of the accumulation of nuclear transcripts for light-harvesting apoproteins does not occur until after a 1-2 hour lag

  13. Nitrogen control of chloroplast development: Progress report

    Energy Technology Data Exchange (ETDEWEB)

    Schmidt, G.W.

    1987-11-01

    A manifestation of nitrogen deficiency in vascular plants and algae is chlorosis, indicating that chloroplast biogenesis can be strongly restricted by direct or indirect effects of nitrogen assimilation products. To define the molecular basis of nitrogen responses we are using Chlamydomonas reinhardtii. Depending on the levels of ammonium, steady-state deficiency conditions are established such that the cellular levels of chlorophylls and xanthophylls are depressed. Chloroplasts in nitrogen-deficient cells contain appreciable levels of carbon assimilation enzyme and thylakoids with high electron transport activities. However, the light harvesting complexes are nearly absent and Photosystem I exhibits unusual characteristics. Studies of rates of protein synthesis by in vivo pulse-chase labeling and levels of RNAs encoded by the chloroplast and nuclear genomes have been initiated: the accumulation of transcripts for the nuclear light-harvesting apoproteins is dramatically altered qualitatively and quantitatively; there is no major effect on chloroplast RNAs but, in general, these are inefficiently utilized for protein synthesis until nitrogen is provided to the cultures. Supplying nitrogen results in an almost immediate release of chloroplast mRNAs from a translational arrest but the stimulation of the accumulation of nuclear transcripts for light-harvesting apoproteins does not occur until after a 1-2 hour lag.

  14. Mergers and acquisitions: malaria and the great chloroplast heist.

    Science.gov (United States)

    McFadden, G I

    2000-01-01

    The origin of the relict chloroplast recently identified in malarial parasites has been mysterious. Several new papers suggest that the parasites obtained their chloroplasts in an ancient endosymbiotic event that also created some major algal groups.

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

    Science.gov (United States)

    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.

  16. Protein methylation reactions in intact pea chloroplasts

    International Nuclear Information System (INIS)

    Niemi, K.J.

    1989-01-01

    Post-translational protein methylation was investigated in Pisum sativum chloroplasts. Intact pea chloroplasts were incubated with ( 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

  17. The demise of chloroplast DNA in Arabidopsis.

    Science.gov (United States)

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

    2004-09-01

    Although it might be expected that chloroplast DNA (cpDNA) would be stably maintained in mature leaves, we report the surprising observation that cpDNA levels decline during plastid development in Arabidopsis thaliana (Col.) until most of the leaves contain little or no DNA long before the onset of senescence. We measured the cpDNA content in developing cotyledons, rosette leaves, and cauline leaves. The amount of cpDNA per chloroplast decreases as the chloroplasts develop, reaching undetectable levels in mature leaves. In young cauline leaves, most individual molecules of cpDNA are found in complex, branched forms. In expanded cauline leaves, cpDNA is present in smaller branched forms only at the base of the leaf and is virtually absent in the distal part of the leaf. We conclude that photosynthetic activity may persist long after the demise of the cpDNA. Copyright 2004 Springer-Verlag

  18. Utilization of complete chloroplast genomes for phylogenetic studies

    NARCIS (Netherlands)

    Ramlee, Shairul Izan Binti

    2016-01-01

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

  19. Non radioactive precursor import into chloroplasts

    International Nuclear Information System (INIS)

    Lombardo, V.A.; Ottado, J.

    2003-01-01

    Full text: Eukaryotic cells have a subcellular organization based on organelles. Protein transport to these organelles is quantitatively important because the majority of cellular proteins are codified in nuclear genes and then delivered to their final destination. Most of the chloroplast proteins are translated on cytoplasmic ribosomes as larger precursors with an amino terminal transit peptide that is necessary and sufficient to direct the precursor to the chloroplast. Once inside the organelle the transit peptide is cleaved and the mature protein adopts its folded form. In this work we developed a system for the expression and purification of the pea ferredoxin-NADP + reductase precursor (preFNR) for its import into chloroplasts in non radioactive conditions. We constructed a preFNR fused in its carboxy terminus to a 6 histidines peptide (preFNR-6xHis) that allows its identification using a commercial specific antibody. The construction was expressed, purified, processed and precipitated, rendering a soluble and active preFNR-6xHis that was used in binding and import into chloroplasts experiments. The reisolated chloroplasts were analyzed by SDS-PAGE, electro-blotting and revealed by immuno-detection using either colorimetric or chemiluminescent reactive. We performed also import experiments labeling preFNR and preFNR-6xHis with radioactive methionine as controls. We conclude that preFNR-6xHis is bound and imported into chloroplasts as the wild type preFNR and that both colorimetric or chemiluminescent detection methods are useful to avoid the manipulation of radioactive material. (author)

  20. A comparison of rice chloroplast genomes

    DEFF Research Database (Denmark)

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

    2004-01-01

    Using high quality sequence reads extracted from our whole genome shotgun repository, we assembled two chloroplast genome sequences from two rice (Oryza sativa) varieties, one from 93-11 (a typical indica variety) and the other from PA64S (an indica-like variety with maternal origin of japonica......), which are both parental varieties of the super-hybrid rice, LYP9. Based on the patterns of high sequence coverage, we partitioned chloroplast sequence variations into two classes, intravarietal and intersubspecific polymorphisms. Intravarietal polymorphisms refer to variations within 93-11 or PA64S...

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

    Science.gov (United States)

    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.

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

    Directory of Open Access Journals (Sweden)

    Marta Brozynska

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

  3. Chloroplast Signaling Gates Thermotolerance in Arabidopsis

    Directory of Open Access Journals (Sweden)

    Patrick J. Dickinson

    2018-02-01

    Full Text Available Temperature is a key environmental variable influencing plant growth and survival. Protection against high temperature stress in eukaryotes is coordinated by heat shock factors (HSFs, transcription factors that activate the expression of protective chaperones such as HEAT SHOCK PROTEIN 70 (HSP70; however, the pathway by which temperature is sensed and integrated with other environmental signals into adaptive responses is not well understood. Plants are exposed to considerable diurnal variation in temperature, and we have found that there is diurnal variation in thermotolerance in Arabidopsis thaliana, with maximal thermotolerance coinciding with higher HSP70 expression during the day. In a forward genetic screen, we identified a key role for the chloroplast in controlling this response, suggesting that light-induced chloroplast signaling plays a key role. Consistent with this, we are able to globally activate binding of HSFA1a to its targets by altering redox status in planta independently of a heat shock.

  4. Chloroplast Chaperonin: An Intricate Protein Folding Machine for Photosynthesis

    Directory of Open Access Journals (Sweden)

    Qian Zhao

    2018-01-01

    Full Text Available Group I chaperonins are large cylindrical-shaped nano-machines that function as a central hub in the protein quality control system in the bacterial cytosol, mitochondria and chloroplasts. In chloroplasts, proteins newly synthesized by chloroplast ribosomes, unfolded by diverse stresses, or translocated from the cytosol run the risk of aberrant folding and aggregation. The chloroplast chaperonin system assists these proteins in folding into their native states. A widely known protein folded by chloroplast chaperonin is the large subunit of ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco, an enzyme responsible for the fixation of inorganic CO2 into organic carbohydrates during photosynthesis. Chloroplast chaperonin was initially identified as a Rubisco-binding protein. All photosynthetic eucaryotes genomes encode multiple chaperonin genes which can be divided into α and β subtypes. Unlike the homo-oligomeric chaperonins from bacteria and mitochondria, chloroplast chaperonins are more complex and exists as intricate hetero-oligomers containing both subtypes. The Group I chaperonin requires proper interaction with a detachable lid-like co-chaperonin in the presence of ATP and Mg2+ for substrate encapsulation and conformational transition. Besides the typical Cpn10-like co-chaperonin, a unique co-chaperonin consisting of two tandem Cpn10-like domains joined head-to-tail exists in chloroplasts. Since chloroplasts were proposed as sensors to various environmental stresses, this diversified chloroplast chaperonin system has the potential to adapt to complex conditions by accommodating specific substrates or through regulation at both the transcriptional and post-translational levels. In this review, we discuss recent progress on the unique structure and function of the chloroplast chaperonin system based on model organisms Chlamydomonas reinhardtii and Arabidopsis thaliana. Knowledge of the chloroplast chaperonin system may ultimately lead

  5. Mechanism of protein import across the chloroplast envelope.

    Science.gov (United States)

    Chen, K; Chen, X; Schnell, D J

    2000-01-01

    The development and maintenance of chloroplasts relies on the contribution of protein subunits from both plastid and nuclear genomes. Most chloroplast proteins are encoded by nuclear genes and are post-translationally imported into the organelle across the double membrane of the chloroplast envelope. Protein import into the chloroplast consists of two essential elements: the specific recognition of the targeting signals (transit sequences) of cytoplasmic preproteins by receptors at the outer envelope membrane and the subsequent translocation of preproteins simultaneously across the double membrane of the envelope. These processes are mediated via the co-ordinate action of protein translocon complexes in the outer (Toc apparatus) and inner (Tic apparatus) envelope membranes.

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

    Directory of Open Access Journals (Sweden)

    Schmitz-Linneweber Christian

    2008-08-01

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

  7. Orientation of the pigment molecules in the chloroplast

    NARCIS (Netherlands)

    Goedheer, J.C.

    1955-01-01

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

  8. Comparative studies on codon usage pattern of chloroplasts and ...

    Indian Academy of Sciences (India)

    Unknown

    different genomic organization and mutation pressures in nuclear and chloroplast genes. The results of Nc-plots and neutrality plots ... As an important organelle of plants, the chloroplast has its own genomic environment and ... leading to the suggestion that the translation mechanism and patterns of codon usage in ...

  9. Carbon dioxide fixation in isolated Kalanchoe chloroplasts

    Energy Technology Data Exchange (ETDEWEB)

    Levi, C.; Gibbs, M.

    1975-07-01

    Chloroplasts isolated from Kalanchoe diagremontiana leaves were capable of photosynthesizing at a rate of 5.4 ..mu..moles of CO/sub 2/ per milligram of chlorophyll per hour. The dark rate of fixation was about 1 percent of the light rate. A high photosynthetic rate was associated with low starch content of the leaves. Ribose 5-phosphate, fructose 1, 6-diphosphate, and dithiothreitol stimulated fixation, whereas phosphoenolpyruvate and azide were inhibitors. The products of CO/sub 2/ fixation were primarily those of the photosynthetic carbon reduction cycle. (auth)

  10. Regulation of Chloroplastic Carbonic Anhydrase 1

    Science.gov (United States)

    Porter, Michael A.; Grodzinski, Bernard

    1983-01-01

    It was previously reported that magnesium ion inhibited carbonic anhydrase (Bamberger and Avron 1975 Plant Physiol 56: 481-485). Studies with partially purified carbonic anhydrase from spinach (Spinacia oleracea L.) chloroplasts show that the effect was the result of the chloride counterion and not the magnesium ion. Enzyme activity was reduced 50% upon addition of 3 to 10 millimolar MgCl2 or KCl while all additions of MgSO4 between 0.3 and 10 millimolar were mildly stimulatory. PMID:16663052

  11. C4 photosynthetic machinery: insights from maize chloroplast proteomics

    Directory of Open Access Journals (Sweden)

    Qi eZhao

    2013-04-01

    Full Text Available C4 plants exhibit much higher CO2 assimilation rates than C3 plants. The specialized differentiation of mesophyll cell (M and bundle sheath cell (BS type chloroplasts is unique to C4 plants and improves photosynthesis efficiency. Maize (Zea mays is an important crop and model with C4 photosynthetic machinery. Current high-throughput quantitative proteomics approaches (e.g., 2DE, iTRAQ, and shotgun proteomics have been employed to investigate maize chloroplast structure and function. These proteomic studies have provided valuable information on C4 chloroplast protein components, photosynthesis, and other metabolic mechanisms underlying chloroplast biogenesis, stromal and membrane differentiation, as well as response to salinity, high/low temperature, and light stress. This review presents an overview of proteomics advances in maize chloroplast biology.

  12. A protocol for expression of foreign genes in chloroplasts.

    Science.gov (United States)

    Verma, Dheeraj; Samson, Nalapalli P; Koya, Vijay; Daniell, Henry

    2008-01-01

    Several major costs associated with the production of biopharmaceuticals or vaccines in fermentation-based systems could be minimized by using plant chloroplasts as bioreactors, which facilitates rapid scale-up. Oral delivery of chloroplast-derived therapeutic proteins through plant cells eliminates expensive purification steps, low temperature storage, transportation and sterile injections for their delivery. Chloroplast transformation technology (CTT) has also been successfully used to engineer valuable agronomic traits and for the production of industrial enzymes and biomaterials. Here, we provide a detailed protocol for the construction of chloroplast expression and integration vectors, selection and regeneration of transformants, evaluation of transgene integration and inheritance, confirmation of transgene expression and extraction, and quantitation and purification of foreign proteins. Integration of appropriate transgenes into chloroplast genomes and the resulting high levels of functional protein expression can be achieved in approximately 6 months in lettuce and tobacco. CTT is eco-friendly because transgenes are maternally inherited in most crop plants.

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

    Science.gov (United States)

    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.

  14. Chloroplasts in anther endothecium of Zea mays (Poaceae).

    Science.gov (United States)

    Murphy, Katherine M; Egger, Rachel L; Walbot, Virginia

    2015-11-01

    Although anthers of Zea mays, Oryza sativa, and Arabidopsis thaliana have been studied intensively using genetic and biochemical analyses in the past 20 years, few updates to anther anatomical and ultrastructural descriptions have been reported. For example, no transmission electron microscopy (TEM) images of the premeiotic maize anther have been published. Here we report the presence of chloroplasts in maize anthers. TEM imaging, electron acceptor photosynthesis assay, in planta photon detection, microarray analysis, and light and fluorescence microscopy were used to investigate the presence of chloroplasts in the maize anther. Most cells of the maize subepidermal endothecium have starch-containing chloroplasts that do not conduct measurable photosynthesis in vitro. The maize anther contains chloroplasts in most subepidermal, endothecial cells. Although maize anthers receive sufficient light to photosynthesize in vivo and the maize anther transcribes >96% of photosynthesis-associated genes found in the maize leaf, no photosynthetic light reaction activity was detected in vitro. The endothecial cell layer should no longer be defined as a complete circle viewed transversely in anther lobes, because chloroplasts are observed only in cells directly beneath the epidermis and not those adjacent to the connective tissue. We propose that chloroplasts be a defining characteristic of differentiated endothecial cells and that nonsubepidermal endothecial cells that lack chloroplasts be defined as a separate cell type, the interendothecium. © 2015 Botanical Society of America.

  15. Development of Chloroplast Genomic Resources in Chinese Yam (Dioscorea polystachya

    Directory of Open Access Journals (Sweden)

    Junling Cao

    2018-01-01

    Full Text Available Chinese yam has been used both as a food and in traditional herbal medicine. Developing more effective genetic markers in this species is necessary to assess its genetic diversity and perform cultivar identification. In this study, new chloroplast genomic resources were developed using whole chloroplast genomes from six genotypes originating from different geographical locations. The Dioscorea polystachya chloroplast genome is a circular molecule consisting of two single-copy regions separated by a pair of inverted repeats. Comparative analyses of six D. polystachya chloroplast genomes revealed 141 single nucleotide polymorphisms (SNPs. Seventy simple sequence repeats (SSRs were found in the six genotypes, including 24 polymorphic SSRs. Forty-three common indels and five small inversions were detected. Phylogenetic analysis based on the complete chloroplast genome provided the best resolution among the genotypes. Our evaluation of chloroplast genome resources among these genotypes led us to consider the complete chloroplast genome sequence of D. polystachya as a source of reliable and valuable molecular markers for revealing biogeographical structure and the extent of genetic variation in wild populations and for identifying different cultivars.

  16. Global RNA association with the transcriptionally active chromosome of chloroplasts.

    Science.gov (United States)

    Lehniger, Marie-Kristin; Finster, Sabrina; Melonek, Joanna; Oetke, Svenja; Krupinska, Karin; Schmitz-Linneweber, Christian

    2017-10-01

    Processed chloroplast RNAs are co-enriched with preparations of the chloroplast transcriptionally active chromosome. Chloroplast genomes are organized as a polyploid DNA-protein structure called the nucleoid. Transcriptionally active chloroplast DNA together with tightly bound protein factors can be purified by gel filtration as a functional entity called the transcriptionally active chromosome (TAC). Previous proteomics analyses of nucleoids and of TACs demonstrated a considerable overlap in protein composition including RNA binding proteins. Therefore the RNA content of TAC preparations from Nicotiana tabacum was determined using whole genome tiling arrays. A large number of chloroplast RNAs was found to be associated with the TAC. The pattern of RNAs attached to the TAC consists of RNAs produced by different chloroplast RNA polymerases and differs from the pattern of RNA found in input controls. An analysis of RNA splicing and RNA editing of selected RNA species demonstrated that TAC-associated RNAs are processed to a similar extent as the RNA in input controls. Thus, TAC fractions contain a specific subset of the processed chloroplast transcriptome.

  17. Complete chloroplast genome of Gracilaria firma (Gracilariaceae, Rhodophyta), with discussion on the use of chloroplast phylogenomics in the subclass Rhodymeniophycidae.

    Science.gov (United States)

    Ng, Poh-Kheng; Lin, Showe-Mei; Lim, Phaik-Eem; Liu, Li-Chia; Chen, Chien-Ming; Pai, Tun-Wen

    2017-01-06

    The chloroplast genome of Gracilaria firma was sequenced in view of its role as an economically important marine crop with wide industrial applications. To date, there are only 15 chloroplast genomes published for the Florideophyceae. Apart from presenting the complete chloroplast genome of G. firma, this study also assessed the utility of genome-scale data to address the phylogenetic relationships within the subclass Rhodymeniophycidae. The synteny and genome structure of the chloroplast genomes across the taxa of Eurhodophytina was also examined. The chloroplast genome of Gracilaria firma maps as a circular molecule of 187,001 bp and contains 252 genes, which are distributed on both strands and consist of 35 RNA genes (3 rRNAs, 30 tRNAs, tmRNA and a ribonuclease P RNA component) and 217 protein-coding genes, including the unidentified open reading frames. The chloroplast genome of G. firma is by far the largest reported for Gracilariaceae, featuring a unique intergenic region of about 7000 bp with discontinuous vestiges of red algal plasmid DNA sequences interspersed between the nblA and cpeB genes. This chloroplast genome shows similar gene content and order to other Florideophycean taxa. Phylogenomic analyses based on the concatenated amino acid sequences of 146 protein-coding genes confirmed the monophyly of the classes Bangiophyceae and Florideophyceae with full nodal support. Relationships within the subclass Rhodymeniophycidae in Florideophyceae received moderate to strong nodal support, and the monotypic family of Gracilariales were resolved with maximum support. Chloroplast genomes hold substantial information that can be tapped for resolving the phylogenetic relationships of difficult regions in the Rhodymeniophycidae, which are perceived to have experienced rapid radiation and thus received low nodal support, as exemplified in this study. The present study shows that chloroplast genome of G. firma could serve as a key link to the full resolution of

  18. Biogeography of the Pistia clade (Araceae): based on chloroplast and mitochondrial DNA sequences and Bayesian divergence time inference.

    Science.gov (United States)

    Renner, Susanne S; Zhang, Li-Bing

    2004-06-01

    Pistia stratiotes (water lettuce) and Lemna (duckweeds) are the only free-floating aquatic Araceae. The geographic origin and phylogenetic placement of these unrelated aroids present long-standing problems because of their highly modified reproductive structures and wide geographical distributions. We sampled chloroplast (trnL-trnF and rpl20-rps12 spacers, trnL intron) and mitochondrial sequences (nad1 b/c intron) for all genera implicated as close relatives of Pistia by morphological, restriction site, and sequencing data, and present a hypothesis about its geographic origin based on the consensus of trees obtained from the combined data, using Bayesian, maximum likelihood, parsimony, and distance analyses. Of the 14 genera closest to Pistia, only Alocasia, Arisaema, and Typhonium are species-rich, and the latter two were studied previously, facilitating the choice of representatives that span the roots of these genera. Results indicate that Pistia and the Seychelles endemic Protarum sechellarum are the basalmost branches in a grade comprising the tribes Colocasieae (Ariopsis, Steudnera, Remusatia, Alocasia, Colocasia), Arisaemateae (Arisaema, Pinellia), and Areae (Arum, Biarum, Dracunculus, Eminium, Helicodiceros, Theriophonum, Typhonium). Unexpectedly, all Areae genera are embedded in Typhonium, which throws new light on the geographic history of Areae. A Bayesian analysis of divergence times that explores the effects of multiple fossil and geological calibration points indicates that the Pistia lineage is 90 to 76 million years (my) old. The oldest fossils of the Pistia clade, though not Pistia itself, are 45-my-old leaves from Germany; the closest outgroup, Peltandreae (comprising a few species in Florida, the Mediterranean, and Madagascar), is known from 60-my-old leaves from Europe, Kazakhstan, North Dakota, and Tennessee. Based on the geographic ranges of close relatives, Pistia likely originated in the Tethys region, with Protarum then surviving on the

  19. Separation of Chloroplast Pigments Using Reverse Phase Chromatography.

    Science.gov (United States)

    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)

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

    International Nuclear Information System (INIS)

    McCarty, R. E.

    2004-01-01

    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

  1. Inhibition of chloroplast protein synthesis following light chilling of tomato

    International Nuclear Information System (INIS)

    Kent, J.; Ort, D.

    1989-01-01

    In the present study we looked at the effects of a high light chill on the pulsed incorporation of 35 S methionine into total, stromal, and thylakoid proteins of lightly abraded leaflets of 18-21 day old tomato (Lycopersicon esculentum Mill ca. Floramerica) seedlings. Based on gel fluorographic patterns of marker proteins that are indicative of the net rates of chloroplast and cytoplasmic protein synthesis, there appears to be a nearly complete cessation of chloroplastic protein synthesis. No labeling is observed for either the stromal large subunit of Rubisco or the thylakoid-bound alpha and beta subunits of the coupling factor. One notable exception, however, appears to be the 32 kd, D1 protein. Its net synthetic rate remains high despite the inhibition of other chloroplastically synthesized proteins. The small subunit of Rubicso, LHCP-II, as well as several other proteins of known cytoplasmic origin, were still synthesized, albeit, at lower than control rates. Light chilling of chill-insensitive spinach produced a similar, but less dramatic differential behavior between chloroplastic and cytoplasmic protein synthesis. It appears, in chilling-sensitive plants, that chloroplast protein synthesis exhibits a greater sensitivity to low temperature inhibition than does cytoplasmic protein synthesis and that recovery of chloroplast protein synthesis may play an important role in recovery of photosynthetic activity following chilling

  2. Chloroplast microsatellite markers for Pseudotaxus chienii developed from the whole chloroplast genome of Taxus chinensis var. mairei (Taxaceae).

    Science.gov (United States)

    Deng, Qi; Zhang, Hanrui; He, Yipeng; Wang, Ting; Su, Yingjuan

    2017-03-01

    Pseudotaxus chienii (Taxaceae) is an old rare species endemic to China that has adapted well to ecological heterogeneity with high genetic diversity in its nuclear genome. However, the genetic variation in its chloroplast genome is unknown. Eighteen chloroplast microsatellite markers (cpSSRs) were developed from the whole chloroplast genome of Taxus chinensis var. mairei and successfully amplified in four P. chienii populations and one T. chinensis var. mairei population. Of these loci, 10 were polymorphic in P. chienii , whereas six were polymorphic in T. chinensis var. mairei . The unbiased haploid diversity per locus ranged from 0.000 to 0.641 and 0.000 to 0.545 for P. chienii and T. chinensis var. mairei , respectively. The 18 cpSSRs will be used to further investigate the chloroplast genetic structure and adaptive evolution in P. chienii populations.

  3. The complete chloroplast genome sequence of Helwingia himalaica (Helwingiaceae, Aquifoliales) and a chloroplast phylogenomic analysis of the Campanulidae

    OpenAIRE

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

    2016-01-01

    Complete chloroplast genome sequences have been very useful for understanding phylogenetic relationships in angiosperms at the family level and above, but there are currently large gaps in coverage. We report the chloroplast genome for Helwingia himalaica, the first in the distinctive family Helwingiaceae and only the second genus to be sequenced in the order Aquifoliales. We then combine this with 36 published sequences in the large (c. 35,000 species) subclass Campanulidae in order to inves...

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

    Directory of Open Access Journals (Sweden)

    Verena Kriechbaumer

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

  5. Insights into the Mechanisms of Chloroplast Division

    Directory of Open Access Journals (Sweden)

    Yamato Yoshida

    2018-03-01

    Full Text Available The endosymbiosis of a free-living cyanobacterium into an ancestral eukaryote led to the evolution of the chloroplast (plastid more than one billion years ago. Given their independent origins, plastid proliferation is restricted to the binary fission of pre-existing plastids within a cell. In the last 25 years, the structure of the supramolecular machinery regulating plastid division has been discovered, and some of its component proteins identified. More recently, isolated plastid-division machineries have been examined to elucidate their structural and mechanistic details. Furthermore, complex studies have revealed how the plastid-division machinery morphologically transforms during plastid division, and which of its component proteins play a critical role in generating the contractile force. Identifying the three-dimensional structures and putative functional domains of the component proteins has given us hints about the mechanisms driving the machinery. Surprisingly, the mechanisms driving plastid division resemble those of mitochondrial division, indicating that these division machineries likely developed from the same evolutionary origin, providing a key insight into how endosymbiotic organelles were established. These findings have opened new avenues of research into organelle proliferation mechanisms and the evolution of organelles.

  6. Polyuridylylation and processing of transcripts from multiple gene minicircles in chloroplasts of the dinoflagellate Amphidinium carterae

    KAUST Repository

    Barbrook, Adrian C.; Dorrell, Richard G.; Burrows, Jennifer; Plenderleith, Lindsey J.; Nisbet, R. Ellen R.; Howe, Christopher J.

    2012-01-01

    -PCR to study transcription and transcript processing in the chloroplasts of Amphidinium carterae, a model peridinin-containing dinoflagellate. These organisms have a highly unusual chloroplast genome, with genes located on multiple small 'minicircle' elements

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

    Directory of Open Access Journals (Sweden)

    Xiang-guang Lv

    Full Text Available A rice chlorophyll-deficient mutant w67 was isolated from an ethyl methane sulfonate (EMS-induced IR64 (Oryza sativa L. ssp. indica mutant bank. The mutant exhibited a distinct yellow-green leaf phenotype in the whole plant growth duration with significantly reduced levels of chlorophyll and carotenoid, impaired chloroplast development and lowered capacity of photosynthesis compared with the wild-type IR64. Expression of a number of genes associated with chlorophyll metabolism, chloroplast biogenesis and photosynthesis was significantly altered in the mutant. Genetic analysis indicated that the yellow-green phenotype was controlled by a single recessive nuclear gene located on the short arm of chromosome 3. Using map-based strategy, the mutation was isolated and predicted to encode a chloroplast signal recognition particle 43 KD protein (cpSRP43 with 388 amino acid residuals. A single base substitution from A to T at position 160 resulted in a premature stop codon. OscpSRP43 was constitutively expressed in various organs with the highest level in the leaf. Functional complementation could rescue the mutant phenotype and subcellular localization showed that the cpSRP43:GFP fusion protein was targeted to the chloroplast. The data suggested that Oryza sativa cpSRP43 (OscpSRP43 was required for the normal development of chloroplasts and photosynthesis in rice.

  8. Conformational changes in spinach (Spinacia oleracea leaves chloroplasts in vivo

    Directory of Open Access Journals (Sweden)

    Janina Godziemba-Czyż

    2015-01-01

    Full Text Available Changes in the surface area of chloroplasts from intact cells of spinach leaves (\tSpinacia oleracea induced by blue (370—500 nm and red (600- 850 nm light of various intensity (102 - 5x105 erg cm-1s-1 were investigated. The changes are deseribed in terms of mean surface area in , μm2 and frequency of oocurrence of surface size classes. Low intensity blue light caused enlargement of the chloroplast surface (as compared with that in darkness, whereas high intensity light markedly reduced it. Exposure of chloroplasts to red light produces an increase of the surface in proportion to the intensity of the light and irradiation time.

  9. Extending the biosynthetic repertoires of cyanobacteria and chloroplasts

    DEFF Research Database (Denmark)

    Nielsen, Agnieszka Janina Zygadlo; Mellor, Silas Busck; Vavitsas, Konstantinos

    2016-01-01

    The chloroplasts found in plants and algae, and photosynthetic microorganisms such as cyanobacteria, are emerging hosts for sustainable production of valuable biochemicals, using only inorganic nutrients, water, CO2 and light as inputs. In the past decade, many bioengineering efforts have focused...... on metabolic engineering and synthetic biology in the chloroplast or in cyanobacteria for the production of fuels, chemicals, as well as complex, high-value bioactive molecules. Biosynthesis of all these compounds can be performed in photosynthetic organelles/organisms by heterologous expression...... of chloroplasts and cyanobacteria as biosynthetic compartments and hosts, and we estimate the production levels to be expected from photosynthetic hosts in light of the fraction of electrons and carbon that can potentially be diverted from photosynthesis. The supply of reducing power, in the form of electrons...

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

    Science.gov (United States)

    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.

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

    Science.gov (United States)

    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.

  12. The action spectrum in chloroplast translocation in multilayer leaf cells

    Directory of Open Access Journals (Sweden)

    Zbigniew Lechowski

    2015-01-01

    Full Text Available By measurement of light transmittance through a leaf as criterion of chloroplast translocation, the action spectrum of Ajuga reptans was established. In the spectrum obtained, a correction was introduced for leaf autoabsorption calculated on the basis of the Beer-Lambert law. The action spectrum has two maxima: at λ= 375 nm and λ= 481 nm. The range above 502 nm has no significant effect on chloroplast translocation. Comparison with other objects examined demonstrated that in multilayer leaf cells riboflavin seems also to be a photoreceptor active in this process.

  13. SKL1 Is Essential for Chloroplast Development in Arabidopsis

    Directory of Open Access Journals (Sweden)

    Huimin Xu

    2018-02-01

    Full Text Available The Arabidopsis shikimate kinase-like 1 (skl1-8 mutant is characterized by a pigment-defective phenotype. Although the related phenotypical defect mainly has been attributed to the blocking of chloroplast development, the molecular functions of SKL1 remain largely unknown. In this study, we combined multiple approaches to investigate the potential functions of SKL1. Results showed that the skl1-8 mutant exhibited an albino phenotype and had dramatically reduced chlorophyll content as a consequence of a single nuclear recessive gene mutation. Chemical complementation analysis indicated that SKL1 does not function as SK enzyme in the shikimate pathway. In addition, by chlorophyll fluorescence parameters and immunoblot analysis, the levels of photosynthetic proteins are substantially reduced. Moreover, by transcriptome analysis, specific groups of nuclear genes involved in photosynthesis, such as light-harvesting complex, pigment metabolism, carbon metabolism, and chloroplast gene expression, were down-regulated, whereas several defense and oxidative stress responsive genes were up-regulated in the skl1-8 mutant compared with the wide type. Furthermore, we found the expression of genes related to auxin transport and response was repressed in the skl1-8 mutant, probable suggesting that SKL1 is involved in auxin-related pathways during chloroplast development. Together, these results provide a useful reference for characterization of SKL1 function during chloroplast biogenesis and development.

  14. Various types of chromoproteins extracted from tobacco chloroplasts

    International Nuclear Information System (INIS)

    Sirchis, Jean; Duranton, Jacques

    1959-01-01

    From tobacco chloroplasts a chroma-proteic complex is isolated; this can be fractionally divided into two different species by the difference in their chemical compositions and their speeds of sedimentation. Reprint of a paper published in 'Comptes Rendus des Seances de l'Academie des Sciences', tome 248, p. 2528-2530, sitting of 27 April 1959 [fr

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

    Science.gov (United States)

    Andrès, Charles; Agne, Birgit; Kessler, Felix

    2010-06-01

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

  16. Extending the biosynthetic repertoires of cyanobacteria and chloroplasts.

    Science.gov (United States)

    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. © 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.

  17. Abscisic acid represses the transcription of chloroplast genes

    Czech Academy of Sciences Publication Activity Database

    Yamburenko, M.V.; Zubo, Y.O.; Vaňková, Radomíra; Kusnetsov, V.; Kulaeva, O.N.; Borner, T.

    2013-01-01

    Roč. 64, č. 14 (2013), s. 4491-4502 ISSN 0022-0957 R&D Projects: GA ČR GA522/09/2058 Institutional research plan: CEZ:AV0Z50380511 Keywords : Abscisic acid (ABA) * chloroplast * cytokinin Subject RIV: ED - Physiology Impact factor: 5.794, year: 2013

  18. Characterization of polymorphic SSRs among Prunus chloroplast genomes

    Science.gov (United States)

    An in silico mining process yielded 80, 75, and 78 microsatellites in the chloroplast genome of Prunus persica, P. kansuensis, and P. mume. A and T repeats were predominant in the three genomes, accounting for 67.8% on average and most of them were successful in primer design. For the 80 P. persica ...

  19. Contribution of chloroplast DNA in the biodiversity of some Aegilops ...

    African Journals Online (AJOL)

    Four Aegilops species (Aegilops longissima, Aegilops speltoides, Aegilops searsii and Aegilops caudata) belonging to the family Poaceae were used in this study. Nucleotides of 1651 bp from 5.8 S rRNA gene and the intergenic spacers trnT-trnL and trnL-trnF from the chloroplast DNA were combined together in order to ...

  20. Functional characterization of recombinant chloroplast signal recognition particle

    NARCIS (Netherlands)

    Groves, M R; Mant, A; Kuhn, A; Koch, J; Dübel, S; Robinson, C; Sinning, I

    2001-01-01

    The signal recognition particle (SRP) is a ubiquitous system for the targeting of membrane and secreted proteins. The chloroplast SRP (cpSRP) is unique among SRPs in that it possesses no RNA and is functional in post-translational as well as co-translational targeting. We have expressed and purified

  1. Protein disorder in plants: a view from the chloroplast

    Directory of Open Access Journals (Sweden)

    Yruela Inmaculada

    2012-09-01

    Full Text Available Abstract Background The intrinsically unstructured state of some proteins, observed in all living organisms, is essential for basic cellular functions. In this field the available information from plants is limited but it has been reached a point where these proteins can be comprehensively classified on the basis of disorder, function and evolution. Results Our analysis of plant genomes confirms that nuclear-encoded proteins follow the same trend than other multi-cellular eukaryotes; however, chloroplast- and mitochondria- encoded proteins conserve the patterns of Archaea and Bacteria, in agreement with their phylogenetic origin. Based on current knowledge about gene transference from the chloroplast to the nucleus, we report a strong correlation between the rate of disorder of transferred and nuclear-encoded proteins, even for polypeptides that play functional roles back in the chloroplast. We further investigate this trend by reviewing the set of chloroplast ribosomal proteins, one of the most representative transferred gene clusters, finding that the ribosomal large subunit, assembled from a majority of nuclear-encoded proteins, is clearly more unstructured than the small one, which integrates mostly plastid-encoded proteins. Conclusions Our observations suggest that the evolutionary dynamics of the plant nucleus adds disordered segments to genes alike, regardless of their origin, with the notable exception of proteins currently encoded in both genomes, probably due to functional constraints.

  2. Engineering the Chloroplast Genome of Oleaginous Marine Microalga Nannochloropsis oceanica

    Directory of Open Access Journals (Sweden)

    Qinhua Gan

    2018-04-01

    Full Text Available Plastid engineering offers an important tool to fill the gap between the technical and the enormous potential of microalgal photosynthetic cell factory. However, to date, few reports on plastid engineering in industrial microalgae have been documented. This is largely due to the small cell sizes and complex cell-wall structures which make these species intractable to current plastid transformation methods (i.e., biolistic transformation and polyethylene glycol-mediated transformation. Here, employing the industrial oleaginous microalga Nannochloropsis oceanica as a model, an electroporation-mediated chloroplast transformation approach was established. Fluorescent microscopy and laser confocal scanning microscopy confirmed the expression of the green fluorescence protein, driven by the endogenous plastid promoter and terminator. Zeocin-resistance selection led to an acquisition of homoplasmic strains of which a stable and site-specific recombination within the chloroplast genome was revealed by sequencing and DNA gel blotting. This demonstration of electroporation-mediated chloroplast transformation opens many doors for plastid genome editing in industrial microalgae, particularly species of which the chloroplasts are recalcitrant to chemical and microparticle bombardment transformation.

  3. Expression of recombinant interferon α-2a in tobacco chloroplasts ...

    African Journals Online (AJOL)

    Chloroplast transformation was accomplished upon bombardment of fully expanded 4 to 6 weeks-old tobacco leaves using helium gun. Green shoots regenerated from single antibiotic resistant cells were subjected to further rounds of selection and regeneration to develop homoplasmic clones. The molecular analysis of ...

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

    Directory of Open Access Journals (Sweden)

    Vivien eRolland

    2016-02-01

    Full Text Available 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.

  5. Manipulation of Glutathione and Amino Acid Biosynthesis in the Chloroplast1

    Science.gov (United States)

    Noctor, Graham; Arisi, Ana-Carolina M.; Jouanin, Lise; Foyer, Christine H.

    1998-01-01

    Poplars (Populus tremula × Populus alba) were transformed to overexpress Escherichia coli γ-glutamylcysteine synthetase (γ-ECS) or glutathione synthetase in the chloroplast. Five independent lines of each transformant strongly expressed the introduced gene and possessed markedly enhanced activity of the gene product. Glutathione (GSH) contents were unaffected by high chloroplastic glutathione synthetase activity. Enhanced chloroplastic γ-ECS activity markedly increased γ-glutamylcysteine and GSH levels. These effects are similar to those previously observed in poplars overexpressing these enzymes in the cytosol. Similar to cytosolic γ-ECS overexpression, chloroplastic overexpression did not deplete foliar cysteine or methionine pools and did not lead to morphological changes. Light was required for maximal accumulation of GSH in poplars overexpressing γ-ECS in the chloroplast. High chloroplastic, but not cytosolic, γ-ECS activities were accompanied by increases in amino acids synthesized in the chloroplast. We conclude that (a) GSH synthesis can occur in the chloroplast and the cytosol and may be up-regulated in both compartments by increased γ-ECS activity, (b) interactions between GSH synthesis and the pathways supplying the necessary substrates are similar in both compartments, and (c) chloroplastic up-regulation of GSH synthesis is associated with an activating effect on the synthesis of specific amino acids formed in the chloroplast. PMID:9765532

  6. Effects and mechanism of acid rain on plant chloroplast ATP synthase.

    Science.gov (United States)

    Sun, Jingwen; Hu, Huiqing; Li, Yueli; Wang, Lihong; Zhou, Qing; Huang, Xiaohua

    2016-09-01

    Acid rain can directly or indirectly affect plant physiological functions, especially photosynthesis. The enzyme ATP synthase is the key in photosynthetic energy conversion, and thus, it affects plant photosynthesis. To clarify the mechanism by which acid rain affects photosynthesis, we studied the effects of acid rain on plant growth, photosynthesis, chloroplast ATP synthase activity and gene expression, chloroplast ultrastructure, intracellular H(+) level, and water content of rice seedlings. Acid rain at pH 4.5 remained the chloroplast structure unchanged but increased the expression of six chloroplast ATP synthase subunits, promoted chloroplast ATP synthase activity, and increased photosynthesis and plant growth. Acid rain at pH 4.0 or less decreased leaf water content, destroyed chloroplast structure, inhibited the expression of six chloroplast ATP synthase subunits, decreased chloroplast ATP synthase activity, and reduced photosynthesis and plant growth. In conclusion, acid rain affected the chloroplast ultrastructure, chloroplast ATPase transcription and activity, and P n by changing the acidity in the cells, and thus influencing the plant growth and development. Finally, the effects of simulated acid rain on the test indices were found to be dose-dependent.

  7. Analysis of chlorophyll fluorescence reveals stage specific patterns of chloroplast-containing cells during Arabidopsis embryogenesis

    Directory of Open Access Journals (Sweden)

    RICARDO I TEJOS

    2010-01-01

    Full Text Available The basic body plan of a plant is established early in embryogenesis when cells differentiate, giving rise to the apical and basal regions of the embryo. Using chlorophyll fluorescence as a marker for chloroplasts, we have detected specific patterns of chloroplast-containing cells at specific stages of embryogenesis. Non-randomly distributed chloroplast-containing cells are seen as early as the globular stage of embryogenesis in Arabidopsis. In the heart stage of embryogenesis, chloroplast containing cells are detected in epidermal cells as well as a central region of the heart stage embryo, forming a triangular septum of chloroplast-containing cells that divides the embryo into three equal sectors. Torpedo stage embryos have chloroplast-containing epidermal cells and a central band of chloroplast-containing cells in the cortex layer, just below the shoot apical meristem. In the walking-stick stage of embryogenesis, chloroplasts are present in the epidermal, cortex and endodermal cells. The chloroplasts appear reduced or absent from the provascular and columella cells of walking-stick stage embryos. These results suggest that there is a tight regulation of plastid differentiation during embryogenesis that generates specific patterns of chloroplast-containing cells in specific cell layers at specific stages of embryogenesis.

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

    Directory of Open Access Journals (Sweden)

    Sławomir Samardakiewicz

    Full Text Available Lead ions are particularly dangerous to the photosynthetic apparatus, but little is known about the effects of trace metals, including Pb, on regulation of chloroplast redistribution. In this study a new effect of lead on chloroplast distribution patterns and movements was demonstrated in mesophyll cells of a small-sized aquatic angiosperm Lemna trisulca L. (star duckweed. An analysis of confocal microscopy images of L. trisulca fronds treated with lead (15 μM Pb2+, 24 h in darkness or in weak white light revealed an enhanced accumulation of chloroplasts in the profile position along the anticlinal cell walls, in comparison to untreated plants. The rearrangement of chloroplasts in their response to lead ions in darkness was similar to the avoidance response of chloroplasts in plants treated with strong white light. Transmission electron microscopy X-ray microanalysis showed that intracellular chloroplast arrangement was independent of the location of Pb deposits, suggesting that lead causes redistribution of chloroplasts, which looks like a light-induced avoidance response, but is not a real avoidance response to the metal. Furthermore, a similar redistribution of chloroplasts in L. trisulca cells in darkness was observed also under the influence of exogenously applied hydrogen peroxide (H2O2. In addition, we detected an enhanced accumulation of endogenous H2O2 after treatment of plants with lead. Interestingly, H2O2-specific scavenger catalase partly abolished the Pb-induced chloroplast response. These results suggest that H2O2 can be involved in the avoidance-like movement of chloroplasts induced by lead. Analysis of photometric measurements revealed also strong inhibition (but not complete of blue-light-induced chloroplast movements by lead. This inhibition may result from disturbances in the actin cytoskeleton, as we observed fragmentation and disappearance of actin filaments around chloroplasts. Results of this study show that the

  9. Chloroplast Dynamics and Photosynthetic Efficiency: Final Technical Report

    Energy Technology Data Exchange (ETDEWEB)

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

    2016-11-03

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

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

    Energy Technology Data Exchange (ETDEWEB)

    Rodermel, Steven

    2015-11-16

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

  11. Chloroplast Genome Evolution in Early Diverged Leptosporangiate Ferns

    OpenAIRE

    Kim, Hyoung Tae; Chung, Myong Gi; Kim, Ki-Joong

    2014-01-01

    In this study, the chloroplast (cp) genome sequences from three early diverged leptosporangiate ferns were completed and analyzed in order to understand the evolution of the genome of the fern lineages. The complete cp genome sequence of Osmunda cinnamomea (Osmundales) was 142,812 base pairs (bp). The cp genome structure was similar to that of eusporangiate ferns. The gene/intron losses that frequently occurred in the cp genome of leptosporangiate ferns were not found in the cp genome of O. c...

  12. Biparental chloroplast inheritance leads to rescue from cytonuclear incompatibility.

    Science.gov (United States)

    Barnard-Kubow, Karen B; McCoy, Morgan A; Galloway, Laura F

    2017-02-01

    Although organelle inheritance is predominantly maternal across animals and plants, biparental chloroplast inheritance has arisen multiple times in the angiosperms. Biparental inheritance has the potential to impact the evolutionary dynamics of cytonuclear incompatibility, interactions between nuclear and organelle genomes that are proposed to be among the earliest types of genetic incompatibility to arise in speciation. We examine the interplay between biparental inheritance and cytonuclear incompatibility in Campanulastrum americanum, a plant species exhibiting both traits. We first determine patterns of chloroplast inheritance in genetically similar and divergent crosses, and then associate inheritance with hybrid survival across multiple generations. There is substantial biparental inheritance in C. americanum. The frequency of biparental inheritance is greater in divergent crosses and in the presence of cytonuclear incompatibility. Biparental inheritance helps to mitigate cytonuclear incompatibility, leading to increased fitness of F 1 hybrids and recovery in the F 2 generation. This study demonstrates the potential for biparental chloroplast inheritance to rescue cytonuclear compatibility, reducing cytonuclear incompatibility's contribution to reproductive isolation and potentially slowing speciation. The efficacy of rescue depended upon the strength of incompatibility, with a greater persistence of weak incompatibilities in later generations. These findings suggest that incompatible plastids may lead to selection for biparental inheritance. © 2016 The Authors. New Phytologist © 2016 New Phytologist Trust.

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

    Science.gov (United States)

    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.

  14. Exploring photosynthesis evolution by comparative analysis of metabolic networks between chloroplasts and photosynthetic bacteria

    Directory of Open Access Journals (Sweden)

    Hou Jing

    2006-04-01

    Full Text Available Abstract Background Chloroplasts descended from cyanobacteria and have a drastically reduced genome following an endosymbiotic event. Many genes of the ancestral cyanobacterial genome have been transferred to the plant nuclear genome by horizontal gene transfer. However, a selective set of metabolism pathways is maintained in chloroplasts using both chloroplast genome encoded and nuclear genome encoded enzymes. As an organelle specialized for carrying out photosynthesis, does the chloroplast metabolic network have properties adapted for higher efficiency of photosynthesis? We compared metabolic network properties of chloroplasts and prokaryotic photosynthetic organisms, mostly cyanobacteria, based on metabolic maps derived from genome data to identify features of chloroplast network properties that are different from cyanobacteria and to analyze possible functional significance of those features. Results The properties of the entire metabolic network and the sub-network that consists of reactions directly connected to the Calvin Cycle have been analyzed using hypergraph representation. Results showed that the whole metabolic networks in chloroplast and cyanobacteria both possess small-world network properties. Although the number of compounds and reactions in chloroplasts is less than that in cyanobacteria, the chloroplast's metabolic network has longer average path length, a larger diameter, and is Calvin Cycle -centered, indicating an overall less-dense network structure with specific and local high density areas in chloroplasts. Moreover, chloroplast metabolic network exhibits a better modular organization than cyanobacterial ones. Enzymes involved in the same metabolic processes tend to cluster into the same module in chloroplasts. Conclusion In summary, the differences in metabolic network properties may reflect the evolutionary changes during endosymbiosis that led to the improvement of the photosynthesis efficiency in higher plants. Our

  15. Phosphorus compounds, proteins, nuclease and acid phosphatase activities in isolated spinach chloroplasts

    Directory of Open Access Journals (Sweden)

    E. Mikulska

    2015-01-01

    Full Text Available This paper deals with attempts to elaborate a simple method of spinach chloroplast isolation ensuring a high proportion of intact chloroplasts. We obtained 3 preparations of isolated chloroplasts. Several preliminary analyses of the obtained chloroplast fraction were also performed. Phosphorus compounds, total protein and the enzyme activities of RNase, DNase and GPase were determined. We found: 0,36-0,59% of RNA, 0,19-0,24% of DNA, 2,1-2,9% of phospholipids and 26-28% of protein. RNase activity was very high.

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

    Energy Technology Data Exchange (ETDEWEB)

    Angelova, Angelina [University of Arizona; Park, Sang-Hycuk [University of Arizona; Kyndt, John [Bellevue University; Fitzsimmons, Kevin [University of Arizona; Brown, Judith K [University of Arizona

    2013-09-01

    With the increasing world demand for biofuel, a number of oleaginous algal species are being considered as renewable sources of oil. Chlorella protothecoides Krüger synthesizes triacylglycerols (TAGs) as storage compounds that can be converted into renewable fuel utilizing an anabolic pathway that is poorly understood. The paucity of algal chloroplast genome sequences has been an important constraint to chloroplast transformation and for studying gene expression in TAGs pathways. In this study, the intact chloroplasts were released from algal cells using sonication followed by sucrose gradient centrifugation, resulting in a 2.36-fold enrichment of chloroplasts from C. protothecoides, based on qPCR analysis. The C. protothecoides chloroplast genome (cpDNA) was determined using the Illumina HiSeq 2000 sequencing platform and found to be 84,576 Kb in size (8.57 Kb) in size, with a GC content of 30.8 %. This is the first report of an optimized protocol that uses a sonication step, followed by sucrose gradient centrifugation, to release and enrich intact chloroplasts from a microalga (C. prototheocoides) of sufficient quality to permit chloroplast genome sequencing with high coverage, while minimizing nuclear genome contamination. The approach is expected to guide chloroplast isolation from other oleaginous algal species for a variety of uses that benefit from enrichment of chloroplasts, ranging from biochemical analysis to genomics studies.

  17. Does a voltage-sensitive outer envelope transport mechanism contributes to the chloroplast iron uptake?

    Science.gov (United States)

    Solti, Ádám; Kovács, Krisztina; Müller, Brigitta; Vázquez, Saúl; Hamar, Éva; Pham, Hong Diep; Tóth, Brigitta; Abadía, Javier; Fodor, Ferenc

    2016-12-01

    Based on the effects of inorganic salts on chloroplast Fe uptake, the presence of a voltage-dependent step is proposed to play a role in Fe uptake through the outer envelope. Although iron (Fe) plays a crucial role in chloroplast physiology, only few pieces of information are available on the mechanisms of chloroplast Fe acquisition. Here, the effect of inorganic salts on the Fe uptake of intact chloroplasts was tested, assessing Fe and transition metal uptake using bathophenantroline-based spectrophotometric detection and plasma emission-coupled mass spectrometry, respectively. The microenvironment of Fe was studied by Mössbauer spectroscopy. Transition metal cations (Cd 2+ , Zn 2+ , and Mn 2+ ) enhanced, whereas oxoanions (NO 3 - , SO 4 2- , and BO 3 3- ) reduced the chloroplast Fe uptake. The effect was insensitive to diuron (DCMU), an inhibitor of chloroplast inner envelope-associated Fe uptake. The inorganic salts affected neither Fe forms in the uptake assay buffer nor those incorporated into the chloroplasts. The significantly lower Zn and Mn uptake compared to that of Fe indicates that different mechanisms/transporters are involved in their acquisition. The enhancing effect of transition metals on chloroplast Fe uptake is likely related to outer envelope-associated processes, since divalent metal cations are known to inhibit Fe 2+ transport across the inner envelope. Thus, a voltage-dependent step is proposed to play a role in Fe uptake through the chloroplast outer envelope on the basis of the contrasting effects of transition metal cations and oxoaninons.

  18. Oxidative damage to chloroplasts from Chlorella vulgaris exposed to ultraviolet-B radiation

    International Nuclear Information System (INIS)

    Malanga, G.; Calmanovici, G.; Puntarulo, S.

    1997-01-01

    Upon UV-B irradiation, Chlorella vulgaris cells and isolated chloroplasts increased in size and starch accumulation. Photosynthetic capacity and chlorophyll content of chloroplasts isolated from irradiated algae decreased by 72 and 66%, as compared to chloroplasts isolated from control cells. Dihydrorhodamine 123 conversion to rhodamine 123 was used as a sensitive method for detection of peroxide (presumably hydrogen peroxide) formation in isolated chloroplasts. The accumulation of rhodamine 123 is higher in irradiated than in nonirradiated chloroplasts and the increased accumulation of rhodamine 123 depended on the UV-B dose. Quantitation of alkyl radical-EPR signals in chloroplasts indicated that UV-B exposure significantly increased radical content in the membranes. The content of an oxidized DNA base (8-hydroxy-2′-deoxyguanosine) in chloroplasts was increased by 72 and 175% after irradiation of the algal culture with 17.3 and 42.6 kJ m −2 , respectively. The chloroplastic activity of superoxide dismutase decreased by 50% as compared with control values after irradiation with 42.6 kJ m −2 and no changes in ascorbate peroxidase activity and ascorbic acid content were detected at the irradiation doses tested. The β-carotene content in chloroplasts was not affected by the irradiation, but the α-tocopherol content increased approximately 4-fold after UV-B irradiation. The results suggest that oxidative damage related to UV-B exposure is responsible for alterations in chloroplasts function and integrity, and that an antioxidant response is triggered in chloroplasts through an increase in α-tocopherol content. (author)

  19. GENETIC POLYMORPHISM IN GYMNODINIUM GALATHEANUM CHLOROPLAST DNA SEQUENCES AND DEVELOPMENT OF A MOLECULAR DETECTION ASSAY. (R827084)

    Science.gov (United States)

    Nuclear and chloroplast-encoded small subunit ribosomal DNA sequences were obtainedfrom several strains of the toxic dinoflagellate Gymnodinium galatheanum. Phylogenetic analyses andcomparison of sequences indicate that the chloroplast sequences show a higher degree of se...

  20. De Novo Assembly of Complete Chloroplast Genomes from Non-model Species Based on a K-mer Frequency-Based Selection of Chloroplast Reads from total DNA Sequences.

    NARCIS (Netherlands)

    Izan, Shairul; Esselink, G.; Visser, R.G.F.; Smulders, M.J.M.; Borm, T.J.A.

    2017-01-01

    Whole Genome Shotgun (WGS) sequences of plant species often contain an abundance of reads that are derived from the chloroplast genome. Up to now these reads have generally been identified and assembled into chloroplast genomes based on homology to chloroplasts from related species. This

  1. Ion and metabolite transport in the chloroplast of algae: lessons from land plants.

    Science.gov (United States)

    Marchand, Justine; Heydarizadeh, Parisa; Schoefs, Benoît; Spetea, Cornelia

    2018-06-01

    Chloroplasts are endosymbiotic organelles and play crucial roles in energy supply and metabolism of eukaryotic photosynthetic organisms (algae and land plants). They harbor channels and transporters in the envelope and thylakoid membranes, mediating the exchange of ions and metabolites with the cytosol and the chloroplast stroma and between the different chloroplast subcompartments. In secondarily evolved algae, three or four envelope membranes surround the chloroplast, making more complex the exchange of ions and metabolites. Despite the importance of transport proteins for the optimal functioning of the chloroplast in algae, and that many land plant homologues have been predicted, experimental evidence and molecular characterization are missing in most cases. Here, we provide an overview of the current knowledge about ion and metabolite transport in the chloroplast from algae. The main aspects reviewed are localization and activity of the transport proteins from algae and/or of homologues from other organisms including land plants. Most chloroplast transporters were identified in the green alga Chlamydomonas reinhardtii, reside in the envelope and participate in carbon acquisition and metabolism. Only a few identified algal transporters are located in the thylakoid membrane and play role in ion transport. The presence of genes for putative transporters in green algae, red algae, diatoms, glaucophytes and cryptophytes is discussed, and roles in the chloroplast are suggested. A deep knowledge in this field is required because algae represent a potential source of biomass and valuable metabolites for industry, medicine and agriculture.

  2. Study of the interaction of cytochrome c and ferredoxine with the double membrane of chloroplast

    International Nuclear Information System (INIS)

    Neuburger, M.; Joyard, J.; Douce, R.

    1975-01-01

    The adsorption of two 59 Fe-labelled proteins on the chloroplast envelope was studied. The former molecule used was ferredoxine extracted from spinach leaves, the latter was cytochrome c, extracted from yeast (Saccharomyces cerevisiae D 261). The chloroplast envelope is thought to be involved in the transport of some proteins such as ferredoxine synthetized in the cytoplasm [fr

  3. Chloroplast microsatellites reveal population genetic diversity in red pine, Pinus resinosa Ait

    Science.gov (United States)

    Craig S. Echt; L.L. DeVerno; M. Anzidei; G.G. Vendramin

    1998-01-01

    Variation in paternally inherited chloroplast microsatellite (cpSSR) DNA was used to study population genetic structure in red pine (Pinus resinosa Ait.), a species characterized by morphological uniformity, no allozyme variation, and limited RAPD variation. Using nine cpSSR loci, a total of 23 chloroplast haplotypes and 25 cpSSR alleles were were...

  4. The diurnal logic of the expression of the chloroplast genome in Chlamydomonas reinhardtii.

    Directory of Open Access Journals (Sweden)

    Adam D Idoine

    Full Text Available Chloroplasts are derived from cyanobacteria and have retained a bacterial-type genome and gene expression machinery. The chloroplast genome encodes many of the core components of the photosynthetic apparatus in the thylakoid membranes. To avoid photooxidative damage and production of harmful reactive oxygen species (ROS by incompletely assembled thylakoid protein complexes, chloroplast gene expression must be tightly regulated and co-ordinated with gene expression in the nucleus. Little is known about the control of chloroplast gene expression at the genome-wide level in response to internal rhythms and external cues. To obtain a comprehensive picture of organelle transcript levels in the unicellular model alga Chlamydomonas reinhardtii in diurnal conditions, a qRT-PCR platform was developed and used to quantify 68 chloroplast, 21 mitochondrial as well as 71 nuclear transcripts in cells grown in highly controlled 12 h light/12 h dark cycles. Interestingly, in anticipation of dusk, chloroplast transcripts from genes involved in transcription reached peak levels first, followed by transcripts from genes involved in translation, and finally photosynthesis gene transcripts. This pattern matches perfectly the theoretical demands of a cell "waking up" from the night. A similar trend was observed in the nuclear transcripts. These results suggest a striking internal logic in the expression of the chloroplast genome and a previously unappreciated complexity in the regulation of chloroplast genes.

  5. Genetic analysis of a Microseris douglasii (Asteraceae) population polymorphic for an alien chloroplast type

    NARCIS (Netherlands)

    Roelofs, Dick; Bachmann, Konrad

    1997-01-01

    Recent evidence suggests chloroplast introgression from Microseris bigelovii into M. douglasii. We have examined 23 plants from a population of M. douglasii polymorphic for M. douglasii and M. bigelovii chloroplast types. All 23 plants were completely homozygous for morphological and RAPD markers,

  6. Effect of alkyl-N-phenylcarbamates on photochemical activity of spinach chloroplasts

    International Nuclear Information System (INIS)

    Sersen, F.; Kralova, K.; Macho, V.

    1999-01-01

    This study is aimed to investigate the effect of alkyl-N-phenylcarbamates on photosynthetic electron transport in spinach chloroplasts, to determine site of action in the photosynthetic apparatus of spinach chloroplasts and to find correlations between their structure and biological activity. (authors)

  7. Characterization of mango (Mangifera indica L.) transcriptome and chloroplast genome.

    Science.gov (United States)

    Azim, M Kamran; Khan, Ishtaiq A; Zhang, Yong

    2014-05-01

    We characterized mango leaf transcriptome and chloroplast genome using next generation DNA sequencing. The RNA-seq output of mango transcriptome generated >12 million reads (total nucleotides sequenced >1 Gb). De novo transcriptome assembly generated 30,509 unigenes with lengths in the range of 300 to ≥3,000 nt and 67× depth of coverage. Blast searching against nonredundant nucleotide databases and several Viridiplantae genomic datasets annotated 24,593 mango unigenes (80% of total) and identified Citrus sinensis as closest neighbor of mango with 9,141 (37%) matched sequences. The annotation with gene ontology and Clusters of Orthologous Group terms categorized unigene sequences into 57 and 25 classes, respectively. More than 13,500 unigenes were assigned to 293 KEGG pathways. Besides major plant biology related pathways, KEGG based gene annotation pointed out active presence of an array of biochemical pathways involved in (a) biosynthesis of bioactive flavonoids, flavones and flavonols, (b) biosynthesis of terpenoids and lignins and (c) plant hormone signal transduction. The mango transcriptome sequences revealed 235 proteases belonging to five catalytic classes of proteolytic enzymes. The draft genome of mango chloroplast (cp) was obtained by a combination of Sanger and next generation sequencing. The draft mango cp genome size is 151,173 bp with a pair of inverted repeats of 27,093 bp separated by small and large single copy regions, respectively. Out of 139 genes in mango cp genome, 91 found to be protein coding. Sequence analysis revealed cp genome of C. sinensis as closest neighbor of mango. We found 51 short repeats in mango cp genome supposed to be associated with extensive rearrangements. This is the first report of transcriptome and chloroplast genome analysis of any Anacardiaceae family member.

  8. Analysis of Acorus calamus chloroplast genome and its phylogenetic implications.

    Science.gov (United States)

    Goremykin, Vadim V; Holland, Barbara; Hirsch-Ernst, Karen I; Hellwig, Frank H

    2005-09-01

    Determining the phylogenetic relationships among the major lines of angiosperms is a long-standing problem, yet the uncertainty as to the phylogenetic affinity of these lines persists. While a number of studies have suggested that the ANITA (Amborella-Nymphaeales-Illiciales-Trimeniales-Aristolochiales) grade is basal within angiosperms, studies of complete chloroplast genome sequences also suggested an alternative tree, wherein the line leading to the grasses branches first among the angiosperms. To improve taxon sampling in the existing chloroplast genome data, we sequenced the chloroplast genome of the monocot Acorus calamus. We generated a concatenated alignment (89,436 positions for 15 taxa), encompassing almost all sequences usable for phylogeny reconstruction within spermatophytes. The data still contain support for both the ANITA-basal and grasses-basal hypotheses. Using simulations we can show that were the ANITA-basal hypothesis true, parsimony (and distance-based methods with many models) would be expected to fail to recover it. The self-evident explanation for this failure appears to be a long-branch attraction (LBA) between the clade of grasses and the out-group. However, this LBA cannot explain the discrepancies observed between tree topology recovered using the maximum likelihood (ML) method and the topologies recovered using the parsimony and distance-based methods when grasses are deleted. Furthermore, the fact that neither maximum parsimony nor distance methods consistently recover the ML tree, when according to the simulations they would be expected to, when the out-group (Pinus) is deleted, suggests that either the generating tree is not correct or the best symmetric model is misspecified (or both). We demonstrate that the tree recovered under ML is extremely sensitive to model specification and that the best symmetric model is misspecified. Hence, we remain agnostic regarding phylogenetic relationships among basal angiosperm lineages.

  9. Biotin Carboxyl Carrier Protein in Barley Chloroplast Membranes

    DEFF Research Database (Denmark)

    Kannangara, C. G.; Jense, C J

    1975-01-01

    Biotin localized in barley chloroplast lamellae is covalently bound to a single protein with an approximate molecular weight of 21000. It contains one mole of biotin per mole of protein and functions as a carboxyl carrier in the acetyl-CoA carboxylase reaction. The protein was obtained by solubil...... by solubilization of the lamellae in phenol/acetic acid/8 M urea. Feeding barley seedlings with [14C]-biotin revealed that the vitamin is not degraded into respiratory substrates by the plant, but is specifically incorporated into biotin carboxyl carrier protein....

  10. Chloroplast Movement May Impact Plant Phenotyping and Photochemistry Results

    Science.gov (United States)

    Malas, J.; Pleban, J. R.; Wang, D. R.; Riley, C.; Mackay, D. S.

    2017-12-01

    Investigating phenotypic responses of crop species across environmental conditions is vital to improving agricultural productivity. Crop production is closely linked with photosynthetic activity, which can be evaluated using parameters such as relative chlorophyll, SPAD, and variable chlorophyll fluorescence. Recently, a handheld device known as the MultispeQ emerged on the market as an open-source instrument that aims to provide high-output, high-quality field data at a low cost to the plant research community. MultispeQ takes measurements of both environmental conditions (light intensity, temperature, humidity, etc.) and photosynthetic parameters (relative chlorophyll, SPAD, photosystem II quantum efficiency (FII), and non-photochemical quenching (NPQ)). Data are automatically backed up and shared on the PhotosynQ network, which serves as a collaborative platform for researchers and professionals. Here, we used the instrument to quantify photosynthetic time-courses of two Brassica rapa genotypes in response to two contrasting nutrient management strategies (Control; High Nitrogen). Previous research found that chloroplast movement is one strategy plants use to optimize photosynthesis across varying light conditions. We were able to detect chloroplast movement throughout the day using the MultispeQ device. Our results support the idea that chloroplast movement serves both as an intrinsic feature of the circadian clock and as a light avoidance strategy. Under low light conditions (PAR 0-300) more light at the near-infrared and red regions was absorbed than under higher light conditions (PAR 500-800). In one genotype by treatment combination, absorbance at 730nm was around 60% at low light, versus only 30% at high light conditions. In light of our results that relative chlorophyll may change throughout a day, we suggest that it is important to take note of these effects when collecting photosynthesis efficiency data in order to avoid bias in measurements. We also

  11. The complete chloroplast genome sequence of Hibiscus syriacus.

    Science.gov (United States)

    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.

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

    Science.gov (United States)

    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.

  13. The complete chloroplast genome sequence of Dendrobium officinale.

    Science.gov (United States)

    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.

  14. Hartmut Lichtenthaler: an authority on chloroplast structure and isoprenoid biochemistry.

    Science.gov (United States)

    Sharkey, Thomas D; Govindjee

    2016-05-01

    We pay tribute to Hartmut Lichtenthaler for making important contributions to the field of photosynthesis research. He was recently recognized for ground-breaking discoveries in chloroplast structure and isoprenoid biochemistry by the Rebeiz Foundation for Basic Research (RFBR; http://vlpbp.org/ ), receiving a 2014 Lifetime Achievement Award for Photosynthesis. The ceremony, held in Champaign, Illinois, was attended by many prominent researchers in the photosynthesis field. We provide below a brief note on his education, and then describe some of the areas in which Hartmut Lichtenthaler has been a pioneer.

  15. Carbon Dioxide Fixation in Isolated Kalanchoe Chloroplasts 1

    Science.gov (United States)

    Levi, Carolyn; Gibbs, Martin

    1975-01-01

    Chloroplasts isolated from Kalanchoe diagremontiana leaves were capable of photosynthesizing at a rate of 5.4 μmoles of CO2 per milligram of chlorophyll per hour. The dark rate of fixation was about 1% of the light rate. A high photosynthetic rate was associated with low starch content of the leaves. Ribose 5-phosphate, fructose 1,6-diphosphate, and dithiothreitol stimulated fixation, whereas phosphoenolpyruvate and azide were inhibitors. The products of CO2 fixation were primarily those of the photosynthetic carbon reduction cycle. PMID:16659249

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

    Science.gov (United States)

    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.

  17. An Arabidopsis chloroplast-targeted Hsp101 homologue, APG6, has an essential role in chloroplast development as well as heat-stress response.

    Science.gov (United States)

    Myouga, Fumiyoshi; Motohashi, Reiko; Kuromori, Takashi; Nagata, Noriko; Shinozaki, Kazuo

    2006-10-01

    Analysis of albino or pale-green (apg) mutants is important for identifying nuclear genes responsible for chloroplast development and pigment synthesis. We have identified 38 apg mutants by screening 11 000 Arabidopsis Ds-tagged lines. One mutant, apg6, contains a Ds insertion in a gene encoding APG6 (ClpB3), a homologue of the heat-shock protein Hsp101 (ClpB1). We isolated somatic revertants and identified two Ds-tagged and one T-DNA-tagged mutant alleles of apg6. All three alleles gave the same pale-green phenotype. These results suggest that APG6 is important for chloroplast development. The APG6 protein contains a transit peptide and is localized in chloroplasts. The plastids of apg6 pale-green cells were smaller than those of the wild type, and contained undeveloped thylakoid membranes. APG6 mRNA accumulated in response to heat shock in various organs, but not in response to other abiotic stresses. Under normal conditions, APG6 is constitutively expressed in the root tips, the organ boundary region, the reproductive tissues of mature plants where plastids exist as proplastids, and slightly in the stems and leaves. In addition, constitutive overexpression of APG6 in transgenic plants inhibited chloroplast development and resulted in a mild pale-green phenotype. The amounts of chloroplast proteins related to photosynthesis were markedly decreased in apg6 mutants. These results suggest that APG6 functions as a molecular chaperone involved in plastid differentiation mediating internal thylakoid membrane formation and conferring thermotolerance to chloroplasts during heat stress. The APG6 protein is not only involved in heat-stress response in chloroplasts, but is also essential for chloroplast development.

  18. PDV2 has a dosage effect on chloroplast division in Arabidopsis.

    Science.gov (United States)

    Chang, Ning; Sun, Qingqing; Li, Yiqiong; Mu, Yajuan; Hu, Jinglei; Feng, Yue; Liu, Xiaomin; Gao, Hongbo

    2017-03-01

    PDV2 has a dosage effect on chloroplast division in Arabidopsis thaliana , but this effect may vary in different plants. Chloroplasts have to be divided as plants grow to maintain an optimized number in the cell. Chloroplasts are divided by protein complexes across the double membranes from the stroma side to the cytosolic side. PDV2 is a chloroplast division protein on the chloroplast outer membrane. It recruits the dynamin-related GTPase ARC5 to the division site. The C-terminus of PDV2 and the C-terminus of ARC6 interact in the intermembrane space, which is important for the localization of PDV2. Previously, it was shown that overexpression of PDV2 can increase the division of chloroplasts in Arabidopsis and moss, so the authors concluded that PDV2 determines the rate of chloroplast division in land plants. PDV2 was also shown to inhibit the GTPase activity of ARC5 by in vitro experiment. These results look to be contradictory. Here, we identified a null allele of PDV2 in Arabidopsis and studied plants with different levels of PDV2. Our results suggested that the chloroplast division phenotype in Arabidopsis is sensitive to the level of PDV2, while this is not the case for ARC6. The level of PDV2 protein is reduced sharply in fast-growing leaves, while the level of ARC6 is not. The levels of PDV2 and ARC6 in several other plant species at different developmental stages were also investigated. The results indicated that their expression pattern varies in different species. Thus, PDV2 is an important positive factor of chloroplast division with an apparent dosage effect in Arabidopsis, but this effect for different chloroplast division proteins in different plants may vary.

  19. The complete chloroplast genome of banana (Musa acuminata, Zingiberales): insight into plastid monocotyledon evolution.

    Science.gov (United States)

    Martin, Guillaume; Baurens, Franc-Christophe; Cardi, Céline; Aury, Jean-Marc; D'Hont, Angélique

    2013-01-01

    Banana (genus Musa) is a crop of major economic importance worldwide. It is a monocotyledonous member of the Zingiberales, a sister group of the widely studied Poales. Most cultivated bananas are natural Musa inter-(sub-)specific triploid hybrids. A Musa acuminata reference nuclear genome sequence was recently produced based on sequencing of genomic DNA enriched in nucleus. The Musa acuminata chloroplast genome was assembled with chloroplast reads extracted from whole-genome-shotgun sequence data. The Musa chloroplast genome is a circular molecule of 169,972 bp with a quadripartite structure containing two single copy regions, a Large Single Copy region (LSC, 88,338 bp) and a Small Single Copy region (SSC, 10,768 bp) separated by Inverted Repeat regions (IRs, 35,433 bp). Two forms of the chloroplast genome relative to the orientation of SSC versus LSC were found. The Musa chloroplast genome shows an extreme IR expansion at the IR/SSC boundary relative to the most common structures found in angiosperms. This expansion consists of the integration of three additional complete genes (rps15, ndhH and ycf1) and part of the ndhA gene. No such expansion has been observed in monocots so far. Simple Sequence Repeats were identified in the Musa chloroplast genome and a new set of Musa chloroplastic markers was designed. The complete sequence of M. acuminata ssp malaccensis chloroplast we reported here is the first one for the Zingiberales order. As such it provides new insight in the evolution of the chloroplast of monocotyledons. In particular, it reinforces that IR/SSC expansion has occurred independently several times within monocotyledons. The discovery of new polymorphic markers within Musa chloroplast opens new perspectives to better understand the origin of cultivated triploid bananas.

  20. The complete chloroplast genome of banana (Musa acuminata, Zingiberales: insight into plastid monocotyledon evolution.

    Directory of Open Access Journals (Sweden)

    Guillaume Martin

    Full Text Available Banana (genus Musa is a crop of major economic importance worldwide. It is a monocotyledonous member of the Zingiberales, a sister group of the widely studied Poales. Most cultivated bananas are natural Musa inter-(sub-specific triploid hybrids. A Musa acuminata reference nuclear genome sequence was recently produced based on sequencing of genomic DNA enriched in nucleus.The Musa acuminata chloroplast genome was assembled with chloroplast reads extracted from whole-genome-shotgun sequence data. The Musa chloroplast genome is a circular molecule of 169,972 bp with a quadripartite structure containing two single copy regions, a Large Single Copy region (LSC, 88,338 bp and a Small Single Copy region (SSC, 10,768 bp separated by Inverted Repeat regions (IRs, 35,433 bp. Two forms of the chloroplast genome relative to the orientation of SSC versus LSC were found. The Musa chloroplast genome shows an extreme IR expansion at the IR/SSC boundary relative to the most common structures found in angiosperms. This expansion consists of the integration of three additional complete genes (rps15, ndhH and ycf1 and part of the ndhA gene. No such expansion has been observed in monocots so far. Simple Sequence Repeats were identified in the Musa chloroplast genome and a new set of Musa chloroplastic markers was designed.The complete sequence of M. acuminata ssp malaccensis chloroplast we reported here is the first one for the Zingiberales order. As such it provides new insight in the evolution of the chloroplast of monocotyledons. In particular, it reinforces that IR/SSC expansion has occurred independently several times within monocotyledons. The discovery of new polymorphic markers within Musa chloroplast opens new perspectives to better understand the origin of cultivated triploid bananas.

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

    Energy Technology Data Exchange (ETDEWEB)

    Saidha, T.; Hanfstingl, U.; Schiff, J.A. (Brandeis Univ., Waltham, MA (USA))

    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.

  2. Is chloroplastic class IIA aldolase a marine enzyme?

    Science.gov (United States)

    Miyasaka, Hitoshi; Ogata, Takeru; Tanaka, Satoshi; Ohama, Takeshi; Kano, Sanae; Kazuhiro, Fujiwara; Hayashi, Shuhei; Yamamoto, Shinjiro; Takahashi, Hiro; Matsuura, Hideyuki; Hirata, Kazumasa

    2016-01-01

    Expressed sequence tag analyses revealed that two marine Chlorophyceae green algae, Chlamydomonas sp. W80 and Chlamydomonas sp. HS5, contain genes coding for chloroplastic class IIA aldolase (fructose-1, 6-bisphosphate aldolase: FBA). These genes show robust monophyly with those of the marine Prasinophyceae algae genera Micromonas, Ostreococcus and Bathycoccus, indicating that the acquisition of this gene through horizontal gene transfer by an ancestor of the green algal lineage occurred prior to the divergence of the core chlorophytes (Chlorophyceae and Trebouxiophyceae) and the prasinophytes. The absence of this gene in some freshwater chlorophytes, such as Chlamydomonas reinhardtii, Volvox carteri, Chlorella vulgaris, Chlorella variabilis and Coccomyxa subellipsoidea, can therefore be explained by the loss of this gene somewhere in the evolutionary process. Our survey on the distribution of this gene in genomic and transcriptome databases suggests that this gene occurs almost exclusively in marine algae, with a few exceptions, and as such, we propose that chloroplastic class IIA FBA is a marine environment-adapted enzyme. This hypothesis was also experimentally tested using Chlamydomonas W80, for which we found that the transcript levels of this gene to be significantly lower under low-salt (that is, simulated terrestrial) conditions. Expression analyses of transcriptome data for two algae, Prymnesium parvum and Emiliania huxleyi, taken from the Sequence Read Archive database also indicated that the expression of this gene under terrestrial conditions (low NaCl and low sulfate) is significantly downregulated. Thus, these experimental and transcriptome data provide support for our hypothesis. PMID:27058504

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

    International Nuclear Information System (INIS)

    Hurewitz, J.; Jagendorf, A.T.

    1986-01-01

    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 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

  4. The molecular architecture of the chloroplast thylakoid membrane

    Energy Technology Data Exchange (ETDEWEB)

    Stefansson, H.

    1996-08-01

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

  5. The complete chloroplast genome sequence of Helwingia himalaica (Helwingiaceae, Aquifoliales) and a chloroplast phylogenomic analysis of the Campanulidae.

    Science.gov (United States)

    Yao, Xin; Liu, Ying-Ying; Tan, Yun-Hong; Song, Yu; Corlett, Richard T

    2016-01-01

    Complete chloroplast genome sequences have been very useful for understanding phylogenetic relationships in angiosperms at the family level and above, but there are currently large gaps in coverage. We report the chloroplast genome for Helwingia himalaica , the first in the distinctive family Helwingiaceae and only the second genus to be sequenced in the order Aquifoliales. We then combine this with 36 published sequences in the large (c. 35,000 species) subclass Campanulidae in order to investigate relationships at the order and family levels. The Helwingia genome consists of 158,362 bp containing a pair of inverted repeat (IR) regions of 25,996 bp separated by a large single-copy (LSC) region and a small single-copy (SSC) region which are 87,810 and 18,560 bp, respectively. There are 142 known genes, including 94 protein-coding genes, eight ribosomal RNA genes, and 40 tRNA genes. The topology of the phylogenetic relationships between Apiales, Asterales, and Dipsacales differed between analyses based on complete genome sequences and on 36 shared protein-coding genes, showing that further studies of campanulid phylogeny are needed.

  6. The complete chloroplast genome sequence of Helwingia himalaica (Helwingiaceae, Aquifoliales and a chloroplast phylogenomic analysis of the Campanulidae

    Directory of Open Access Journals (Sweden)

    Xin Yao

    2016-11-01

    Full Text Available Complete chloroplast genome sequences have been very useful for understanding phylogenetic relationships in angiosperms at the family level and above, but there are currently large gaps in coverage. We report the chloroplast genome for Helwingia himalaica, the first in the distinctive family Helwingiaceae and only the second genus to be sequenced in the order Aquifoliales. We then combine this with 36 published sequences in the large (c. 35,000 species subclass Campanulidae in order to investigate relationships at the order and family levels. The Helwingia genome consists of 158,362 bp containing a pair of inverted repeat (IR regions of 25,996 bp separated by a large single-copy (LSC region and a small single-copy (SSC region which are 87,810 and 18,560 bp, respectively. There are 142 known genes, including 94 protein-coding genes, eight ribosomal RNA genes, and 40 tRNA genes. The topology of the phylogenetic relationships between Apiales, Asterales, and Dipsacales differed between analyses based on complete genome sequences and on 36 shared protein-coding genes, showing that further studies of campanulid phylogeny are needed.

  7. Chloroplast Translation: Structural and Functional Organization, Operational Control, and Regulation[OPEN

    Science.gov (United States)

    2018-01-01

    Chloroplast translation is essential for cellular viability and plant development. Its positioning at the intersection of organellar RNA and protein metabolism makes it a unique point for the regulation of gene expression in response to internal and external cues. Recently obtained high-resolution structures of plastid ribosomes, the development of approaches allowing genome-wide analyses of chloroplast translation (i.e., ribosome profiling), and the discovery of RNA binding proteins involved in the control of translational activity have greatly increased our understanding of the chloroplast translation process and its regulation. In this review, we provide an overview of the current knowledge of the chloroplast translation machinery, its structure, organization, and function. In addition, we summarize the techniques that are currently available to study chloroplast translation and describe how translational activity is controlled and which cis-elements and trans-factors are involved. Finally, we discuss how translational control contributes to the regulation of chloroplast gene expression in response to developmental, environmental, and physiological cues. We also illustrate the commonalities and the differences between the chloroplast and bacterial translation machineries and the mechanisms of protein biosynthesis in these two prokaryotic systems. PMID:29610211

  8. The chloroplast genome of a symbiodinium sp. clade C3 isolate

    KAUST Repository

    Barbrook, Adrian C.

    2014-01-01

    Dinoflagellate algae of the genus Symbiodinium form important symbioses within corals and other benthic marine animals. Dinoflagellates possess an extremely reduced plastid genome relative to those examined in plants and other algae. In dinoflagellates the plastid genes are located on small plasmids, commonly referred to as \\'minicircles\\'. However, the chloroplast genomes of dinoflagellates have only been extensively characterised from a handful of species. There is also evidence of considerable variation in the chloroplast genome organisation across those species that have been examined. We therefore characterised the chloroplast genome from an environmental coral isolate, in this case containing a symbiont belonging to the Symbiodinium sp. clade C3. The gene content of the genome is well conserved with respect to previously characterised genomes. However, unlike previously characterised dinoflagellate chloroplast genomes we did not identify any \\'empty\\' minicircles. The sequences of this chloroplast genome show a high rate of evolution relative to other algal species. Particularly notable was a surprisingly high level of sequence divergence within the core polypeptides of photosystem I, the reasons for which are currently unknown. This chloroplast genome also possesses distinctive codon usage and GC content. These features suggest that chloroplast genomes in Symbiodinium are highly plastic. © 2013 Adrian C. Barbrook.

  9. Chloroplast two-component systems: evolution of the link between photosynthesis and gene expression.

    Science.gov (United States)

    Puthiyaveetil, Sujith; Allen, John F

    2009-06-22

    Two-component signal transduction, consisting of sensor kinases and response regulators, is the predominant signalling mechanism in bacteria. This signalling system originated in prokaryotes and has spread throughout the eukaryotic domain of life through endosymbiotic, lateral gene transfer from the bacterial ancestors and early evolutionary precursors of eukaryotic, cytoplasmic, bioenergetic organelles-chloroplasts and mitochondria. Until recently, it was thought that two-component systems inherited from an ancestral cyanobacterial symbiont are no longer present in chloroplasts. Recent research now shows that two-component systems have survived in chloroplasts as products of both chloroplast and nuclear genes. Comparative genomic analysis of photosynthetic eukaryotes shows a lineage-specific distribution of chloroplast two-component systems. The components and the systems they comprise have homologues in extant cyanobacterial lineages, indicating their ancient cyanobacterial origin. Sequence and functional characteristics of chloroplast two-component systems point to their fundamental role in linking photosynthesis with gene expression. We propose that two-component systems provide a coupling between photosynthesis and gene expression that serves to retain genes in chloroplasts, thus providing the basis of cytoplasmic, non-Mendelian inheritance of plastid-associated characters. We discuss the role of this coupling in the chronobiology of cells and in the dialogue between nuclear and cytoplasmic genetic systems.

  10. Structure-Function Analysis of Chloroplast Proteins via Random Mutagenesis Using Error-Prone PCR.

    Science.gov (United States)

    Dumas, Louis; Zito, Francesca; Auroy, Pascaline; Johnson, Xenie; Peltier, Gilles; Alric, Jean

    2018-06-01

    Site-directed mutagenesis of chloroplast genes was developed three decades ago and has greatly advanced the field of photosynthesis research. Here, we describe a new approach for generating random chloroplast gene mutants that combines error-prone polymerase chain reaction of a gene of interest with chloroplast complementation of the knockout Chlamydomonas reinhardtii mutant. As a proof of concept, we targeted a 300-bp sequence of the petD gene that encodes subunit IV of the thylakoid membrane-bound cytochrome b 6 f complex. By sequencing chloroplast transformants, we revealed 149 mutations in the 300-bp target petD sequence that resulted in 92 amino acid substitutions in the 100-residue target subunit IV sequence. Our results show that this method is suited to the study of highly hydrophobic, multisubunit, and chloroplast-encoded proteins containing cofactors such as hemes, iron-sulfur clusters, and chlorophyll pigments. Moreover, we show that mutant screening and sequencing can be used to study photosynthetic mechanisms or to probe the mutational robustness of chloroplast-encoded proteins, and we propose that this method is a valuable tool for the directed evolution of enzymes in the chloroplast. © 2018 American Society of Plant Biologists. All rights reserved.

  11. Functional Disruption of a Chloroplast Pseudouridine Synthase Desensitizes Arabidopsis Plants to Phosphate Starvation

    Directory of Open Access Journals (Sweden)

    Shan Lu

    2017-08-01

    Full Text Available Phosphate (Pi deficiency is a common nutritional stress of plants in both agricultural and natural ecosystems. Plants respond to Pi starvation in the environment by triggering a suite of biochemical, physiological, and developmental changes that increase survival and growth. The key factors that determine plant sensitivity to Pi starvation, however, are unclear. In this research, we identified an Arabidopsis mutant, dps1, with greatly reduced sensitivity to Pi starvation. The dps1 phenotypes are caused by a mutation in the previously characterized SVR1 (SUPPRESSION OF VARIAGATION 1 gene, which encodes a chloroplast-localized pseudouridine synthase. The mutation of SVR1 results in defects in chloroplast rRNA biogenesis, which subsequently reduces chloroplast translation. Another mutant, rps5, which contains a mutation in the chloroplast ribosomal protein RPS5 and has reduced chloroplast translation, also displayed decreased sensitivity to Pi starvation. Furthermore, wild type plants treated with lincomycin, a chemical inhibitor of chloroplast translation, showed similar growth phenotypes and Pi starvation responses as dps1 and rps5. These results suggest that impaired chloroplast translation desensitizes plants to Pi starvation. Combined with previously published results showing that enhanced leaf photosynthesis augments plant responses to Pi starvation, we propose that the decrease in responses to Pi starvation in dps1, rps5, and lincomycin-treated plants is due to their reduced demand for Pi input from the environment.

  12. The chloroplast genome of a symbiodinium sp. clade C3 isolate

    KAUST Repository

    Barbrook, Adrian C.; Voolstra, Christian R.; Howe, Christopher J.

    2014-01-01

    Dinoflagellate algae of the genus Symbiodinium form important symbioses within corals and other benthic marine animals. Dinoflagellates possess an extremely reduced plastid genome relative to those examined in plants and other algae. In dinoflagellates the plastid genes are located on small plasmids, commonly referred to as 'minicircles'. However, the chloroplast genomes of dinoflagellates have only been extensively characterised from a handful of species. There is also evidence of considerable variation in the chloroplast genome organisation across those species that have been examined. We therefore characterised the chloroplast genome from an environmental coral isolate, in this case containing a symbiont belonging to the Symbiodinium sp. clade C3. The gene content of the genome is well conserved with respect to previously characterised genomes. However, unlike previously characterised dinoflagellate chloroplast genomes we did not identify any 'empty' minicircles. The sequences of this chloroplast genome show a high rate of evolution relative to other algal species. Particularly notable was a surprisingly high level of sequence divergence within the core polypeptides of photosystem I, the reasons for which are currently unknown. This chloroplast genome also possesses distinctive codon usage and GC content. These features suggest that chloroplast genomes in Symbiodinium are highly plastic. © 2013 Adrian C. Barbrook.

  13. Cadmium Disrupts Subcellular Organelles, Including Chloroplasts, Resulting in Melatonin Induction in Plants

    Directory of Open Access Journals (Sweden)

    Hyoung-Yool Lee

    2017-10-01

    Full Text Available Cadmium is a well-known elicitor of melatonin synthesis in plants, including rice. However, the mechanisms by which cadmium induces melatonin induction remain elusive. To investigate whether cadmium influences physical integrities in subcellular organelles, we treated tobacco leaves with either CdCl2 or AlCl3 and monitored the structures of subcellular organelles—such as chloroplasts, mitochondria, and the endoplasmic reticulum (ER—using confocal microscopic analysis. Unlike AlCl3 treatment, CdCl2 (0.5 mM treatment significantly disrupted chloroplasts, mitochondria, and ER. In theory, the disruption of chloroplasts enabled chloroplast-expressed serotonin N-acetyltransferase (SNAT to encounter serotonin in the cytoplasm, leading to the synthesis of N-acetylserotonin followed by melatonin synthesis. In fact, the disruption of chloroplasts by cadmium, not by aluminum, gave rise to a huge induction of melatonin in rice leaves, which suggests that cadmium-treated chloroplast disruption plays an important role in inducing melatonin in plants by removing physical barriers, such as chloroplast double membranes, allowing SNAT to gain access to the serotonin substrate enriched in the cytoplasm.

  14. Dated tribe-wide whole chloroplast genome phylogeny indicates recurrent hybridizations within Triticeae.

    Science.gov (United States)

    Bernhardt, Nadine; Brassac, Jonathan; Kilian, Benjamin; Blattner, Frank R

    2017-06-16

    Triticeae, the tribe of wheat grasses, harbours the cereals barley, rye and wheat and their wild relatives. Although economically important, relationships within the tribe are still not understood. We analysed the phylogeny of chloroplast lineages among nearly all monogenomic Triticeae taxa and polyploid wheat species aiming at a deeper understanding of the tribe's evolution. We used on- and off-target reads of a target-enrichment experiment followed by Illumina sequencing. The read data was used to assemble the plastid locus ndhF for 194 individuals and the whole chloroplast genome for 183 individuals, representing 53 Triticeae species and 15 genera. We conducted Bayesian and multispecies coalescent analyses to infer relationships and estimate divergence times of the taxa. We present the most comprehensive dated Triticeae chloroplast phylogeny and review previous hypotheses in the framework of our results. Monophyly of Triticeae chloroplasts could not be confirmed, as either Bromus or Psathyrostachys captured a chloroplast from a lineage closely related to a Bromus-Triticeae ancestor. The most recent common ancestor of Triticeae occurred approximately between ten and 19 million years ago. The comparison of the chloroplast phylogeny with available nuclear data in several cases revealed incongruences indicating past hybridizations. Recent events of chloroplast capture were detected as individuals grouped apart from con-specific accessions in otherwise monopyhletic groups.

  15. Complete sequencing of five araliaceae chloroplast genomes and the phylogenetic implications.

    Directory of Open Access Journals (Sweden)

    Rong Li

    Full Text Available BACKGROUND: The ginseng family (Araliaceae includes a number of economically important plant species. Previously phylogenetic studies circumscribed three major clades within the core ginseng plant family, yet the internal relationships of each major group have been poorly resolved perhaps due to rapid radiation of these lineages. Recent studies have shown that phyogenomics based on chloroplast genomes provides a viable way to resolve complex relationships. METHODOLOGY/PRINCIPAL FINDINGS: We report the complete nucleotide sequences of five Araliaceae chloroplast genomes using next-generation sequencing technology. The five chloroplast genomes are 156,333-156,459 bp in length including a pair of inverted repeats (25,551-26,108 bp separated by the large single-copy (86,028-86,566 bp and small single-copy (18,021-19,117 bp regions. Each chloroplast genome contains the same 114 unique genes consisting of 30 transfer RNA genes, four ribosomal RNA genes, and 80 protein coding genes. Gene size, content, and order, AT content, and IR/SC boundary structure are similar among all Araliaceae chloroplast genomes. A total of 140 repeats were identified in the five chloroplast genomes with palindromic repeat as the most common type. Phylogenomic analyses using parsimony, likelihood, and Bayesian inference based on the complete chloroplast genomes strongly supported the monophyly of the Asian Palmate group and the Aralia-Panax group. Furthermore, the relationships among the sampled taxa within the Asian Palmate group were well resolved. Twenty-six DNA markers with the percentage of variable sites higher than 5% were identified, which may be useful for phylogenetic studies of Araliaceae. CONCLUSION: The chloroplast genomes of Araliaceae are highly conserved in all aspects of genome features. The large-scale phylogenomic data based on the complete chloroplast DNA sequences is shown to be effective for the phylogenetic reconstruction of Araliaceae.

  16. Combined analysis of the chloroplast genome and transcriptome of the Antarctic vascular plant Deschampsia antarctica Desv.

    Science.gov (United States)

    Lee, Jungeun; Kang, Yoonjee; Shin, Seung Chul; Park, Hyun; Lee, Hyoungseok

    2014-01-01

    Antarctic hairgrass (Deschampsia antarctica Desv.) is the only natural grass species in the maritime Antarctic. It has been researched as an important ecological marker and as an extremophile plant for studies on stress tolerance. Despite its importance, little genomic information is available for D. antarctica. Here, we report the complete chloroplast genome, transcriptome profiles of the coding/noncoding genes, and the posttranscriptional processing by RNA editing in the chloroplast system. The complete chloroplast genome of D. antarctica is 135,362 bp in length with a typical quadripartite structure, including the large (LSC: 79,881 bp) and small (SSC: 12,519 bp) single-copy regions, separated by a pair of identical inverted repeats (IR: 21,481 bp). It contains 114 unique genes, including 81 unique protein-coding genes, 29 tRNA genes, and 4 rRNA genes. Sequence divergence analysis with other plastomes from the BEP clade of the grass family suggests a sister relationship between D. antarctica, Festuca arundinacea and Lolium perenne of the Poeae tribe, based on the whole plastome. In addition, we conducted high-resolution mapping of the chloroplast-derived transcripts. Thus, we created an expression profile for 81 protein-coding genes and identified ndhC, psbJ, rps19, psaJ, and psbA as the most highly expressed chloroplast genes. Small RNA-seq analysis identified 27 small noncoding RNAs of chloroplast origin that were preferentially located near the 5'- or 3'-ends of genes. We also found >30 RNA-editing sites in the D. antarctica chloroplast genome, with a dominance of C-to-U conversions. We assembled and characterized the complete chloroplast genome sequence of D. antarctica and investigated the features of the plastid transcriptome. These data may contribute to a better understanding of the evolution of D. antarctica within the Poaceae family for use in molecular phylogenetic studies and may also help researchers understand the characteristics of the chloroplast

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

    Directory of Open Access Journals (Sweden)

    Zhou Xiangjun

    2011-11-01

    Full Text Available Abstract Background Chloroplasts are the green plastids where photosynthesis takes place. The biogenesis of chloroplasts requires the coordinate expression of both nuclear and chloroplast genes and is regulated by developmental and environmental signals. Despite extensive studies of this process, the genetic basis and the regulatory control of chloroplast biogenesis and development remain to be elucidated. Results Green cauliflower mutant causes ectopic development of chloroplasts in the curd tissue of the plant, turning the otherwise white curd green. To investigate the transcriptional control of chloroplast development, we compared gene expression between green and white curds using the RNA-seq approach. Deep sequencing produced over 15 million reads with lengths of 86 base pairs from each cDNA library. A total of 7,155 genes were found to exhibit at least 3-fold changes in expression between green and white curds. These included light-regulated genes, genes encoding chloroplast constituents, and genes involved in chlorophyll biosynthesis. Moreover, we discovered that the cauliflower ELONGATED HYPOCOTYL5 (BoHY5 was expressed higher in green curds than white curds and that 2616 HY5-targeted genes, including 1600 up-regulated genes and 1016 down-regulated genes, were differently expressed in green in comparison to white curd tissue. All these 1600 up-regulated genes were HY5-targeted genes in the light. Conclusions The genome-wide profiling of gene expression by RNA-seq in green curds led to the identification of large numbers of genes associated with chloroplast development, and suggested the role of regulatory genes in the high hierarchy of light signaling pathways in mediating the ectopic chloroplast development in the green curd cauliflower mutant.

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

    Science.gov (United States)

    Zhou, Xiangjun; Fei, Zhangjun; Thannhauser, Theodore W; Li, Li

    2011-11-23

    Chloroplasts are the green plastids where photosynthesis takes place. The biogenesis of chloroplasts requires the coordinate expression of both nuclear and chloroplast genes and is regulated by developmental and environmental signals. Despite extensive studies of this process, the genetic basis and the regulatory control of chloroplast biogenesis and development remain to be elucidated. Green cauliflower mutant causes ectopic development of chloroplasts in the curd tissue of the plant, turning the otherwise white curd green. To investigate the transcriptional control of chloroplast development, we compared gene expression between green and white curds using the RNA-seq approach. Deep sequencing produced over 15 million reads with lengths of 86 base pairs from each cDNA library. A total of 7,155 genes were found to exhibit at least 3-fold changes in expression between green and white curds. These included light-regulated genes, genes encoding chloroplast constituents, and genes involved in chlorophyll biosynthesis. Moreover, we discovered that the cauliflower ELONGATED HYPOCOTYL5 (BoHY5) was expressed higher in green curds than white curds and that 2616 HY5-targeted genes, including 1600 up-regulated genes and 1016 down-regulated genes, were differently expressed in green in comparison to white curd tissue. All these 1600 up-regulated genes were HY5-targeted genes in the light. The genome-wide profiling of gene expression by RNA-seq in green curds led to the identification of large numbers of genes associated with chloroplast development, and suggested the role of regulatory genes in the high hierarchy of light signaling pathways in mediating the ectopic chloroplast development in the green curd cauliflower mutant.

  19. Destruction of pigments and lipids in isolated chloroplasts under the effect of visible radiation

    International Nuclear Information System (INIS)

    Merzlyak, M.N.; Pogosyan, S.I.

    1988-01-01

    The results of experiments on the effect of light radiation on lipid and pigment destruction in isolated chloroplasts are generalized. Substrates and products of oxidation destruction of lipid and pigments, the role of photosynthetic electron transport in photodestruction, the participation of activated oxygen and free-radical intermediate forms in it are considered. The role of antioxidants, carotenoids and enzymatic systems in protection of chloroplast membranes from destructive light effect is discussed. A general scheme of possible ways of photodestruction in chloroplasts is presented. 53 refs

  20. The whole chloroplast genome of wild rice (Oryza australiensis).

    Science.gov (United States)

    Wu, Zhiqiang; Ge, Song

    2016-01-01

    The whole chloroplast genome of wild rice (Oryza australiensis) is characterized in this study. The genome size is 135,224  bp, exhibiting a typical circular structure including a pair of 25,776  bp inverted repeats (IRa,b) separated by a large single-copy region (LSC) of 82,212  bp and a small single-copy region (SSC) of 12,470  bp. The overall GC content of the genome is 38.95%. 110 unique genes were annotated, including 76 protein-coding genes, 4 ribosomal RNA genes, and 30t RNA genes. Among these, 18 are duplicated in the inverted repeat regions, 13 genes contain one intron, and 2 genes (rps12 and ycf3) have two introns.

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

    Science.gov (United States)

    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.

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

    Science.gov (United States)

    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.

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

    Science.gov (United States)

    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.

  4. Photosynthetic production of diterpenoids in chloroplasts and cyanobacteria

    DEFF Research Database (Denmark)

    Vavitsas, Konstantinos

    Terpenoids are one of the largest classes of chemical compounds, some of them with industrial interest as nutraceuticals, biofuels, or chemical feedstocks. Diterpenoids are a large terpenoid subclass, and their chemical structure consists of a core skeleton of 20 carbon atoms. This skeleton can...... be further modified by cyclizing enzymes, and be decorated by the addition of chemical groups. Even though they are mainly plant-derived compounds, diterpenoid production in photosynthetic organisms is rather unexplored, with a few successful studies reported in the literature. In this thesis, I elaborate...... on the potential of using plant chloroplasts and cyanobacteria as biosynthetic vessels, with a focus on diterpenoid production, and on the potential direct linking of photosynthesis to drive electron-consuming enzymes, such as the monooxygenases cytochrome P450s. I subsequently present the full localization...

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

    Energy Technology Data Exchange (ETDEWEB)

    Wetzel, Carolyn M

    2005-02-22

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

  6. The complete chloroplast genome of Sinopodophyllum hexandrum Ying (Berberidaceae).

    Science.gov (United States)

    Meng, Lihua; Liu, Ruijuan; Chen, Jianbing; Ding, Chenxu

    2017-05-01

    The complete nucleotide sequence of the Sinopodophyllum hexandrum Ying chloroplast genome (cpDNA) was determined based on next-generation sequencing technologies in this study. The genome was 157 203 bp in length, containing a pair of inverted repeat (IRa and IRb) regions of 25 960 bp, which were separated by a large single-copy (LSC) region of 87 065 bp and a small single-copy (SSC) region of 18 218 bp, respectively. The cpDNA contained 148 genes, including 96 protein-coding genes, 8 ribosomal RNA genes, and 44 tRNA genes. In these genes, eight harbored a single intron, and two (ycf3 and clpP) contained a couple of introns. The cpDNA AT content of S. hexandrum cpDNA is 61.5%.

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

    Science.gov (United States)

    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.

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

    Science.gov (United States)

    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.

  9. The complete chloroplast genome sequence of Dendrobium nobile.

    Science.gov (United States)

    Yan, Wenjin; Niu, Zhitao; Zhu, Shuying; Ye, Meirong; Ding, Xiaoyu

    2016-11-01

    The complete chloroplast (cp) genome sequence of Dendrobium nobile, an endangered and traditional Chinese medicine with important economic value, is presented in this article. The total genome size is 150,793 bp, containing a large single copy (LSC) region (84,939 bp) and a small single copy region (SSC) (13,310 bp) which were separated by two inverted repeat (IRs) regions (26,272 bp). The overall GC contents of the plastid genome were 38.8%. In total, 130 unique genes were annotated and they were consisted of 76 protein-coding genes, 30 tRNA genes and 4 rRNA genes. Fourteen genes contained one or two introns.

  10. Metallothionein expression in chloroplasts enhances mercury accumulation and phytoremediation capability.

    Science.gov (United States)

    Ruiz, Oscar N; Alvarez, Derry; Torres, Cesar; Roman, Laura; Daniell, Henry

    2011-06-01

    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 metallothioneins in plants for mercury phytoremediation. Chloroplast genetic engineering approach is useful to express metal-scavenging proteins for phytoremediation. © 2011 The Authors. Plant Biotechnology Journal © 2011 Society for Experimental Biology, Association of Applied Biologists and Blackwell Publishing Ltd.

  11. The complete chloroplast genomes of two Wisteria species, W. floribunda and W. sinensis (Fabaceae).

    Science.gov (United States)

    Kim, Na-Rae; Kim, Kyunghee; Lee, Sang-Choon; Lee, Jung-Hoon; Cho, Seong-Hyun; Yu, Yeisoo; Kim, Young-Dong; Yang, Tae-Jin

    2016-11-01

    Wisteria floribunda and Wisteria sinensis are ornamental woody vines in the Fabaceae. The complete chloroplast genome sequences of the two species were generated by de novo assembly using whole genome next generation sequences. The chloroplast genomes of W. floribunda and W. sinensis were 130 960 bp and 130 561 bp long, respectively, and showed inverted repeat (IR)-lacking structures as those reported in IRLC in the Fabaceae. The chloroplast genomes of both species contained same number of protein-coding sequences (77), tRNA genes (30), and rRNA genes (4). The phylogenetic analysis with the reported chloroplast genomes confirmed close taxonomical relationship of W. floribunda and W. sinensis.

  12. Formation and scavenging of superoxide in chloroplasts, with relation to injury by sulfur dioxide

    Energy Technology Data Exchange (ETDEWEB)

    Asada, K

    1980-01-01

    Injury of plant leaf cells by sulfur dioxide-exposure is greater in day time than in night. A hypothesis is proposed that the free radical chain oxidation of sulfite is initiated by the superoxide radicals (O/sub 2//sup -/) produced in illuminated chloroplasts, and that the resulting amplified production of O/sub 2//sup -/, the hydroxyl radicals and the bisulfite radicals causes the injury of leaf tissues. In this review, the production of O/sub 2//sup -/ in illuminated chloroplasts and scavenging of O/sub 2//sup -/ by superoxide dismutase and their relation to oxidation of sulfite in chloroplasts are discussed. Superoxide dismutase in chloroplasts plays an important role in protecting leaf cells from injury by sulfur dioxide.

  13. Comparative analyses of chloroplast genome data representing nine green algae in Sphaeropleales (Chlorophyceae, Chlorophyta

    Directory of Open Access Journals (Sweden)

    Karolina Fučíková

    2016-06-01

    Full Text Available 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].

  14. Chloroplasts activity and PAP-signaling regulate programmed cell death in Arabidopsis

    KAUST Repository

    Bruggeman, Quentin; Mazubert, Christelle; Prunier, Florence; Lugan, Raphael; Chan, Kai Xun; Phua, Su Yin; Pogson, Barry J.; Krieger-Liszkay, Anja; Delarue, Marianne; Benhamed, Moussa; Bergounioux, Catherine; Raynaud, Cé cile

    2016-01-01

    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

  15. The metabolism of sorbitol and fructose in isolated chloroplasts of Santa Rosa plum leaves

    International Nuclear Information System (INIS)

    De Villiers, O.T.

    1979-01-01

    Aqueously as well as non-aqueously isolated chloroplasts from Santa Rosa plum leaves readily metabolised sorbitol- 14 C to fructose, glucose and sucrose. Likewise, fructose- 14 C was converted to sorbitol, glucose and sucrose [af

  16. Genetic polymorphism in Gymnodinium galatheanum chloroplast DNA sequences and development of a molecular detection assay.

    Science.gov (United States)

    Tengs, T; Bowers, H A; Ziman, A P; Stoecker, D K; Oldach, D W

    2001-02-01

    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 sequence divergence than the nuclear homologue. The chloroplast sequences were chosen as targets for the development of a 5'--3' exonuclease assay for detection of the organism. The assay has a very high degree of specificity and has been used to screen environmental water samples from a fish farm where the presence of this dinoflagellate species has previously been associated with fish kills. Various hypotheses for the derived nature of the chloroplast sequences are discussed, as well as what is known about the toxicity of the species.

  17. Confocal laser scanning microscopy detection of chlorophylls and carotenoids in chloroplasts and chromoplasts of tomato fruit.

    Science.gov (United States)

    D'Andrea, Lucio; Amenós, Montse; Rodríguez-Concepción, Manuel

    2014-01-01

    Plant cells are unique among eukaryotic cells because of the presence of plastids, including chloroplasts and chromoplasts. Chloroplasts are found in green tissues and harbor the photosynthetic machinery (including chlorophyll molecules), while chromoplasts are present in non-photosynthetic tissues and accumulate large amounts of carotenoids. During tomato fruit development, chloroplasts are converted into chromoplasts that accumulate high levels of lycopene, a linear carotenoid responsible for the characteristic red color of ripe fruit. Here, we describe a simple and fast method to detect both types of fully differentiated plastids (chloroplasts and chromoplasts), as well as intermediate stages, in fresh tomato fruits. The method is based on the differential autofluorescence of chlorophylls and carotenoids (lycopene) detected by Confocal Laser Scanning Microscopy.

  18. Regulation of chloroplast number and DNA synthesis in higher plants. Final report

    Energy Technology Data Exchange (ETDEWEB)

    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.

  19. Frataxin Is Localized to Both the Chloroplast and Mitochondrion and Is Involved in Chloroplast Fe-S Protein Function in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Valeria R Turowski

    Full Text Available 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.

  20. Enzymic synthesis of γ-coniceine in Conium maculatum chloroplasts and mitochondria.

    Science.gov (United States)

    Roberts, M F

    1981-08-01

    Further studies of the transaminase responsible for the first committed step in alkaloid formation in Conium maculatum have shown the L-alanine: 5-ketooctanal transaminase to occur in both the mitochondria and chloroplast. Experiments suggest that these enzymes are the isoenzymes Transaminase A and B respectively previously isolated by the author. It is suggested that the chloroplast enzyme is normally responsible for alkaloid production.

  1. Release of proteins from intact chloroplasts induced by reactive oxygen species during biotic and abiotic stress.

    Science.gov (United States)

    Kwon, Kwang-Chul; Verma, Dheeraj; Jin, Shuangxia; Singh, Nameirakpam D; Daniell, Henry

    2013-01-01

    Plastids sustain life on this planet by providing food, feed, essential biomolecules and oxygen. Such diverse metabolic and biosynthetic functions require efficient communication between plastids and the nucleus. However, specific factors, especially large molecules, released from plastids that regulate nuclear genes have not yet been fully elucidated. When tobacco and lettuce transplastomic plants expressing GFP within chloroplasts, were challenged with Erwinia carotovora (biotic stress) or paraquat (abiotic stress), GFP was released into the cytoplasm. During this process GFP moves gradually towards the envelope, creating a central red zone of chlorophyll fluorescence. GFP was then gradually released from intact chloroplasts into the cytoplasm with an intact vacuole and no other visible cellular damage. Different stages of GFP release were observed inside the same cell with a few chloroplasts completely releasing GFP with detection of only red chlorophyll fluorescence or with no reduction in GFP fluorescence or transitional steps between these two phases. Time lapse imaging by confocal microscopy clearly identified sequence of these events. Intactness of chloroplasts during this process was evident from chlorophyll fluorescence emanated from thylakoid membranes and in vivo Chla fluorescence measurements (maximum quantum yield of photosystem II) made before or after infection with pathogens to evaluate their photosynthetic competence. Hydrogen peroxide and superoxide anion serve as signal molecules for generation of reactive oxygen species and Tiron, scavenger of superoxide anion, blocked release of GFP from chloroplasts. Significant increase in ion leakage in the presence of paraquat and light suggests changes in the chloroplast envelope to facilitate protein release. Release of GFP-RC101 (an antimicrobial peptide), which was triggered by Erwinia infection, ceased after conferring protection, further confirming this export phenomenon. These results suggest a

  2. Release of proteins from intact chloroplasts induced by reactive oxygen species during biotic and abiotic stress.

    Directory of Open Access Journals (Sweden)

    Kwang-Chul Kwon

    Full Text Available Plastids sustain life on this planet by providing food, feed, essential biomolecules and oxygen. Such diverse metabolic and biosynthetic functions require efficient communication between plastids and the nucleus. However, specific factors, especially large molecules, released from plastids that regulate nuclear genes have not yet been fully elucidated. When tobacco and lettuce transplastomic plants expressing GFP within chloroplasts, were challenged with Erwinia carotovora (biotic stress or paraquat (abiotic stress, GFP was released into the cytoplasm. During this process GFP moves gradually towards the envelope, creating a central red zone of chlorophyll fluorescence. GFP was then gradually released from intact chloroplasts into the cytoplasm with an intact vacuole and no other visible cellular damage. Different stages of GFP release were observed inside the same cell with a few chloroplasts completely releasing GFP with detection of only red chlorophyll fluorescence or with no reduction in GFP fluorescence or transitional steps between these two phases. Time lapse imaging by confocal microscopy clearly identified sequence of these events. Intactness of chloroplasts during this process was evident from chlorophyll fluorescence emanated from thylakoid membranes and in vivo Chla fluorescence measurements (maximum quantum yield of photosystem II made before or after infection with pathogens to evaluate their photosynthetic competence. Hydrogen peroxide and superoxide anion serve as signal molecules for generation of reactive oxygen species and Tiron, scavenger of superoxide anion, blocked release of GFP from chloroplasts. Significant increase in ion leakage in the presence of paraquat and light suggests changes in the chloroplast envelope to facilitate protein release. Release of GFP-RC101 (an antimicrobial peptide, which was triggered by Erwinia infection, ceased after conferring protection, further confirming this export phenomenon. These

  3. Chloroplast and mitochondrial microsatellites for Millettia pinnata (Fabaceae) and cross-amplification in related species 1

    OpenAIRE

    Wang, Yanling; Xie, Hongxian; Yang, Yi; Huang, Yelin; Wang, Jianwu; Tan, Fengxiao

    2017-01-01

    Premise of the study: Chloroplast and mitochondrial microsatellites were identified to study the population genetics of Millettia pinnata (Fabaceae). Methods and Results: Based on publicly available plastid genome sequence data of M. pinnata, 42 primer pairs were developed, of which 17 displayed polymorphisms across 89 individuals from four populations. For chloroplast loci, two to six alleles were recovered and the unbiased haploid diversity per locus ranged from 0.391 to 0.857. For mitochon...

  4. Transcriptional regulation and DNA methylation in plastids during transitional conversion of chloroplasts to chromoplasts.

    OpenAIRE

    Kobayashi, H; Ngernprasirtsiri, J; Akazawa, T

    1990-01-01

    During transitional conversion of chloroplasts to chromoplasts in ripening tomato (Lycopersicon esculentum) fruits, transcripts for several plastid genes for photosynthesis decreased to undetectable levels. Run-on transcription of plastids indicated that transcriptional regulation operated as a predominant factor. We found that most of the genes in chloroplasts were actively transcribed in vitro by Escherichia coli and soluble plastid RNA polymerases, but some genes in chromoplasts seemed to ...

  5. Effect of Radiation Dosage on Efficiency of Chloroplast Transfer by Protoplast Fusion in Nicotiana

    OpenAIRE

    Menczel, László; Galiba, Gábor; Nagy, Ferenc; Maliga, Pál

    1982-01-01

    Chloroplasts of Nicotiana tabacum SR1 were transferred into Nicotiana plumbaginifolia by protoplast fusion. The protoplasts of the organelle donor were irradiated with different lethal doses using a 60Co source, to facilitate the elimination of their nuclei from the fusion products. After fusion induction, clones derived from fusion products and containing streptomycin-resistant N. tabacum SR1 chloroplasts were selected by their ability to green on a selective medium. When N. tabacum protopla...

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

    Directory of Open Access Journals (Sweden)

    Kalpana eManandhar-Shrestha

    2013-03-01

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

  7. Characterization of the snowy cotyledon 1 mutant of Arabidopsis thaliana: the impact of chloroplast elongation factor G on chloroplast development and plant vitality.

    Science.gov (United States)

    Albrecht, Verónica; Ingenfeld, Anke; Apel, Klaus

    2006-03-01

    During seedling development chloroplast formation marks the transition from heterotrophic to autotrophic growth. The development and activity of chloroplasts may differ in cotyledons that initially serve as a storage organ and true leaves whose primary function is photosynthesis. A genetic screen was used for the identification of genes that affect selectively chloroplast function in cotyledons of Arabidopsis thaliana. Several mutants exhibiting pale cotyledons and green true leaves were isolated and dubbed snowy cotyledon (sco). One of the mutants, sco1, was characterized in more detail. The mutated gene was identified using map-based cloning. The mutant contains a point mutation in a gene encoding the chloroplast elongation factor G, leading to an amino acid exchange within the predicted 70S ribosome-binding domain. The mutation results in a delay in the onset of germination. At this early developmental stage embryos still contain undifferentiated proplastids, whose proper function seems necessary for seed germination. In light-grown sco1 seedlings the greening of cotyledons is severely impaired, whereas the following true leaves develop normally as in wild-type plants. Despite this apparent similarity of chloroplast development in true leaves of mutant and wild-type plants various aspects of mature plant development are also affected by the sco1 mutation such as the onset of flowering, the growth rate, and seed production. The onset of senescence in the mutant and the wild-type plants occurs, however, at the same time, suggesting that in the mutant this particular developmental step does not seem to suffer from reduced protein translation efficiency in chloroplasts.

  8. Chloroplast overexpression of rice caffeic acid O-methyltransferase increases melatonin production in chloroplasts via the 5-methoxytryptamine pathway in transgenic rice plants.

    Science.gov (United States)

    Choi, Geun-Hee; Lee, Hyoung Yool; Back, Kyoungwhan

    2017-08-01

    Recent analyses of the enzymatic features of various melatonin biosynthetic genes from bacteria, animals, and plants have led to the hypothesis that melatonin could be synthesized via the 5-methoxytryptamine (5-MT) pathway. 5-MT is known to be synthesized in vitro from serotonin by the enzymatic action of O-methyltransferases, including N-acetylserotonin methyltransferase (ASMT) and caffeic acid O-methyltransferase (COMT), leading to melatonin synthesis by the subsequent enzymatic reaction with serotonin N-acetyltransferase (SNAT). Here, we show that 5-MT was produced and served as a precursor for melatonin synthesis in plants. When rice seedlings were challenged with senescence treatment, 5-MT levels and melatonin production were increased in transgenic rice seedlings overexpressing the rice COMT in chloroplasts, while no such increases were observed in wild-type or transgenic seedlings overexpressing the rice COMT in the cytosol, suggesting a 5-MT transport limitation from the cytosol to chloroplasts. In contrast, cadmium treatment led to results different from those in senescence. The enhanced melatonin production was not observed in the chloroplast COMT lines relative over the cytosol COMT lines although 5-MT levels were equally induced in all genotypes upon cadmium treatment. The transgenic seedlings with enhanced melatonin in their chloroplasts exhibited improved seedling growth vs the wild type under continuous light conditions. This is the first report describing enhanced melatonin production in chloroplasts via the 5-MT pathway with the ectopic overexpression of COMT in chloroplasts in plants. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  9. Chloroplast behaviour and interactions with other organelles in Arabidopsis thaliana pavement cells.

    Science.gov (United States)

    Barton, Kiah A; Wozny, Michael R; Mathur, Neeta; Jaipargas, Erica-Ashley; Mathur, Jaideep

    2018-01-29

    Chloroplasts are a characteristic feature of green plants. Mesophyll cells possess the majority of chloroplasts and it is widely believed that, with the exception of guard cells, the epidermal layer in most higher plants does not contain chloroplasts. However, recent observations on Arabidopsis thaliana have shown a population of chloroplasts in pavement cells that are smaller than mesophyll chloroplasts and have a high stroma to grana ratio. Here, using stable transgenic lines expressing fluorescent proteins targeted to the plastid stroma, plasma membrane, endoplasmic reticulum, tonoplast, nucleus, mitochondria, peroxisomes, F-actin and microtubules, we characterize the spatiotemporal relationships between the pavement cell chloroplasts (PCCs) and their subcellular environment. Observations on the PCCs suggest a source-sink relationship between the epidermal and the mesophyll layers, and experiments with the Arabidopsis mutants glabra2 ( gl2 ) and immutans ( im ), which show altered epidermal plastid development, underscored their developmental plasticity. Our findings lay down the foundation for further investigations aimed at understanding the precise role and contributions of PCCs in plant interactions with the environment. © 2018. Published by The Company of Biologists Ltd.

  10. Conflict amongst chloroplast DNA sequences obscures the phylogeny of a group of Asplenium ferns.

    Science.gov (United States)

    Shepherd, Lara D; Holland, Barbara R; Perrie, Leon R

    2008-07-01

    A previous study of the relationships amongst three subgroups of the Austral Asplenium ferns found conflicting signal between the two chloroplast loci investigated. Because organelle genomes like those of chloroplasts and mitochondria are thought to be non-recombining, with a single evolutionary history, we sequenced four additional chloroplast loci with the expectation that this would resolve these relationships. Instead, the conflict was only magnified. Although tree-building analyses favoured one of the three possible trees, one of the alternative trees actually had one more supporting site (six versus five) and received greater support in spectral and neighbor-net analyses. Simulations suggested that chance alone was unlikely to produce strong support for two of the possible trees and none for the third. Likelihood permutation tests indicated that the concatenated chloroplast sequence data appeared to have experienced recombination. However, recombination between the chloroplast genomes of different species would be highly atypical, and corollary supporting observations, like chloroplast heteroplasmy, are lacking. Wider taxon sampling clarified the composition of the Austral group, but the conflicting signal meant analyses (e.g., morphological evolution, biogeographic) conditional on a well-supported phylogeny could not be performed.

  11. Comparative analysis of complete chloroplast genome sequence and inversion variation in Lasthenia burkei (Madieae, Asteraceae).

    Science.gov (United States)

    Walker, Joseph F; Zanis, Michael J; Emery, Nancy C

    2014-04-01

    Complete chloroplast genome studies can help resolve relationships among large, complex plant lineages such as Asteraceae. We present the first whole plastome from the Madieae tribe and compare its sequence variation to other chloroplast genomes in Asteraceae. We used high throughput sequencing to obtain the Lasthenia burkei chloroplast genome. We compared sequence structure and rates of molecular evolution in the small single copy (SSC), large single copy (LSC), and inverted repeat (IR) regions to those for eight Asteraceae accessions and one Solanaceae accession. The chloroplast sequence of L. burkei is 150 746 bp and contains 81 unique protein coding genes and 4 coding ribosomal RNA sequences. We identified three major inversions in the L. burkei chloroplast, all of which have been found in other Asteraceae lineages, and a previously unreported inversion in Lactuca sativa. Regions flanking inversions contained tRNA sequences, but did not have particularly high G + C content. Substitution rates varied among the SSC, LSC, and IR regions, and rates of evolution within each region varied among species. Some observed differences in rates of molecular evolution may be explained by the relative proportion of coding to noncoding sequence within regions. Rates of molecular evolution vary substantially within and among chloroplast genomes, and major inversion events may be promoted by the presence of tRNAs. Collectively, these results provide insight into different mechanisms that may promote intramolecular recombination and the inversion of large genomic regions in the plastome.

  12. Short-term effects of salt exposure on the maize chloroplast protein pattern.

    Science.gov (United States)

    Zörb, Christian; Herbst, Ramona; Forreiter, Christoph; Schubert, Sven

    2009-09-01

    It is of fundamental importance to understand the physiological differences leading to salt resistance and to get access to the molecular mechanisms underlying this physiological response. The aim of this work was to investigate the effects of short-term salt exposure on the proteome of maize chloroplasts in the initial phase of salt stress (up to 4 h). It could be shown that sodium ions accumulate quickly and excessively in chloroplasts in the initial phase of moderate salt stress. A change in the chloroplast protein pattern was observed without a change in water potential of the leaves. 2-DE revealed that 12 salt-responsive chloroplast proteins increased while eight chloroplast proteins decreased. Some of the maize chloroplast proteins such as CF1e and a Ca(2+)-sensing receptor show a rather transient response for the first 4 h of salt exposure. The enhanced abundance of the ferredoxin NADPH reductase, the 23 kDa polypeptide of the photosystem II, and the FtsH-like protein might reflect mechanism to attenuate the detrimental effects of Na(+) on the photosynthetic machinery. The observed transient increase and subsequent decrease of selected proteins may exhibit a counterbalancing effect of target proteins in this context. Intriguingly, several subunits of the CF1-CF0 complex are unequally affected, whereas others do not respond at all.

  13. A database of PCR primers for the chloroplast genomes of higher plants

    Science.gov (United States)

    Heinze, Berthold

    2007-01-01

    Background Chloroplast genomes evolve slowly and many primers for PCR amplification and analysis of chloroplast sequences can be used across a wide array of genera. In some cases 'universal' primers have been designed for the purpose of working across species boundaries. However, the essential information on these primer sequences is scattered throughout the literature. Results A database is presented here which assembles published primer information for chloroplast DNA. Additional primers were designed to fill gaps where little or no primer information could be found. Amplicons are either the genes themselves (typically useful in studies of sequence variation in higher-order phylogeny) or they are spacers, introns, and intergenic regions (for studies of phylogeographic patterns within and among species). The current list of 'generic' primers consists of more than 700 sequences. Wherever possible, we give the locations of the primers in the thirteen fully sequenced chloroplast genomes (Nicotiana tabacum, Atropa belladonna, Spinacia oleracea, Arabidopsis thaliana, Populus trichocarpa, Oryza sativa, Pinus thunbergii, Marchantia polymorpha, Zea mays, Oenothera elata, Acorus calamus, Eucalyptus globulus, Medicago trunculata). Conclusion The database described here is designed to serve as a resource for researchers who are venturing into the study of poorly described chloroplast genomes, whether for large- or small-scale DNA sequencing projects, to study molecular variation or to investigate chloroplast evolution. PMID:17326828

  14. A database of PCR primers for the chloroplast genomes of higher plants

    Directory of Open Access Journals (Sweden)

    Heinze Berthold

    2007-02-01

    Full Text Available Abstract Background Chloroplast genomes evolve slowly and many primers for PCR amplification and analysis of chloroplast sequences can be used across a wide array of genera. In some cases 'universal' primers have been designed for the purpose of working across species boundaries. However, the essential information on these primer sequences is scattered throughout the literature. Results A database is presented here which assembles published primer information for chloroplast DNA. Additional primers were designed to fill gaps where little or no primer information could be found. Amplicons are either the genes themselves (typically useful in studies of sequence variation in higher-order phylogeny or they are spacers, introns, and intergenic regions (for studies of phylogeographic patterns within and among species. The current list of 'generic' primers consists of more than 700 sequences. Wherever possible, we give the locations of the primers in the thirteen fully sequenced chloroplast genomes (Nicotiana tabacum, Atropa belladonna, Spinacia oleracea, Arabidopsis thaliana, Populus trichocarpa, Oryza sativa, Pinus thunbergii, Marchantia polymorpha, Zea mays, Oenothera elata, Acorus calamus, Eucalyptus globulus, Medicago trunculata. Conclusion The database described here is designed to serve as a resource for researchers who are venturing into the study of poorly described chloroplast genomes, whether for large- or small-scale DNA sequencing projects, to study molecular variation or to investigate chloroplast evolution.

  15. Chloroplasts activity and PAP-signaling regulate programmed cell death in Arabidopsis

    KAUST Repository

    Bruggeman, Quentin

    2016-01-09

    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. © 2016 American Society of Plant Biologists. All Rights Reserved.

  16. Arabidopsis VARIEGATED 3 encodes a chloroplast-targeted, zinc-finger protein required for chloroplast and palisade cell development

    DEFF Research Database (Denmark)

    Næsted, Henrik; Holm, Agnethe; Jenkins, Tom

    2004-01-01

    The stable, recessive Arabidopsis variegated 3 (var3) mutant exhibits a variegated phenotype due to somatic areas lacking or containing developmentally retarded chloroplasts and greatly reduced numbers of palisade cells. The VAR3 gene, isolated by transposon tagging, encodes the 85.9 kDa VAR3...... that pigment profiles are qualitatively similar in wild type and var3, although var3 accumulates lower levels of chlorophylls and carotenoids. These results indicate that VAR3 is a part of a protein complex required for normal chloroplast and palisade cell development....

  17. Intersectional gene flow between insular endemics of Ilex (Aquifoliaceae) on the Bonin Islands and the Ryukyu Islands.

    Science.gov (United States)

    Setoguchi, H; Watanabe, I

    2000-06-01

    Hybridization and introgression play important roles in plant evolution, and their occurrence on the oceanic islands provides good examples of plant speciation and diversification. Restriction fragment length polymorphisms (RFLPs) and trnL (UAA) 3'exon-trnF (GAA) intergenic spacer (IGS) sequences of chloroplast DNA (cpDNA), and the sequences of internal transcribed spacer (ITS) of nuclear ribosomal DNA were examined to investigate the occurrence of gene transfer in Ilex species on the Bonin Islands and the Ryukyu Islands in Japan. A gene phylogeny for the plastid genome is in agreement with the morphologically based taxonomy, whereas the nuclear genome phylogeny clusters putatively unrelated endemics both on the Bonin and the Ryukyu Islands. Intersectional hybridization and nuclear gene flow were independently observed in insular endemics of Ilex on both sets of islands without evidence of plastid introgression. Gene flow observed in these island systems can be explained by ecological features of insular endemics, i.e., limits of distribution range or sympatric distribution in a small land area.

  18. Plant DNA Detection from Grasshopper Guts: A Step-by-Step Protocol, from Tissue Preparation to Obtaining Plant DNA Sequences

    Directory of Open Access Journals (Sweden)

    Alina Avanesyan

    2014-02-01

    Full Text Available Premise of the study: A PCR-based method of identifying ingested plant DNA in gut contents of Melanoplus grasshoppers was developed. Although previous investigations have focused on a variety of insects, there are no protocols available for plant DNA detection developed for grasshoppers, agricultural pests that significantly influence plant community composition. Methods and Results: The developed protocol successfully used the noncoding region of the chloroplast trnL (UAA gene and was tested in several feeding experiments. Plant DNA was obtained at seven time points post-ingestion from whole guts and separate gut sections, and was detectable up to 12 h post-ingestion in nymphs and 22 h post-ingestion in adult grasshoppers. Conclusions: The proposed protocol is an effective, relatively quick, and low-cost method of detecting plant DNA from the grasshopper gut and its different sections. This has important applications, from exploring plant “movement” during food consumption, to detecting plant–insect interactions.

  19. Plant DNA detection from grasshopper guts: A step-by-step protocol, from tissue preparation to obtaining plant DNA sequences1

    Science.gov (United States)

    Avanesyan, Alina

    2014-01-01

    • Premise of the study: A PCR-based method of identifying ingested plant DNA in gut contents of Melanoplus grasshoppers was developed. Although previous investigations have focused on a variety of insects, there are no protocols available for plant DNA detection developed for grasshoppers, agricultural pests that significantly influence plant community composition. • Methods and Results: The developed protocol successfully used the noncoding region of the chloroplast trnL (UAA) gene and was tested in several feeding experiments. Plant DNA was obtained at seven time points post-ingestion from whole guts and separate gut sections, and was detectable up to 12 h post-ingestion in nymphs and 22 h post-ingestion in adult grasshoppers. • Conclusions: The proposed protocol is an effective, relatively quick, and low-cost method of detecting plant DNA from the grasshopper gut and its different sections. This has important applications, from exploring plant “movement” during food consumption, to detecting plant–insect interactions. PMID:25202604

  20. Determination of locust bean gum and guar gum by polymerase chain reaction and restriction fragment length polymorphism analysis.

    Science.gov (United States)

    Meyer, K; Rosa, C; Hischenhuber, C; Meyer, R

    2001-01-01

    A polymerase chain reaction (PCR) was developed to differentiate the thickening agents locust bean gum (LBG) and the cheaper guar gum in finished food products. Universal primers for amplification of the intergenic spacer region between trnL 3' (UAA) exon and trnF (GAA) gene in the chloroplast (cp) genome and subsequent restriction analysis were applied to differentiate guar gum and LBG. The presence of guar gum powder added to LBG powder was detectable. Based on data obtained from sequencing this intergenic spacer region, a second PCR method for the specific detection of guar gum DNA was also developed. This assay detected guar gum powder in LBG in amounts as low as 1% (w/w). Both methods successfully detected guar gum and/or LBG in ice cream stabilizers and in foodstuffs, such as dairy products, ice cream, dry seasoning mixes, a finished roasting sauce, and a fruit jelly product, but not in products with highly degraded DNA, such as tomato ketchup and sterilized chocolate cream. Both methods detected guar gum and LBG in ice cream and fresh cheese at levels <0.1%.

  1. RNA transcription in isolated chloroplasts during senescence and rejuvenation of intact cotyledons of CUCURBITA PEPO L. (ZUCCHINI)

    International Nuclear Information System (INIS)

    Mishev, K.; Ananiev, E.; Denev, L.; Radeva, G.

    2006-01-01

    RNA transcription was studied in intact chloroplasts isolated from cotyledons of Cucurbita pepoL. (zucchini) during their growth and development including natural senescence and rejuvenation. Rejuvenation of cotyledons was studied after decapitation of the epicotyl above the senescing yellow cotyledons. Maximal incorporation of [32P] UTP into overall chloroplast RNA was measured two days after exposure of seedlings to light (day 6 th after the onset of germination), followed by a gradual decrease reaching minimal values at the age of 25-28 days when cotyledons began to yellow and eventually die. Rejuvenation of cotyledons completely restored chloroplast RNA synthesis and fifteen days after decapitation (at the age of 40 days), the values of chloroplast transcription even exceeded that of the maximal transcriptional activity in young cotyledons. Inhibitory analysis with tagetitoxin (a specific inhibitor of plastid encoded chloroplast RNA polymerase (PEP)) showed that in young and rejuvenated cotyledons about 85% of chloroplast RNA polymerase activity was due to PEP and only 15% corresponded to the nuclear encoded plastid RNA polymerase (NEP). Definite regions of two chloroplast encoded genes were amplified by means of PCR technique using specific DNA primers for Rubisco large subunit gene (rbcL) and the housekeeping gene for chloroplast 16S rRNA as well as chloroplast DNA as a template. The appropriate lengths of the amplified DNA fragments were checked by restriction analysis

  2. Chloroplasts as source and target of cellular redox regulation: a discussion on chloroplast redox signals in the context of plant physiology.

    Science.gov (United States)

    Baier, Margarete; Dietz, Karl-Josef

    2005-06-01

    During the evolution of plants, chloroplasts have lost the exclusive genetic control over redox regulation and antioxidant gene expression. Together with many other genes, all genes encoding antioxidant enzymes and enzymes involved in the biosynthesis of low molecular weight antioxidants were transferred to the nucleus. On the other hand, photosynthesis bears a high risk for photo-oxidative damage. Concomitantly, an intricate network for mutual regulation by anthero- and retrograde signals has emerged to co-ordinate the activities of the different genetic and metabolic compartments. A major focus of recent research in chloroplast regulation addressed the mechanisms of redox sensing and signal transmission, the identification of regulatory targets, and the understanding of adaptation mechanisms. In addition to redox signals communicated through signalling cascades also used in pathogen and wounding responses, specific chloroplast signals control nuclear gene expression. Signalling pathways are triggered by the redox state of the plastoquinone pool, the thioredoxin system, and the acceptor availability at photosystem I, in addition to control by oxolipins, tetrapyrroles, carbohydrates, and abscisic acid. The signalling function is discussed in the context of regulatory circuitries that control the expression of antioxidant enzymes and redox modulators, demonstrating the principal role of chloroplasts as the source and target of redox regulation.

  3. Insights from the complete chloroplast genome into the evolution of Sesamum indicum L.

    Directory of Open Access Journals (Sweden)

    Haiyang Zhang

    Full Text Available Sesame (Sesamum indicum L. is one of the oldest oilseed crops. In order to investigate the evolutionary characters according to the Sesame Genome Project, apart from sequencing its nuclear genome, we sequenced the complete chloroplast genome of S. indicum cv. Yuzhi 11 (white seeded using Illumina and 454 sequencing. Comparisons of chloroplast genomes between S. indicum and the 18 other higher plants were then analyzed. The chloroplast genome of cv. Yuzhi 11 contains 153,338 bp and a total of 114 unique genes (KC569603. The number of chloroplast genes in sesame is the same as that in Nicotiana tabacum, Vitis vinifera and Platanus occidentalis. The variation in the length of the large single-copy (LSC regions and inverted repeats (IR in sesame compared to 18 other higher plant species was the main contributor to size variation in the cp genome in these species. The 77 functional chloroplast genes, except for ycf1 and ycf2, were highly conserved. The deletion of the cp ycf1 gene sequence in cp genomes may be due either to its transfer to the nuclear genome, as has occurred in sesame, or direct deletion, as has occurred in Panax ginseng and Cucumis sativus. The sesame ycf2 gene is only 5,721 bp in length and has lost about 1,179 bp. Nucleotides 1-585 of ycf2 when queried in BLAST had hits in the sesame draft genome. Five repeats (R10, R12, R13, R14 and R17 were unique to the sesame chloroplast genome. We also found that IR contraction/expansion in the cp genome alters its rate of evolution. Chloroplast genes and repeats display the signature of convergent evolution in sesame and other species. These findings provide a foundation for further investigation of cp genome evolution in Sesamum and other higher plants.

  4. Fatty acid synthesis by spinach chloroplasts, 2. The path from PGA to fatty acids

    Energy Technology Data Exchange (ETDEWEB)

    Yamada, Mitsuhiro; Nakamura, Yasunori [Tokyo Univ. (Japan). Coll. of General Education

    1975-02-01

    By incorporation of /sup 3/H/sub 2/O into the fatty acid chain in the presence of unlabelled precursor, we showed that fatty acids are synthesized from PGA, PEP and pyruvate by intact spinach chloroplasts in the light. /sup 13/C-tracer experiments confirmed that 1-C of pyruvate is decarboxylated and 2-C is incorporated into fatty acids by the chloroplasts. The patterns of fatty acids synthesized from PGA and pyruvate were the same as that from acetate. The highest rate of fatty acid synthesis was reached at the physiological concentration of PGA (3 mM) and pyruvate (1 mM). These results indicate the operation of the following path in the chloroplasts in light: PGA..-->..PEP..-->..pyruvate..-->..acetylCoA..-->..fatty acids. Since citrate and OAA were much less active and malate and glyoxylate were inert as precursors for fatty acid synthesis, PEP or pyruvate carboxylation, citrate lyase reaction and malate synthetase reaction are not involved in the formation of acetylCoA and fatty acids. Since pyruvate was much more effective as a substrate for fatty acid synthesis than lactate, acetaldehyde or acetate, direct decarboxylation path is considered to be the primary path from pyruvate to acetylCoA. The insignificant effect of chloroplast-washing on fatty acid synthesis from PGA and pyruvate indicates that the glycolytic path from PGA to pyruvate is associated with the chloroplasts. Since pyruvate was more effectively incorporated into fatty acids than acetylCoA, it is unlikely that pyruvate decarboxylation to acetylCoA is due to mitochondria contaminating the chloroplast preparation. On the basis of measurements of /sup 3/H/sub 2/O incorporation in the light and dark, the activity of fatty acid synthesis in spincah leaves appears to be shared by the activities in chloroplasts (87%) and other organelles (13%).

  5. A simple low-cost microcontroller-based photometric instrument for monitoring chloroplast movement.

    Science.gov (United States)

    Berg, Robert; Königer, Martina; Schjeide, Brit-Maren; Dikmak, George; Kohler, Susan; Harris, Gary C

    2006-03-01

    A new microcontroller-based photometric instrument for monitoring blue light dependent changes in leaf transmission (chloroplast movement) was developed based on a modification of the double-beam technique developed by Walzcak and Gabrys [(1980) Photosynthetica 14: 65-72]. A blue and red bicolor light emitting diode (LED) provided both a variable intensity blue actinic light and a low intensity red measuring beam. A phototransistor detected the intensity of the transmitted measuring light. An inexpensive microcontroller independently and precisely controlled the light emission of the bicolor LED. A typical measurement event involved turning off the blue actinic light for 100 mus to create a narrow temporal window for turning on and measuring the transmittance of the red light. The microcontroller was programmed using LogoChip Logo (http://www.wellesley.edu/Physics/Rberg/logochip/) to record fluence rate response curves. Laser scanning confocal microscopy was utilized to correlate the changes in leaf transmission with intercellular chloroplast position. In the dark, the chloroplasts in the spongy mesophyll exhibited no evident asymmetries in their distribution, however, in the palisade layer the cell surface in contact with the overlying epidermis was devoid of chloroplasts. The low light dependent decrease in leaf transmittance in dark acclimated leaves was correlated with the movement of chloroplasts within the palisade layer into the regions previously devoid of chloroplasts. Changes in leaf transmittance were evident within one minute following the onset of illumination. Minimal leaf transmittance was correlated with chloroplasts having retreated from cell surfaces perpendicular to the incident light (avoidance reaction) in both spongy and palisade layers.

  6. The Complete Chloroplast Genome Sequences of Six Rehmannia Species

    Directory of Open Access Journals (Sweden)

    Shuyun Zeng

    2017-03-01

    Full Text Available Rehmannia is a non-parasitic genus in Orobanchaceae including six species mainly distributed in central and north China. Its phylogenetic position and infrageneric relationships remain uncertain due to potential hybridization and polyploidization. In this study, we sequenced and compared the complete chloroplast genomes of six Rehmannia species using Illumina sequencing technology to elucidate the interspecific variations. Rehmannia plastomes exhibited typical quadripartite and circular structures with good synteny of gene order. The complete genomes ranged from 153,622 bp to 154,055 bp in length, including 133 genes encoding 88 proteins, 37 tRNAs, and 8 rRNAs. Three genes (rpoA, rpoC2, accD have potentially experienced positive selection. Plastome size variation of Rehmannia was mainly ascribed to the expansion and contraction of the border regions between the inverted repeat (IR region and the single-copy (SC regions. Despite of the conserved structure in Rehmannia plastomes, sequence variations provide useful phylogenetic information. Phylogenetic trees of 23 Lamiales species reconstructed with the complete plastomes suggested that Rehmannia was monophyletic and sister to the clade of Lindenbergia and the parasitic taxa in Orobanchaceae. The interspecific relationships within Rehmannia were completely different with the previous studies. In future, population phylogenomic works based on plastomes are urgently needed to clarify the evolutionary history of Rehmannia.

  7. Radiation inactivation analysis of chloroplast CF0-CF1 ATPase

    International Nuclear Information System (INIS)

    Wang, M.Y.; Chien, L.F.; Pan, R.L.

    1988-01-01

    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 3 (2-) and CO 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

  8. Stromal serine protein kinase activity in spinach chloroplasts

    International Nuclear Information System (INIS)

    Cortez, N.; Lucero, H.A.; Vallejos, R.H.

    1987-01-01

    At least twelve 32 P-labeled stromal proteins were detected by electrophoresis under denaturing conditions when intact chloroplasts were incubated with 32 Pi, in the light but only three were detected in the presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) or in the dark. Incubation of isolated stroma with [gamma- 32 P]ATP resulted in the preferential phosphorylation of one of them, a 70-kDa polypeptide, in serine residues. Thylakoid membranes in the dark promoted the phosphorylation of two additional stromal polypeptides of 55 and 40 kDa. Illumination during the phosphorylation of stroma in the presence of thylakoids stimulated severalfold the labeling of the 40-kDa polypeptide but not when DCMU was added. The protein kinase activity present in isolated stroma phosphorylated exogenous substrates like histone III, phosvitin, histone II, and casein with specific activities of 3, 1.8, 0.7, and 0.2 pmol X mg-1 X min-1. Histone III polypeptides were phosphorylated differently by stroma and by thylakoids in the dark. Moreover, histone III phosphorylated by thylakoids in the dark yielded a pattern of phosphopeptides after V8 protease treatment that was different from the pattern obtained when histone III was phosphorylated by stroma

  9. Two complete chloroplast genome sequences of Cannabis sativa varieties.

    Science.gov (United States)

    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.

  10. The complete chloroplast genome sequence of Abies nephrolepis (Pinaceae: Abietoideae

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    Dong-Keun Yi

    2016-06-01

    Full Text Available The plant chloroplast (cp genome has maintained a relatively conserved structure and gene content throughout evolution. Cp genome sequences have been used widely for resolving evolutionary and phylogenetic issues at various taxonomic levels of plants. Here, we report the complete cp genome of Abies nephrolepis. The A. nephrolepis cp genome is 121,336 base pairs (bp in length including a pair of short inverted repeat regions (IRa and IRb of 139 bp each separated by a small single copy (SSC region of 54,323 bp (SSC and a large single copy region of 66,735 bp (LSC. It contains 114 genes, 68 of which are protein coding genes, 35 tRNA and four rRNA genes, six open reading frames, and one pseudogene. Seventeen repeat units and 64 simple sequence repeats (SSR have been detected in A. nephrolepis cp genome. Large IR sequences locate in 42-kb inversion points (1186 bp. The A. nephrolepis cp genome is identical to Abies koreana’s which is closely related to taxa. Pairwise comparison between two cp genomes revealed 140 polymorphic sites in each. Complete cp genome sequence of A. nephrolepis has a significant potential to provide information on the evolutionary pattern of Abietoideae and valuable data for development of DNA markers for easy identification and classification.

  11. Chloroplast DNA footprints of postglacial recolonization by oaks

    Science.gov (United States)

    Petit, Rémy J.; Pineau, Emmanuel; Demesure, Brigitte; Bacilieri, Roberto; Ducousso, Alexis; Kremer, Antoine

    1997-01-01

    Recolonization of Europe by forest tree species after the last glaciation is well documented in the fossil pollen record. This spread may have been achieved at low densities by rare events of long-distance dispersal, rather than by a compact wave of advance, generating a patchy genetic structure through founder effects. In long-lived oak species, this structure could still be discernible by using maternally transmitted genetic markers. To test this hypothesis, a fine-scale study of chloroplast DNA (cpDNA) variability of two sympatric oak species was carried out in western France. The distributions of six cpDNA length variants were analyzed at 188 localities over a 200 × 300 km area. A cpDNA map was obtained by applying geostatistics methods to the complete data set. Patches of several hundred square kilometers exist which are virtually fixed for a single haplotype for both oak species. This local systematic interspecific sharing of the maternal genome strongly suggests that long-distance seed dispersal events followed by interspecific exchanges were involved at the time of colonization, about 10,000 years ago. PMID:11038572

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

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    Adyasha Bharati

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

  13. Complete chloroplast genome sequence of a tree fern Alsophila spinulosa: insights into evolutionary changes in fern chloroplast genomes.

    Science.gov (United States)

    Gao, Lei; Yi, Xuan; Yang, Yong-Xia; Su, Ying-Juan; Wang, Ting

    2009-06-11

    Ferns have generally been neglected in studies of chloroplast genomics. Before this study, only one polypod and two basal ferns had their complete chloroplast (cp) genome reported. Tree ferns represent an ancient fern lineage that first occurred in the Late Triassic. In recent phylogenetic analyses, tree ferns were shown to be the sister group of polypods, the most diverse group of living ferns. Availability of cp genome sequence from a tree fern will facilitate interpretation of the evolutionary changes of fern cp genomes. Here we have sequenced the complete cp genome of a scaly tree fern Alsophila spinulosa (Cyatheaceae). The Alsophila cp genome is 156,661 base pairs (bp) in size, and has a typical quadripartite structure with the large (LSC, 86,308 bp) and small single copy (SSC, 21,623 bp) regions separated by two copies of an inverted repeat (IRs, 24,365 bp each). This genome contains 117 different genes encoding 85 proteins, 4 rRNAs and 28 tRNAs. Pseudogenes of ycf66 and trnT-UGU are also detected in this genome. A unique trnR-UCG gene (derived from trnR-CCG) is found between rbcL and accD. The Alsophila cp genome shares some unusual characteristics with the previously sequenced cp genome of the polypod fern Adiantum capillus-veneris, including the absence of 5 tRNA genes that exist in most other cp genomes. The genome shows a high degree of synteny with that of Adiantum, but differs considerably from two basal ferns (Angiopteris evecta and Psilotum nudum). At one endpoint of an ancient inversion we detected a highly repeated 565-bp-region that is absent from the Adiantum cp genome. An additional minor inversion of the trnD-GUC, which is possibly shared by all ferns, was identified by comparison between the fern and other land plant cp genomes. By comparing four fern cp genome sequences it was confirmed that two major rearrangements distinguish higher leptosporangiate ferns from basal fern lineages. The Alsophila cp genome is very similar to that of the

  14. Complete chloroplast genome sequence of a tree fern Alsophila spinulosa: insights into evolutionary changes in fern chloroplast genomes

    Directory of Open Access Journals (Sweden)

    Yang Yong-Xia

    2009-06-01

    Full Text Available Abstract Background Ferns have generally been neglected in studies of chloroplast genomics. Before this study, only one polypod and two basal ferns had their complete chloroplast (cp genome reported. Tree ferns represent an ancient fern lineage that first occurred in the Late Triassic. In recent phylogenetic analyses, tree ferns were shown to be the sister group of polypods, the most diverse group of living ferns. Availability of cp genome sequence from a tree fern will facilitate interpretation of the evolutionary changes of fern cp genomes. Here we have sequenced the complete cp genome of a scaly tree fern Alsophila spinulosa (Cyatheaceae. Results The Alsophila cp genome is 156,661 base pairs (bp in size, and has a typical quadripartite structure with the large (LSC, 86,308 bp and small single copy (SSC, 21,623 bp regions separated by two copies of an inverted repeat (IRs, 24,365 bp each. This genome contains 117 different genes encoding 85 proteins, 4 rRNAs and 28 tRNAs. Pseudogenes of ycf66 and trnT-UGU are also detected in this genome. A unique trnR-UCG gene (derived from trnR-CCG is found between rbcL and accD. The Alsophila cp genome shares some unusual characteristics with the previously sequenced cp genome of the polypod fern Adiantum capillus-veneris, including the absence of 5 tRNA genes that exist in most other cp genomes. The genome shows a high degree of synteny with that of Adiantum, but differs considerably from two basal ferns (Angiopteris evecta and Psilotum nudum. At one endpoint of an ancient inversion we detected a highly repeated 565-bp-region that is absent from the Adiantum cp genome. An additional minor inversion of the trnD-GUC, which is possibly shared by all ferns, was identified by comparison between the fern and other land plant cp genomes. Conclusion By comparing four fern cp genome sequences it was confirmed that two major rearrangements distinguish higher leptosporangiate ferns from basal fern lineages. The

  15. Dissecting the chloroplast proteome of chickpea (Cicer arietinum L.) provides new insights into classical and non-classical functions.

    Science.gov (United States)

    Lande, Nilesh Vikram; Subba, Pratigya; Barua, Pragya; Gayen, Dipak; Keshava Prasad, T S; Chakraborty, Subhra; Chakraborty, Niranjan

    2017-08-08

    Chloroplast, the energy organelle unique to plant cells, is a dynamic entity which integrates an array of metabolic pathways and serves as first level for energy conversion for the entire ecological hierarchy. Increasing amount of sequence data and evolution of mass spectrometric approaches has opened up new avenues for opportune exploration of the global proteome of this organelle. In our study, we aimed at generation of a comprehensive catalogue of chloroplast proteins in a grain legume, chickpea and provided a reference proteome map. To accurately assign the identified proteins, purity of chloroplast-enriched fraction was stringently monitored by multiple chemical and immunological indexes, besides pigment and enzyme analyses. The proteome analysis led to the identification of 2451 proteins, including 27 isoforms, which include predicted and novel chloroplast constituents. The identified proteins were validated through their sequence analysis. Extensive sequence based localization prediction revealed more than 50% proteins to be chloroplast resident by at least two different algorithms. Chromosomal distribution of identified proteins across nuclear and chloroplast genome unveiled the presence of 55 chloroplast encoded gene. In depth comparison of our dataset with the non-redundant set of chloroplast proteins identified so far across other species revealed novel as well as overlapping candidates. Pulses add large amount of nitrogen to the soil and has very low water footprint and therefore, contributes to fortification of sustainable agriculture. Chickpea is one of the earliest cultivated legumes and serves as an energy and protein source for humans and animals. Chloroplasts are the unique organelles which conduct photosynthesis. Investigation on chloroplast proteome is of particular significance, especially to plant biologists, as it would allow a better understanding of chloroplast function in plants. Generation of a saturated proteome map would not only

  16. Expression of the Native Cholera Toxin B Subunit Gene and Assembly as Functional Oligomers in Transgenic Tobacco Chloroplasts

    Science.gov (United States)

    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

  17. Polyuridylylation and processing of transcripts from multiple gene minicircles in chloroplasts of the dinoflagellate Amphidinium carterae

    KAUST Repository

    Barbrook, Adrian C.

    2012-05-05

    Although transcription and transcript processing in the chloroplasts of plants have been extensively characterised, the RNA metabolism of other chloroplast lineages across the eukaryotes remains poorly understood. In this paper, we use RT-PCR to study transcription and transcript processing in the chloroplasts of Amphidinium carterae, a model peridinin-containing dinoflagellate. These organisms have a highly unusual chloroplast genome, with genes located on multiple small \\'minicircle\\' elements, and a number of idiosyncratic features of RNA metabolism including transcription via a rolling circle mechanism, and 3′ terminal polyuridylylation of transcripts. We demonstrate that transcription occurs in A. carterae via a rolling circle mechanism, as previously shown in the dinoflagellate Heterocapsa, and present evidence for the production of both polycistronic and monocistronic transcripts from A. carterae minicircles, including several regions containing ORFs previously not known to be expressed. We demonstrate the presence of both polyuridylylated and non-polyuridylylated transcripts in A. carterae, and show that polycistronic transcripts can be terminally polyuridylylated. We present a model for RNA metabolism in dinoflagellate chloroplasts where long polycistronic precursors are processed to form mature transcripts. Terminal polyuridylylation may mark transcripts with the correct 3′ end. © 2012 Springer Science+Business Media B.V.

  18. Light-stimulated accumulation of transcripts of nuclear and chloroplast genes for ribulosebisphosphate carboxylase

    Energy Technology Data Exchange (ETDEWEB)

    Smith, S M; Ellis, R J

    1981-01-01

    The chloroplast enzyme, ribulosebisphosphate carboxylase, consists of large subunit polypeptides encoded in the chloroplast genome and small subunit polypeptides encoded in the nuclear genome. Cloned DNA complementary to the small subunit mRNA hybridizes to a single RNA species of 900-1000 nucleotides in both total and poly(A)-containing RNA from leaves of Pisum sativum, but does not hybridize to chloroplast RNA. Small subunit cDNA hybridizes to at least three RNA species from nuclei, two of which are of higher molecular weight than the mature mRNA. A cloned large subunit DNA sequence hybridizes to a single species of Pisum chloroplast RNA containing approximately 1700 nucleotides, but does not hybridize to nuclear RNA. The light-stimulation of carboxylase accumulation reflects increases in the amounts of transcripts for both subunits in total leaf RNA. Transcripts of the small subunit gene are more abundant in nuclear RNA from light-grown leaves than in that from dark-grown leaves. These results suggest that the stimulation of carboxylase accumulation by light is mediated at the level of either transcription or RNA turnover in both nucleus and chloroplast.

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

    Science.gov (United States)

    Jouhet, Juliette; Gray, John C

    2009-10-01

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

  20. Chloroplast-Derived Vaccine Antigens and Biopharmaceuticals: Expression, Folding, Assembly and Functionality

    Science.gov (United States)

    Chebolu, S.; Daniell, H.

    2009-01-01

    Chloroplast genetic engineering offers several advantages, including high levels of transgene expression, transgene containment via maternal inheritance, and multi-gene expression in a single transformation event. Oral delivery is facilitated by hyperexpression of vaccine antigens against cholera, tetanus, anthrax, plague, or canine parvovirus (4%–31% of total soluble protein, TSP) in transgenic chloroplasts (leaves) or non-green plastids (carrots, tomato) as well as the availability of antibiotic free selectable markers or the ability to excise selectable marker genes. Hyperexpression of several therapeutic proteins, including human serum albumin (11.1% TSP), somatotropin (7% TSP), interferon-alpha (19% TSP), interferon-gamma (6% TSP), and antimicrobial peptide (21.5% TSP), facilitates efficient and economic purification. Also, the presence of chaperones and enzymes in chloroplasts facilitates assembly of complex multisubunit proteins and correct folding of human blood proteins with proper disulfide bonds. Functionality of chloroplast-derived vaccine antigens and therapeutic proteins has been demonstrated by several assays, including the macrophage lysis assay, GM1-ganglioside binding assay, protection of HeLA cells or human lung carcinoma cells against encephalomyocarditis virus, systemic immune response, protection against pathogen challenge, and growth or inhibition of cell cultures. Purification of human proinsulin has been achieved using novel purification strategies (inverse temperature transition property) that do not require expensive column chromatography techniques. Thus, transgenic chloroplasts are ideal bioreactors for production of functional human and animal therapeutic proteins in an environmentally friendly manner. PMID:19401820

  1. The complete chloroplast genome of Cinnamomum camphora and its comparison with related Lauraceae species

    Directory of Open Access Journals (Sweden)

    Caihui Chen

    2017-09-01

    Full Text Available Cinnamomum camphora, a member of the Lauraceae family, is a valuable aromatic and timber tree that is indigenous to the south of China and Japan. All parts of Cinnamomum camphora have secretory cells containing different volatile chemical compounds that are utilized as herbal medicines and essential oils. Here, we reported the complete sequencing of the chloroplast genome of Cinnamomum camphora using illumina technology. The chloroplast genome of Cinnamomum camphora is 152,570 bp in length and characterized by a relatively conserved quadripartite structure containing a large single copy region of 93,705 bp, a small single copy region of 19,093 bp and two inverted repeat (IR regions of 19,886 bp. Overall, the genome contained 123 coding regions, of which 15 were repeated in the IR regions. An analysis of chloroplast sequence divergence revealed that the small single copy region was highly variable among the different genera in the Lauraceae family. A total of 40 repeat structures and 83 simple sequence repeats were detected in both the coding and non-coding regions. A phylogenetic analysis indicated that Calycanthus is most closely related to Lauraceae, both being members of Laurales, which forms a sister group to Magnoliids. The complete sequence of the chloroplast of Cinnamomum camphora will aid in in-depth taxonomical studies of the Lauraceae family in the future. The genetic sequence information will also have valuable applications for chloroplast genetic engineering.

  2. Overoxidation of chloroplast 2-Cys peroxiredoxins: balancing toxic and signaling activities of hydrogen peroxide

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    Leonor ePuerto-Galán

    2013-08-01

    Full Text Available 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 (ROS, 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.

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

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    John F Allen

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

  4. Enzyme-Triggered Defined Protein Nanoarrays: Efficient Light-Harvesting Systems to Mimic Chloroplasts.

    Science.gov (United States)

    Zhao, Linlu; Zou, Haoyang; Zhang, Hao; Sun, Hongcheng; Wang, Tingting; Pan, Tiezheng; Li, Xiumei; Bai, Yushi; Qiao, Shanpeng; Luo, Quan; Xu, Jiayun; Hou, Chunxi; Liu, Junqiu

    2017-01-24

    The elegance and efficiency by which chloroplasts harvest solar energy and conduct energy transfer have been a source of inspiration for chemists to mimic such process. However, precise manipulation to obtain orderly arranged antenna chromophores in constructing artificial chloroplast mimics was a great challenge, especially from the structural similarity and bioaffinity standpoints. Here we reported a design strategy that combined covalent and noncovalent interactions to prepare a protein-based light-harvesting system to mimic chloroplasts. Cricoid stable protein one (SP1) was utilized as a building block model. Under enzyme-triggered covalent protein assembly, mutant SP1 with tyrosine (Tyr) residues at the designated sites can couple together to form nanostructures. Through controlling the Tyr sites on the protein surface, we can manipulate the assembly orientation to respectively generate 1D nanotubes and 2D nanosheets. The excellent stability endowed the self-assembled protein architectures with promising applications. We further integrated quantum dots (QDs) possessing optical and electronic properties with the 2D nanosheets to fabricate chloroplast mimics. By attaching different sized QDs as donor and acceptor chromophores to the negatively charged surface of SP1-based protein nanosheets via electrostatic interactions, we successfully developed an artificial light-harvesting system. The assembled protein nanosheets structurally resembled the natural thylakoids, and the QDs can achieve pronounced FRET phenomenon just like the chlorophylls. Therefore, the coassembled system was meaningful to explore the photosynthetic process in vitro, as it was designed to mimic the natural chloroplast.

  5. The effect of UV-B radiation on chloroplast translation in Pisum sativum

    International Nuclear Information System (INIS)

    Raab, M.M.; Jagendorf, A.T.

    1990-01-01

    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 ( 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 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 - 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

  6. Purification of intact chloroplasts from marine plant Posidonia oceanica suitable for organelle proteomics.

    Science.gov (United States)

    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. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. Effects of lead on enzymes of porphyrine biosynthesis in chloroplasts and erythrocytes

    Energy Technology Data Exchange (ETDEWEB)

    Hampp, R.; Kriebitzsch, C.; Ziegler, H.

    1974-01-01

    Two enzymes of the chlorophyll biosynthesis pathway, delta-aminolevulinic acid dehydratase (ALAD) and prophobilinogenase (PBGA), show a pronounced sensitivity to lead ion, as was shown in isolated chloroplasts of spinach. It has been reported by several authors that the activity of ALAD involved in the hemoglobine-biosynthesis in erythrocytes is also very sensitive to lead ions. Spinach chloroplasts were isolated and sonicated and the enzyme activity tested. Calf blood was collected with heparin and kept at 0/sup 0/C until enzyme determination. Hemolyzed erythrocytes (rapid freezing and thawing twice) were used as the source of enzymes. The incubation mixture was the same as for chloroplasts; the hemoglobin content per test was about 44 mg (ALAD) and 91 mg (PBGA). ALAD in erythrocytes is somewhat more sensitive to lead ions than ALAD in chloroplasts. PBGA in erythrocytes is also inhibited by Pb/sup 2 +/ ions, again more than the chloroplast enzyme. At all concentrations of Pb/sup 2 +/ checked in our experiments the percentage of inhibition was higher with PBGA. 3 references, 1 figure.

  8. Chloroplast genome evolution in early diverged leptosporangiate ferns.

    Science.gov (United States)

    Kim, Hyoung Tae; Chung, Myong Gi; Kim, Ki-Joong

    2014-05-01

    In this study, the chloroplast (cp) genome sequences from three early diverged leptosporangiate ferns were completed and analyzed in order to understand the evolution of the genome of the fern lineages. The complete cp genome sequence of Osmunda cinnamomea (Osmundales) was 142,812 base pairs (bp). The cp genome structure was similar to that of eusporangiate ferns. The gene/intron losses that frequently occurred in the cp genome of leptosporangiate ferns were not found in the cp genome of O. cinnamomea. In addition, putative RNA editing sites in the cp genome were rare in O. cinnamomea, even though the sites were frequently predicted to be present in leptosporangiate ferns. The complete cp genome sequence of Diplopterygium glaucum (Gleicheniales) was 151,007 bp and has a 9.7 kb inversion between the trnL-CAA and trnVGCA genes when compared to O. cinnamomea. Several repeated sequences were detected around the inversion break points. The complete cp genome sequence of Lygodium japonicum (Schizaeales) was 157,142 bp and a deletion of the rpoC1 intron was detected. This intron loss was shared by all of the studied species of the genus Lygodium. The GC contents and the effective numbers of codons (ENCs) in ferns varied significantly when compared to seed plants. The ENC values of the early diverged leptosporangiate ferns showed intermediate levels between eusporangiate and core leptosporangiate ferns. However, our phylogenetic tree based on all of the cp gene sequences clearly indicated that the cp genome similarity between O. cinnamomea (Osmundales) and eusporangiate ferns are symplesiomorphies, rather than synapomorphies. Therefore, our data is in agreement with the view that Osmundales is a distinct early diverged lineage in the leptosporangiate ferns.

  9. Towards resolving Lamiales relationships: insights from rapidly evolving chloroplast sequences

    Directory of Open Access Journals (Sweden)

    Heubl Günther

    2010-11-01

    Full Text Available Abstract 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

  10. Towards resolving Lamiales relationships: insights from rapidly evolving chloroplast sequences

    Science.gov (United States)

    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

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

    Directory of Open Access Journals (Sweden)

    Susumu eUehara

    2016-02-01

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

  12. The novel chloroplast outer membrane kinase KOC1 is a required component of the plastid protein import machinery.

    Science.gov (United States)

    Zufferey, Mónica; Montandon, Cyrille; Douet, Véronique; Demarsy, Emilie; Agne, Birgit; Baginsky, Sacha; Kessler, Felix

    2017-04-28

    The biogenesis and maintenance of cell organelles such as mitochondria and chloroplasts require the import of many proteins from the cytosol, a process that is controlled by phosphorylation. In the case of chloroplasts, the import of hundreds of different proteins depends on translocons at the outer and inner chloroplast membrane (TOC and TIC, respectively) complexes. The essential protein TOC159 functions thereby as an import receptor. It has an N-terminal acidic (A-) domain that extends into the cytosol, controls receptor specificity, and is highly phosphorylated in vivo However, kinases that phosphorylate the TOC159 A-domain to enable protein import have remained elusive. Here, using co-purification with TOC159 from Arabidopsis , we discovered a novel component of the chloroplast import machinery, the regulatory kinase at the outer chloroplast membrane 1 (KOC1). We found that KOC1 is an integral membrane protein facing the cytosol and stably associates with TOC. Moreover, KOC1 phosphorylated the A-domain of TOC159 in vitro , and in mutant koc1 chloroplasts, preprotein import efficiency was diminished. koc1 Arabidopsis seedlings had reduced survival rates after transfer from the dark to the light in which protein import into plastids is required to rapidly complete chloroplast biogenesis. In summary, our data indicate that KOC1 is a functional component of the TOC machinery that phosphorylates import receptors, supports preprotein import, and contributes to efficient chloroplast biogenesis. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  13. β-Carotene as a factor in the reconstitution of cyclic phospho rylation in damaged chloroplast membranes

    Directory of Open Access Journals (Sweden)

    Anna Tukendorf

    2014-01-01

    Full Text Available Phenazine methosulphate mediated cyclic phosphorylation suppressed by heptane extraction or galactolipase treatment of spinach chloroplasts is restored by β -carotene, in 100% and 50%, respectively. Xanthophylls are not able to reconstitute this reaction. β-Carotene replaces galactolipids in reactivation of galactolipase treated chloroplasts, indicating a nonspecific effect of lipids in photosystem I dependent reactions.

  14. Formation and Change of Chloroplast-Located Plant Metabolites in Response to Light Conditions

    Directory of Open Access Journals (Sweden)

    Yiyong Chen

    2018-02-01

    Full Text Available Photosynthesis is the central energy conversion process for plant metabolism and occurs within mature chloroplasts. Chloroplasts are also the site of various metabolic reactions involving amino acids, lipids, starch, and sulfur, as well as where the production of some hormones takes place. Light is one of the most important environmental factors, acting as an essential energy source for plants, but also as an external signal influencing their growth and development. Plants experience large fluctuations in the intensity and spectral quality of light, and many attempts have been made to improve or modify plant metabolites by treating them with different light qualities (artificial lighting or intensities. In this review, we discuss how changes in light intensity and wavelength affect the formation of chloroplast-located metabolites in plants.

  15. Effect of gamma irradiation of some quantitative indices of wheat and the ultrastructure of wheat chloroplasts

    Energy Technology Data Exchange (ETDEWEB)

    Petrovic, J; Marek, J; Hraska, S [Vysoka Skola Polnohospodarska, Nitra (Czechoslovakia). Katedra Slachtenia a Obrany Rastlin

    1977-01-01

    The effects were observed of acute gamma irradiation on dry seeds of Triticum aestivum var. erythrospermum (Koern.) Mansf., variety Kosutska, and Triticum monococcum (L.). Gamma radiation doses ranged from 0.258 C.kg/sup -1/ (1 kR) to 5.160 C.kg/sup -1/ (20 kR). Plant samples from pot experiments were analyzed as to the lengths of the first three leaves, production of dry matter and chloroplast ultrastructure. The studied quantitative indices, their variability and correlations are substantially dependent on the genotype of the tested species and on the applied radiation dose. Gamma radiation caused vesiculation and increase of chloroplasts, an increase in the grain number, a decline in the number of disks, a reduction of stroma thylakoids and a grouping of grana in the chloroplasts. The frequency of these changes is significantly influenced by the genotype of the tested species and by the applied radiation dose level.

  16. Expression and Chloroplast Targeting of Cholesterol Oxidase in Transgenic Tobacco Plants

    Science.gov (United States)

    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

  17. Development of 12 Chloroplast Microsatellite Markers in Vigna unguiculata (Fabaceae and Amplification in Phaseolus vulgaris

    Directory of Open Access Journals (Sweden)

    Lei Pan

    2014-03-01

    Full Text Available Premise of the study: Vigna unguiculata is an economically important legume, and the complexity of its variability and evolution needs to be further understood. Based on publicly available databases, we developed chloroplast microsatellite primers to investigate genetic diversity within V. unguiculata and its related species Phaseolus vulgaris. Methods and Results: Twelve polymorphic chloroplast microsatellite markers were developed and characterized in 62 V. unguiculata individuals. The number of alleles per locus varied between two and four, the unbiased haploid diversity per locus ranged from 0.123 to 0.497, and the polymorphism information content varied from 0.114 to 0.369. In cross-species amplifications, nine of these markers showed polymorphism in 29 P. vulgaris individuals. Conclusions: The newly developed chloroplast microsatellite markers exhibit variation in V. unguiculata as well as their transferability in P. vulgaris. These markers can be used to investigate genetic diversity and evolution in V. unguiculata and P. vulgaris.

  18. The complete chloroplast genome of a medicinal plant Epimedium koreanum Nakai (Berberidaceae).

    Science.gov (United States)

    Lee, Jung-Hoon; Kim, Kyunghee; Kim, Na-Rae; Lee, Sang-Choon; Yang, Tae-Jin; Kim, Young-Dong

    2016-11-01

    Epimedium koreanum is a perennial medicinal plant distributed in Eastern Asia. The complete chloroplast genome sequences of E. koreanum was obtained by de novo assembly using whole genome next-generation sequences. The chloroplast genome of E. koreanum was 157 218 bp in length and separated into four distinct regions such as large single copy region (89 600 bp), small single copy region (17 222 bp) and a pair of inverted repeat regions (25 198 bp). The genome contained a total of 112 genes including 78 protein-coding genes, 30 tRNA genes, and 4 rRNA genes. Phylogenetic analysis with the reported chloroplast genomes revealed that E. koreanum is most closely related to Berberis bealei, a traditional medicinal plant in the Berberidaceae family.

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

    Science.gov (United States)

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

    2012-11-01

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

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

    Science.gov (United States)

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

    2016-02-01

    Plastid DNA engineering is a well-established research area of plant biotechnology, and plastid transgenes often give high expression levels. However, it is still almost impossible to predict the accumulation rate of heterologous protein in transplastomic plants, and there are many cases of unsuccessful transgene expression. Chloroplasts regulate their proteome at the post-transcriptional level, mainly through translation control. One of the mechanisms to modulate the translation has been described in plant chloroplasts for the chloroplast-encoded subunits of multiprotein complexes, and the autoregulation of the translation initiation of these subunits depends on the availability of their assembly partners [control by epistasy of synthesis (CES)]. In Chlamydomonas reinhardtii, autoregulation of endogenous proteins recruited in the assembly of functional complexes has also been reported. In this study, we revealed a self-regulation mechanism triggered by the accumulation of a soluble recombinant protein, phaseolin, in the stroma of chloroplast-transformed tobacco plants. Immunoblotting experiments showed that phaseolin could avoid this self-regulation mechanism when targeted to the thylakoids in transplastomic plants. To inhibit the thylakoid-targeted phaseolin translation as well, this protein was expressed in the presence of a nuclear version of the phaseolin gene with a transit peptide. Pulse-chase and polysome analysis revealed that phaseolin mRNA translation on plastid ribosomes was repressed due to the accumulation in the stroma of the same soluble polypeptide imported from the cytosol. We suggest that translation autoregulation in chloroplast is not limited to heteromeric protein subunits but also involves at least some of the foreign soluble recombinant proteins, leading to the inhibition of plastome-encoded transgene expression in chloroplast. © 2015 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

  1. Towards an understanding of wheat chloroplasts: a methodical investigation of thylakoid proteome.

    Science.gov (United States)

    Kamal, Abu Hena Mostafa; Cho, Kun; Komatsu, Setsuko; Uozumi, Nobuyuki; Choi, Jong-Soon; Woo, Sun Hee

    2012-05-01

    We utilized Percoll density gradient centrifugation to isolate and fractionate chloroplasts of Korean winter wheat cultivar cv. Kumgang (Triticum aestivum L.). The resulting protein fractions were separated by one dimensional polyacrylamide gel electrophoresis (1D-PAGE) coupled with LTQ-FTICR mass spectrometry. This enabled us to detect and identify 767 unique proteins. Our findings represent the most comprehensive exploration of a proteome to date. Based on annotation information from the UniProtKB/Swiss-Prot database and our analyses via WoLF PSORT and PSORT, these proteins are localized in the chloroplast (607 proteins), chloroplast stroma (145), thylakoid membrane (342), lumens (163), and integral membranes (166). In all, 67% were confirmed as chloroplast thylakoid proteins. Although nearly complete protein coverage (89% proteins) has been accomplished for the key chloroplast pathways in wheat, such as for photosynthesis, many other proteins are involved in regulating carbon metabolism. The identified proteins were assigned to 103 functional categories according to a classification system developed by the iProClass database and provided through Protein Information Resources. Those functions include electron transport, energy, cellular organization and biogenesis, transport, stress responses, and other metabolic processes. Whereas most of these proteins are associated with known complexes and metabolic pathways, about 13% of the proteins have unknown functions. The chloroplast proteome contains many proteins that are localized to the thylakoids but as yet have no known function. We propose that some of these familiar proteins participate in the photosynthetic pathway. Thus, our new and comprehensive protein profile may provide clues for better understanding that photosynthetic process in wheat.

  2. Structure of cells chloroplasts and mitochondria of cotton leaves following gamma irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Arslanova, S V [AN Uzbekskoj SSR, Tashkent. Inst. Ehksperimental' noj Biologii Rastenij

    1975-01-01

    The article investigates the structural changes in the plastides and mitochondria of cotton leaf cells after irradiation. Cotton seeds that had been moistened for 24 hours were irradiated by a gamma source with a dose of 10 kR (intensity: 19 R/s.). For the study of the plastides and mitochondria of the leaf cells samples were taken in the cotyledonous leaf and flowering phases of the cotton. The cells of the cotton leaf mesophillum in the standard consists of chloroplast with developed interior structures. Study of the ultrastructure of the cells of the mesophilic tissue of the cotyledonous leaf in irradiated cotton plants showed that the chloroplastide membranes are not damaged. A change in the form of the chloroplasts, an accumulation of starch and plastic substances in the chloroplasts, and a reduction in the number of inter-grain bonds were noted. It was discovered that gamma irradiation produces an excessive build-up of starch in the chloroplasts. The mitochondria are often located close to the plastides. The optical density is typical of the matrix of the mitochondria in non-irradiated plants. After cotton seeds that have sprouted are irradiated with a dose of 10 kR in the cotyledonous leaf phase, part of the mitochondria swells. The matrix becomes more transparent, and the number of chrysts decreases. Part of the mitochondria remains intact. The optical density and internal membranes of the mitochondria remain the same as in the control group. The disturbances of the chloroplast and the mitochondria are also observed in the budding and flowering phases (under conditions of a natural day). It was noted that a shortened day facilitated to some extent a normalization of metabolism, and this produced in turn a normal development of the chloroplasts, leaf mitochondria and ATF generation, which reduces the final biological effect of the radiation.

  3. Complete chloroplast DNA sequence from a Korean endemic genus, Megaleranthis saniculifolia, and its evolutionary implications.

    Science.gov (United States)

    Kim, Young-Kyu; Park, Chong-wook; Kim, Ki-Joong

    2009-03-31

    The chloroplast DNA sequences of Megaleranthis saniculifolia, an endemic and monotypic endangered plant species, were completed in this study (GenBank FJ597983). The genome is 159,924 bp in length. It harbors a pair of IR regions consisting of 26,608 bp each. The lengths of the LSC and SSC regions are 88,326 bp and 18,382 bp, respectively. The structural organizations, gene and intron contents, gene orders, AT contents, codon usages, and transcription units of the Megaleranthis chloroplast genome are similar to those of typical land plant cp DNAs. However, the detailed features of Megaleranthis chloroplast genomes are substantially different from that of Ranunculus, which belongs to the same family, the Ranunculaceae. First, the Megaleranthis cp DNA was 4,797 bp longer than that of Ranunculus due to an expanded IR region into the SSC region and duplicated sequence elements in several spacer regions of the Megaleranthis cp genome. Second, the chloroplast genomes of Megaleranthis and Ranunculus evidence 5.6% sequence divergence in the coding regions, 8.9% sequence divergence in the intron regions, and 18.7% sequence divergence in the intergenic spacer regions, respectively. In both the coding and noncoding regions, average nucleotide substitution rates differed markedly, depending on the genome position. Our data strongly implicate the positional effects of the evolutionary modes of chloroplast genes. The genes evidencing higher levels of base substitutions also have higher incidences of indel mutations and low Ka/Ks ratios. A total of 54 simple sequence repeat loci were identified from the Megaleranthis cp genome. The existence of rich cp SSR loci in the Megaleranthis cp genome provides a rare opportunity to study the population genetic structures of this endangered species. Our phylogenetic trees based on the two independent markers, the nuclear ITS and chloroplast matK sequences, strongly support the inclusion of the Megaleranthis to the Trollius. Therefore, our

  4. Identification and Characterization of a Chloroplast-Targeted Obg GTPase in Dendrobium officinale.

    Science.gov (United States)

    Chen, Ji; Deng, Feng; Deng, Mengsheng; Han, Jincheng; Chen, Jianbin; Wang, Li; Yan, Shen; Tong, Kai; Liu, Fan; Tian, Mengliang

    2016-12-01

    Bacterial homologous chloroplast-targeted Obg GTPases (ObgCs) belong to the plant-typical Obg group, which is involved in diverse physiological processes during chloroplast development. However, the evolutionarily conserved function of ObgC in plants remains elusive and requires further investigation. In this study, we identified DoObgC from an epiphytic plant Dendrobium officinale and demonstrated the characteristics of DoObgC. Sequence analysis indicated that DoObgC is highly conserved with other plant ObgCs, which contain the chloroplast transit peptide (cTP), Obg fold, G domain, and OCT regions. The C terminus of DoObgC lacking the chloroplast-targeting cTP region, DoObgC Δ1-160 , showed strong similarity to ObgE and other bacterial Obgs. Overexpression of DoObgC Δ1-160 in Escherichia coli caused slow cell growth and an increased number of elongated cells. This phenotype was consistent with the phenotype of cells overexpressing ObgE. Furthermore, the expression of recombinant DoObgC Δ1-160 enhanced the cell persistence of E. coli to streptomycin. Results of transient expression assays revealed that DoObgC was localized to chloroplasts. Moreover, we demonstrated that DoObgC could rescue the embryotic lethal phenotype of the Arabidopsis obgc-t mutant, suggesting that DoObgC is a functional homolog to Arabidopsis AtObgC in D. officinale. Gene expression profiles showed that DoObgC was expressed in leaf-specific and light-dependent patterns and that DoObgC responded to wounding treatments. Our previous and present studies reveal that ObgC has an evolutionarily conserved role in ribosome biogenesis to adapt chloroplast development to the environment.

  5. Photosynthesis by isolated chloroplasts. VIII. Photosynthetic phosphorylation and the generation of assimilatory power

    Energy Technology Data Exchange (ETDEWEB)

    Arnon, D I; Whatley, F R; Allen, M B

    1959-01-01

    Photochemical ATP formation by isolated chloroplasts was coupled with a reduction of ferricyanide or TPN. Esterification of two moles of orthophosphate was coupled with the formation of two moles of TPNH/sub 2/ and the evolution of one mole of oxygen. The addition of catalytic amounts of FMN, vitamin K or phenazine methosulfate to the TPN phosphorylating system suppressed TPNH/sub 2/ accumulation as well as oxygen evolution and greatly increased the light-dependent ATP formation. A revised general scheme is presented for photosynthesis by isolated chloroplasts. 35 references, 9 figures, 4 tables.

  6. The DCL gene of tomato is required for chloroplast development and palisade cell morphogenesis in leaves.

    OpenAIRE

    Keddie, J S; Carroll, B; Jones, J D; Gruissem, W

    1996-01-01

    The defective chloroplasts and leaves-mutable (dcl-m) mutation of tomato was identified in a Ds mutagenesis screen. This unstable mutation affects both chloroplast development and palisade cell morphogenesis in leaves. Mutant plants are clonally variegated as a result of somatic excision of Ds and have albino leaves with green sectors. Leaf midribs and stems are light green with sectors of dark green tissue but fruit and petals are wild-type in appearance. Within dark green sectors of dcl-m l...

  7. Noncoding RNA mediated traffic of foreign mRNA into chloroplasts reveals a novel signaling mechanism in plants.

    Directory of Open Access Journals (Sweden)

    Gustavo Gómez

    Full Text Available Communication between chloroplasts and the nucleus is one of the milestones of the evolution of plants on earth. Proteins encoded by ancestral chloroplast-endogenous genes were transferred to the nucleus during the endosymbiotic evolution and originated this communication, which is mainly dependent on specific transit-peptides. However, the identification of nuclear-encoded proteins targeted to the chloroplast lacking these canonical signals suggests the existence of an alternative cellular pathway tuning this metabolic crosstalk. Non-coding RNAS (NcRNAs are increasingly recognized as regulators of gene expression as they play roles previously believed to correspond to proteins. Avsunviroidae family viroids are the only noncoding functional RNAs that have been reported to traffic inside the chloroplasts. Elucidating mechanisms used by these pathogens to enter this organelle will unearth novel transport pathways in plant cells. Here we show that a viroid-derived NcRNA acting as a 5'UTR-end mediates the functional import of Green Fluorescent Protein (GFP mRNA into chloroplast. This claim is supported by the observation at confocal microscopy of a selective accumulation of GFP in the chloroplast of the leaves expressing the chimeric vd-5'UTR/GFP and by the detection of the GFP mRNA in chloroplasts isolated from cells expressing this construct. These results support the existence of an alternative signaling mechanism in plants between the host cell and chloroplasts, where an ncRNA functions as a key regulatory molecule to control the accumulation of nuclear-encoded proteins in this organelle. In addition, our findings provide a conceptual framework to develop new biotechnological tools in systems using plant chloroplast as bioreactors. Finally, viroids of the family Avsunviroidae have probably evolved to subvert this signaling mechanism to regulate their differential traffic into the chloroplast of infected cells.

  8. A model for tetrapyrrole synthesis as the primary mechanism for plastid-to-nucleus signaling during chloroplast biogenesis

    Directory of Open Access Journals (Sweden)

    Matthew J. Terry

    2013-02-01

    Full Text Available Chloroplast biogenesis involves the co-ordinated expression of the chloroplast and nuclear genomes, requiring information to be sent from the developing chloroplasts to the nucleus. This is achieved through retrograde signaling pathways and can be demonstrated experimentally using the photobleaching herbicide, Norflurazon, which results in chloroplast damage and the reduced expression of many photosynthesis-related, nuclear genes in seedlings. Genetic analysis of this pathway points to a major role for tetrapyrrole synthesis in retrograde signaling, as well as a strong interaction with light-signaling pathways. Currently, the best model to explain the genetic data is that a specific heme pool generated by flux through ferrochelatase-1 functions as a positive signal to promote the expression of genes required for chloroplast development. We propose that this heme-related signal is the primary positive signal during chloroplast biogenesis, and that treatments and mutations affecting chloroplast transcription, RNA editing, translation, or protein import all impact on the synthesis and/or processing of this signal. A positive signal is consistent with the need to provide information on chloroplast status at all times. We further propose that GUN1 normally serves to restrict the production of the heme signal. In addition to a positive signal re-enforcing chloroplast development under normal conditions, aberrant chloroplast development may produce a negative signal due to accumulation of unbound chlorophyll biosynthesis intermediates, such as Mg-porphyrins. Under these conditions a rapid shut-down of tetrapyrrole synthesis is required. We propose that accumulation of these intermediates results in a rapid light-dependent inhibition of nuclear gene expression that is most likely mediated via singlet oxygen generated by photo-excitation of Mg-porphyrins. Thus, the tetrapyrrole pathway may provide both positive and inhibitory signals to control

  9. De Novo Assembly of Complete Chloroplast Genomes from Non-model Species Based on a K-mer Frequency-Based Selection of Chloroplast Reads from Total DNA Sequences

    Directory of Open Access Journals (Sweden)

    Shairul Izan

    2017-08-01

    Full Text Available Whole Genome Shotgun (WGS sequences of plant species often contain an abundance of reads that are derived from the chloroplast genome. Up to now these reads have generally been identified and assembled into chloroplast genomes based on homology to chloroplasts from related species. This re-sequencing approach may select against structural differences between the genomes especially in non-model species for which no close relatives have been sequenced before. The alternative approach is to de novo assemble the chloroplast genome from total genomic DNA sequences. In this study, we used k-mer frequency tables to identify and extract the chloroplast reads from the WGS reads and assemble these using a highly integrated and automated custom pipeline. Our strategy includes steps aimed at optimizing assemblies and filling gaps which are left due to coverage variation in the WGS dataset. We have successfully de novo assembled three complete chloroplast genomes from plant species with a range of nuclear genome sizes to demonstrate the universality of our approach: Solanum lycopersicum (0.9 Gb, Aegilops tauschii (4 Gb and Paphiopedilum henryanum (25 Gb. We also highlight the need to optimize the choice of k and the amount of data used. This new and cost-effective method for de novo short read assembly will facilitate the study of complete chloroplast genomes with more accurate analyses and inferences, especially in non-model plant genomes.

  10. Electron cryomicroscopy of two-dimensional crystals of the H+-ATPase from chloroplasts

    NARCIS (Netherlands)

    Böttcher, Bettina; Gräber, Peter; Boekema, Egbert J.; Lücken, Uwe

    1995-01-01

    The H+-ATPase from spinach chloroplasts was isolated and purified. Two-dimensional crystals were obtained from the protein/lipid/detergent micelles by treatment with phospholipase and simultaneous removal of detergent and fatty acids by Biobeads. The resulting two-dimensionally ordered arrays were

  11. ELECTRON CRYOMICROSCOPY OF 2-DIMENSIONAL CRYSTALS OF THE H+-ATPASE FROM CHLOROPLASTS

    NARCIS (Netherlands)

    BOTTCHER, B; GRABER, P; BOEKEMA, EJ; LUCKEN, U

    1995-01-01

    The H+-ATPase from spinach chloroplasts was isolated and purified, Two-dimensional crystals were obtained from the protein/lipid/detergent micelles by treatment with phospholipase and simultaneous removal of detergent and fatty acids by Biobeads. The resulting two-dimensionally ordered arrays were

  12. Electrochromic effects in relation to energy transduction and energy coupling in chloroplast membranes

    NARCIS (Netherlands)

    Peters, R.L.A.

    1986-01-01

    A study was made on the kinetics of the flash-induced P515 electrochromic bandshift signal in spinach leaves and isolated chloroplasts. It was found that part of the signal (i.e. the slow component, also called reaction 2), normally present in dark-adapted membranes is absent from the signal under

  13. Diversity of chloroplast genome among local clones of cocoa (Theobroma cacao, L.) from Central Sulawesi

    Science.gov (United States)

    Suwastika, I. Nengah; Pakawaru, Nurul Aisyah; Rifka, Rahmansyah, Muslimin, Ishizaki, Yoko; Cruz, André Freire; Basri, Zainuddin; Shiina, Takashi

    2017-02-01

    Chloroplast genomes typically range in size from 120 to 170 kilo base pairs (kb), which relatively conserved among plant species. Recent evaluation on several species, certain unique regions showed high variability which can be utilized in the phylogenetic analysis. Many fragments of coding regions, introns, and intergenic spacers, such as atpB-rbcL, ndhF, rbcL, rpl16, trnH-psbA, trnL-F, trnS-G, etc., have been used for phylogenetic reconstructions at various taxonomic levels. Based on that status, we would like to analysis the diversity of chloroplast genome within species of local cacao (Theobroma cacao L.) from Central Sulawesi. Our recent data showed, there were more than 20 clones from local farming in Central Sulawesi, and it can be detected based on phenotypic and nuclear-genome-based characterization (RAPD- Random Amplified Polymorphic DNA and SSR- Simple Sequences Repeat) markers. In developing DNA marker for this local cacao, here we also included analysis based on the variation of chloroplast genome. At least several regions such as rpl32-TurnL, it can be considered as chloroplast markers on our local clone of cocoa. Furthermore, we could develop phylogenetic analysis in between clones of cocoa.

  14. A set of 100 chloroplast DNA primer pairs to study population genetics and phylogeny in monocotylenons

    DEFF Research Database (Denmark)

    Scarcelli, Nora; Bernaud, Adeline; Eiserhardt, Wolf L.

    2011-01-01

    Chloroplast DNA sequences are of great interest for population genetics and phylogenetic studies. However, only a small set of markers are commonly used. Most of them have been designed for amplification in a large range of Angiosperms and are located in the Large Single Copy (LSC). Here we...... anticipate that it will also be useful for phylogeny and bar-coding studies....

  15. Sequencing of chloroplast genome using whole cellular DNA and Solexa sequencing technology

    Directory of Open Access Journals (Sweden)

    Jian eWu

    2012-11-01

    Full Text Available Sequencing of the chloroplast genome using traditional sequencing methods has been difficult because of its size (>120 kb and the complicated procedures required to prepare templates. To explore the feasibility of sequencing the chloroplast genome using DNA extracted from whole cells and Solexa sequencing technology, we sequenced whole cellular DNA isolated from leaves of three Brassica rapa accessions with one lane per accession. In total, 246 Mb, 362Mb, 361 Mb sequence data were generated for the three accessions Chiifu-401-42, Z16 and FT, respectively. Microreads were assembled by reference-guided assembly using the cpDNA sequences of B. rapa, Arabidopsis thaliana, and Nicotiana tabacum. We achieved coverage of more than 99.96% of the cp genome in the three tested accessions using the B. rapa sequence as the reference. When A. thaliana or N. tabacum sequences were used as references, 99.7–99.8% or 95.5–99.7% of the B. rapa chloroplast genome was covered, respectively. These results demonstrated that sequencing of whole cellular DNA isolated from young leaves using the Illumina Genome Analyzer is an efficient method for high-throughput sequencing of chloroplast genome.

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

    Science.gov (United States)

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

    2008-01-01

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

  17. Regulation of photosynthetic electron flow in isolated chloroplasts by bicarbonate, formate and herbicides

    NARCIS (Netherlands)

    Snel, J.F.H.

    1985-01-01

    This thesis describes some efforts that were made to gain a better understanding of the processes involved in the regulation of photosynthetic electron flow by bicarbonate, formate and herbicides in chloroplasts. In the past decade a large amount of research has been devoted to get insight into the

  18. Increasing phylogenetic resolution at low taxonomic levels using massively parallel sequencing of chloroplast genomes

    Science.gov (United States)

    Matthew Parks; Richard Cronn; Aaron Liston

    2009-01-01

    We reconstruct the infrageneric phylogeny of Pinus from 37 nearly-complete chloroplast genomes (average 109 kilobases each of an approximately 120 kilobase genome) generated using multiplexed massively parallel sequencing. We found that 30/33 ingroup nodes resolved wlth > 95-percent bootstrap support; this is a substantial improvement relative...

  19. A tiling microarray for global analysis of chloroplast genome expression in cucumber and other plants

    Directory of Open Access Journals (Sweden)

    Pląder Wojciech

    2011-09-01

    Full Text Available Abstract Plastids are small organelles equipped with their own genomes (plastomes. Although these organelles are involved in numerous plant metabolic pathways, current knowledge about the transcriptional activity of plastomes is limited. To solve this problem, we constructed a plastid tiling microarray (PlasTi-microarray consisting of 1629 oligonucleotide probes. The oligonucleotides were designed based on the cucumber chloroplast genomic sequence and targeted both strands of the plastome in a non-contiguous arrangement. Up to 4 specific probes were designed for each gene/exon, and the intergenic regions were covered regularly, with 70-nt intervals. We also developed a protocol for direct chemical labeling and hybridization of as little as 2 micrograms of chloroplast RNA. We used this protocol for profiling the expression of the cucumber chloroplast plastome on the PlasTi-microarray. Owing to the high sequence similarity of plant plastomes, the newly constructed microarray can be used to study plants other than cucumber. Comparative hybridization of chloroplast transcriptomes from cucumber, Arabidopsis, tomato and spinach showed that the PlasTi-microarray is highly versatile.

  20. BEL1-LIKE HOMEODOMAIN 11 regulated chloroplast development and chlorophyll synthesis in tomato fruit

    Science.gov (United States)

    Chloroplast development and chlorophyll content and metabolism in unripe tomato contribute to the growth and development of the fruit, and also the ripe fruit quality, but the mechanism is poorly understood. In this work, seven homeobox-containing transcription factors (TFs) with specific ripening-a...

  1. Chloroplast evolution in the Pinus montezumae complex: a coalescent approach to hybridization.

    Science.gov (United States)

    Matos, J A; Schaal, B A

    2000-08-01

    This study addresses the evolutionary history of the chloroplast genomes of two closely related pine species, Pinus hartwegii Lindl. and P. montezumae Lamb (subsect. Ponderosae) using coalescent theory and some of the statistical tools that have been developed from it during the past two decades. Pinus hartwegii and P. montezumae are closely related species in the P. montezumae complex (subsect. Ponderosae) of Mexico and Central America. Pinus hartwegii is a high elevation species, whereas P. montezumae occurs at lower elevations. The two species occur on many of the same mountains throughout Mexico. A total of 350 individuals of P. hartwegii and P. montezumae were collected from Nevado de Colima (Jalisco), Cerro Potosí (Nuevo León), Iztaccihuatl/Popocatepetl (México), and Nevado de Toluca (México). The chloroplast genome of P. hartwegii and P. montezumae was mapped using eight restriction enzymes. Fifty-one different haplotypes were characterized; 38 of 160 restriction sites were polymorphic. Clades of most parsimoniously related chloroplast haplotypes are geographically localized and do not overlap in distribution, and the geographically localized clades of haplotypes include both P. hartwegii and P. montezumae. Some haplotypes in the clades occur in only one of the two species, whereas other haplotypes occur in both species. These data strongly suggest ancient and/or ongoing hybridization between P. hartwegii and P. montezumae and a shared chloroplast genome history within geographic regions of Mexico.

  2. Rangewide Genetic Variation in Coast Redwood Populations at a Chloroplast Microsatellite Locus

    Science.gov (United States)

    Chris Brinegar

    2012-01-01

    Old growth and second growth populations of coast redwood (Sequoia sempervirens) were sampled at 10 locations throughout its range and analyzed at a highly variable chloroplast microsatellite locus. Very low FST values indicated that there was no significant genetic differentiation between adjacent old growth and second growth populations at each location. Genetic...

  3. Inner structure of intact chloroplasts observed by a low temperature laser scanning microscope

    Czech Academy of Sciences Publication Activity Database

    Vácha, František; Vácha, M.; Bumba, L.; Hashizume, K.; Tani, T.

    2000-01-01

    Roč. 38, - (2000), s. 493-496 ISSN 0300-3604 R&D Projects: GA MŠk ME 156; GA MŠk VS96085 Keywords : chloroplasts * physiology * scanning microscopy Subject RIV: EE - Microbiology, Virology Impact factor: 0.482, year: 2000

  4. Photosynthesis by isolated chloroplasts. IV. General concept and comparison of three photochemical reactions

    Energy Technology Data Exchange (ETDEWEB)

    Arnon, D I; Allen, M B; Whatley, F R

    1956-01-01

    Procedures are described for the preparation of chloroplasts capable of carrying out three photochemical reactions, each representing an increasingly complex phase of photosynthesis: photolysis of water (Hill reaction), esterification of inorganic phosphate into adenosine triphosphate (photosynthetic phosphorylation) and the reduction of carbon dioxide to the level of carbohydrates with a simultaneous evolution of oxygen. The three photochemical reactions were separable by variations in the technique for preparation of chloroplasts and by differential inhibition by several reagents. Inhibition of a more complex phase of photosynthesis does not affect the simpler one which precedes it and, conversely, the inhibition of a simpler phase of photosynthesis is paralleled by an inhibition of the more complex phase which follows. Reversible inhibition of CO/sub 2/ fixation and photosynthetic phosphorylation, but not of photolysis, by sulfhydryl group inhibitors suggests that sulfhydryl compounds (enzymes, cofactors, or both) are involved in phosphorylation and CO/sub 2/ fixation, but not in the primary conversion of light into chemical energy as measured by the Hill reaction. Evidence is presented in support of the conclusion that the synthesis of ATP by green cells occurs at two distinct sites: anaerobically in chloroplasts by photosynthetic phosphorylation, and acrobically in smaller cytoplasmic particles, presumably mitochondria, by oxidative phosphorylation independent of light. A general scheme of photosynthesis by chloroplasts, consistent with these findings, is presented. 44 references, 8 figures, 4 tables.

  5. Differences in thermal acclimation of chloroplast functioning in two ecotypes of Valonia utricularis (Chlorophyta)

    NARCIS (Netherlands)

    Eggert, A.; van Hasselt, P.R; Breeman, Arno

    Chloroplast functioning in two temperature ecotypes of the tropical to warm-temperate green macrophyte Valonia ultricularis was monitored by measuring chlorophyll a fluorescence parameters. One ecotype from the Mediterranean Sea is, with respect to growth and survival, more cold-adapted and

  6. Complete Chloroplast Genomes of Papaver rhoeas and Papaver orientale: Molecular Structures, Comparative Analysis, and Phylogenetic Analysis

    Directory of Open Access Journals (Sweden)

    Jianguo Zhou

    2018-02-01

    Full Text Available Papaver rhoeas L. and P. orientale L., which belong to the family Papaveraceae, are used as ornamental and medicinal plants. The chloroplast genome has been used for molecular markers, evolutionary biology, and barcoding identification. In this study, the complete chloroplast genome sequences of P. rhoeas and P. orientale are reported. Results show that the complete chloroplast genomes of P. rhoeas and P. orientale have typical quadripartite structures, which are comprised of circular 152,905 and 152,799-bp-long molecules, respectively. A total of 130 genes were identified in each genome, including 85 protein-coding genes, 37 tRNA genes, and 8 rRNA genes. Sequence divergence analysis of four species from Papaveraceae indicated that the most divergent regions are found in the non-coding spacers with minimal differences among three Papaver species. These differences include the ycf1 gene and intergenic regions, such as rpoB-trnC, trnD-trnT, petA-psbJ, psbE-petL, and ccsA-ndhD. These regions are hypervariable regions, which can be used as specific DNA barcodes. This finding suggested that the chloroplast genome could be used as a powerful tool to resolve the phylogenetic positions and relationships of Papaveraceae. These results offer valuable information for future research in the identification of Papaver species and will benefit further investigations of these species.

  7. The ultrastructure of chloroplasts in variegata irregulare mutants of garden petunias (Petunia hybrida hort. superbissima

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    Stanisław Muszyński

    2015-01-01

    Full Text Available The ultrastructure of mutated chloroplasts in tetraploid garden petunias (Petunia hybrida hort. superbissima was analyzed by electron microscopy. The formation of grana structure is inhibited after secondary thylacoids start forming. Rapid dezintegration of the structure is observed. It is suggested that a substance responsible for photostabilization of grana structure is lacking.

  8. Chloroplast DNA variation of oaks in western Central Europe and genetic consequences of human influences

    NARCIS (Netherlands)

    König, A.O.; Ziegenhagen, B.; Dam, van B.C.; Csaikl, U.M.; Coart, E.; Degen, B.; Burg, K.; Vries, de S.M.G.; Petit, R.J.

    2002-01-01

    Oak chloroplast DNA (cpDNA) variation was studied in a grid-based inventory in western Central Europe, including Belgium, The Netherlands, Luxembourg, Germany, the Czech Republic, and the northern parts of Upper and Lower Austria. A total of 2155 trees representing 426 populations of Quercus robur

  9. Insights into phylogeny, sex function and age of Fragaria based on whole chloroplast genome sequencing

    Science.gov (United States)

    Wambui Njunguna; Aaron Liston; Richard Cronn; Tia-Lynn Ashman; Nahla Bassil

    2013-01-01

    The cultivated strawberry is one of the youngest domesticated plants, developed in France in the 1700s from chance hybridization between two western hemisphere octoploid species. However, little is known about the evolution of the species that gave rise to this important fruit crop. Phylogenetic analysis of chloroplast genome sequences of 21 Fragaria...

  10. The complete chloroplast DNA sequence of the green alga Oltmannsiellopsis viridis reveals a distinctive quadripartite architecture in the chloroplast genome of early diverging ulvophytes

    Directory of Open Access Journals (Sweden)

    Lemieux Claude

    2006-02-01

    Full Text Available Abstract Background The phylum Chlorophyta contains the majority of the green algae and is divided into four classes. The basal position of the Prasinophyceae has been well documented, but the divergence order of the Ulvophyceae, Trebouxiophyceae and Chlorophyceae is currently debated. The four complete chloroplast DNA (cpDNA sequences presently available for representatives of these classes have revealed extensive variability in overall structure, gene content, intron composition and gene order. The chloroplast genome of Pseudendoclonium (Ulvophyceae, in particular, is characterized by an atypical quadripartite architecture that deviates from the ancestral type by a large inverted repeat (IR featuring an inverted rRNA operon and a small single-copy (SSC region containing 14 genes normally found in the large single-copy (LSC region. To gain insights into the nature of the events that led to the reorganization of the chloroplast genome in the Ulvophyceae, we have determined the complete cpDNA sequence of Oltmannsiellopsis viridis, a representative of a distinct, early diverging lineage. Results The 151,933 bp IR-containing genome of Oltmannsiellopsis differs considerably from Pseudendoclonium and other chlorophyte cpDNAs in intron content and gene order, but shares close similarities with its ulvophyte homologue at the levels of quadripartite architecture, gene content and gene density. Oltmannsiellopsis cpDNA encodes 105 genes, contains five group I introns, and features many short dispersed repeats. As in Pseudendoclonium cpDNA, the rRNA genes in the IR are transcribed toward the single copy region featuring the genes typically found in the ancestral LSC region, and the opposite single copy region harbours genes characteristic of both the ancestral SSC and LSC regions. The 52 genes that were transferred from the ancestral LSC to SSC region include 12 of those observed in Pseudendoclonium cpDNA. Surprisingly, the overall gene organization of

  11. Photosynthesis-dependent H2O2 transfer from chloroplasts to nuclei provides a high-light signalling mechanism.

    Science.gov (United States)

    Exposito-Rodriguez, Marino; Laissue, Pierre Philippe; Yvon-Durocher, Gabriel; Smirnoff, Nicholas; Mullineaux, Philip M

    2017-06-29

    Chloroplasts communicate information by signalling to nuclei during acclimation to fluctuating light. Several potential operating signals originating from chloroplasts have been proposed, but none have been shown to move to nuclei to modulate gene expression. One proposed signal is hydrogen peroxide (H 2 O 2 ) produced by chloroplasts in a light-dependent manner. Using HyPer2, a genetically encoded fluorescent H 2 O 2 sensor, we show that in photosynthetic Nicotiana benthamiana epidermal cells, exposure to high light increases H 2 O 2 production in chloroplast stroma, cytosol and nuclei. Critically, over-expression of stromal ascorbate peroxidase (H 2 O 2 scavenger) or treatment with DCMU (photosynthesis inhibitor) attenuates nuclear H 2 O 2 accumulation and high light-responsive gene expression. Cytosolic ascorbate peroxidase over-expression has little effect on nuclear H 2 O 2 accumulation and high light-responsive gene expression. This is because the H 2 O 2 derives from a sub-population of chloroplasts closely associated with nuclei. Therefore, direct H 2 O 2 transfer from chloroplasts to nuclei, avoiding the cytosol, enables photosynthetic control over gene expression.Multiple plastid-derived signals have been proposed but not shown to move to the nucleus to promote plant acclimation to fluctuating light. Here the authors use a fluorescent hydrogen peroxide sensor to provide evidence that H 2 O 2 is transferred directly from chloroplasts to nuclei to control nuclear gene expression.

  12. Origins of the amphiploid species Brassica napus L. investigated by chloroplast and nuclear molecular markers

    Directory of Open Access Journals (Sweden)

    Allender Charlotte J

    2010-03-01

    Full Text Available Abstract Background The amphiploid species Brassica napus (oilseed rape, Canola is a globally important oil crop yielding food, biofuels and industrial compounds such as lubricants and surfactants. Identification of the likely ancestors of each of the two genomes (designated A and C found in B. napus would facilitate incorporation of novel alleles from the wider Brassica genepool in oilseed rape crop genetic improvement programmes. Knowledge of the closest extant relatives of the genotypes involved in the initial formation of B. napus would also allow further investigation of the genetic factors required for the formation of a stable amphiploid and permit the more efficient creation of fully fertile re-synthesised B. napus. We have used a combination of chloroplast and nuclear genetic markers to investigate the closest extant relatives of the original maternal progenitors of B. napus. This was based on a comprehensive sampling of the relevant genepools, including 83 accessions of A genome B. rapa L. (both wild and cultivated types, 94 accessions of B. napus and 181 accessions of C genome wild and cultivated B. oleracea L. and related species. Results Three chloroplast haplotypes occurred in B. napus. The most prevalent haplotype (found in 79% of accessions was not present within the C genome accessions but was found at low frequencies in B. rapa. Chloroplast haplotypes characteristic of B. napus were found in a small number of wild and weedy B. rapa populations, and also in two accessions of cultivated B. rapa 'brocoletto'. Whilst introgression of the B. napus chloroplast type in the wild and weedy B. rapa populations has been proposed by other studies, the presence of this haplotype within the two brocoletto accessions is unexplained. Conclusions The distribution of chloroplast haplotypes eliminate any of the C genome species as being the maternal ancestor of the majority of the B. napus accessions. The presence of multiple chloroplast

  13. Transcriptome and proteomic analyses reveal multiple differences associated with chloroplast development in the spaceflight-induced wheat albino mutant mta.

    Directory of Open Access Journals (Sweden)

    Kui Shi

    Full Text Available Chloroplast development is an integral part of plant survival and growth, and occurs in parallel with chlorophyll biosynthesis. However, little is known about the mechanisms underlying chloroplast development in hexaploid wheat. Here, we obtained a spaceflight-induced wheat albino mutant mta. Chloroplast ultra-structural observation showed that chloroplasts of mta exhibit abnormal morphology and distribution compared to wild type. Photosynthetic pigments content was also significantly decreased in mta. Transcriptome and chloroplast proteome profiling of mta and wild type were done to identify differentially expressed genes (DEGs and proteins (DEPs, respectively. In total 4,588 DEGs including 1,980 up- and 2,608 down-regulated, and 48 chloroplast DEPs including 15 up- and 33 down-regulated were identified in mta. Classification of DEGs revealed that most were involved in chloroplast development, chlorophyll biosynthesis, or photosynthesis. Besides, transcription factors such as PIF3, GLK and MYB which might participate in those pathways were also identified. The correlation analysis between DEGs and DEPs revealed that the transcript-to-protein in abundance was functioned into photosynthesis and chloroplast relevant groups. Real time qPCR analysis validated that the expression level of genes encoding photosynthetic proteins was significantly decreased in mta. Together, our results suggest that the molecular mechanism for albino leaf color formation in mta is a thoroughly regulated and complicated process. The combined analysis of transcriptome and proteome afford comprehensive information for further research on chloroplast development mechanism in wheat. And spaceflight provides a potential means for mutagenesis in crop breeding.

  14. Stable megadalton TOC-TIC supercomplexes as major mediators of protein import into chloroplasts.

    Science.gov (United States)

    Chen, Lih-Jen; Li, Hsou-Min

    2017-10-01

    Preproteins are believed to be imported into chloroplasts through membrane contact sites where the translocon complexes of the outer (TOC) and inner (TIC) envelope membranes are assembled together. However, a single TOC-TIC supercomplex containing preproteins undergoing active import has not yet been directly observed. We optimized the blue native polyacrylamide gel electrophoresis (PAGE) (BN-PAGE) system to detect and resolve megadalton (MD)-sized complexes. Using this optimized system, the outer-membrane channel Toc75 from pea chloroplasts was found in at least two complexes: the 880-kD TOC complex and a previously undetected 1-MD complex. Two-dimensional BN-PAGE immunoblots further showed that Toc75, Toc159, Toc34, Tic20, Tic56 and Tic110 were all located in the 880-kD to 1.3-MD region. During active preprotein import, preproteins were transported mostly through the 1-MD complex and a smaller amount of preproteins was also detected in a complex of 1.25 MD. Antibody-shift assays showed that the 1-MD complex is a TOC-TIC supercomplex containing at least Toc75, Toc159, Toc34 and Tic110. Results from crosslinking and import with Arabidopsis chloroplasts suggest that the 1.25-MD complex is also a supercomplex. Our data provide direct evidence supporting that chloroplast preproteins are imported through TOC-TIC supercomplexes, and also provide the first size estimation of these supercomplexes. Furthermore, unlike in mitochondria where translocon supercomplexes are only transiently assembled during preprotein import, in chloroplasts at least some of the supercomplexes are preassembled stable structures. © 2017 The Authors The Plant Journal published by John Wiley & Sons Ltd and Society for Experimental Biology.

  15. Chloroplast genome resources and molecular markers differentiate rubber dandelion species from weedy relatives.

    Science.gov (United States)

    Zhang, Yingxiao; Iaffaldano, Brian J; Zhuang, Xiaofeng; Cardina, John; Cornish, Katrina

    2017-02-02

    Rubber dandelion (Taraxacum kok-saghyz, TK) is being developed as a domestic source of natural rubber to meet increasing global demand. However, the domestication of TK is complicated by its colocation with two weedy dandelion species, Taraxacum brevicorniculatum (TB) and the common dandelion (Taraxacum officinale, TO). TB is often present as a seed contaminant within TK accessions, while TO is a pandemic weed, which may have the potential to hybridize with TK. To discriminate these species at the molecular level, and facilitate gene flow studies between the potential rubber crop, TK, and its weedy relatives, we generated genomic and marker resources for these three dandelion species. Complete chloroplast genome sequences of TK (151,338 bp), TO (151,299 bp), and TB (151,282 bp) were obtained using the Illumina GAII and MiSeq platforms. Chloroplast sequences were analyzed and annotated for all the three species. Phylogenetic analysis within Asteraceae showed that TK has a closer genetic distance to TB than to TO and Taraxacum species were most closely related to lettuce (Lactuca sativa). By sequencing multiple genotypes for each species and testing variants using gel-based methods, four chloroplast Single Nucleotide Polymorphism (SNP) variants were found to be fixed between TK and TO in large populations, and between TB and TO. Additionally, Expressed Sequence Tag (EST) resources developed for TO and TK permitted the identification of five nuclear species-specific SNP markers. The availability of chloroplast genomes of these three dandelion species, as well as chloroplast and nuclear molecular markers, will provide a powerful genetic resource for germplasm differentiation and purification, and the study of potential gene flow among Taraxacum species.

  16. The First Complete Chloroplast Genome Sequences in Actinidiaceae: Genome Structure and Comparative Analysis.

    Science.gov (United States)

    Yao, Xiaohong; Tang, Ping; Li, Zuozhou; Li, Dawei; Liu, Yifei; Huang, Hongwen

    2015-01-01

    Actinidia chinensis is an important economic plant belonging to the basal lineage of the asterids. Availability of a complete Actinidia chloroplast genome sequence is crucial to understanding phylogenetic relationships among major lineages of angiosperms and facilitates kiwifruit genetic improvement. We report here the complete nucleotide sequences of the chloroplast genomes for Actinidia chinensis and A. chinensis var deliciosa obtained through de novo assembly of Illumina paired-end reads produced by total DNA sequencing. The total genome size ranges from 155,446 to 157,557 bp, with an inverted repeat (IR) of 24,013 to 24,391 bp, a large single copy region (LSC) of 87,984 to 88,337 bp and a small single copy region (SSC) of 20,332 to 20,336 bp. The genome encodes 113 different genes, including 79 unique protein-coding genes, 30 tRNA genes and 4 ribosomal RNA genes, with 16 duplicated in the inverted repeats, and a tRNA gene (trnfM-CAU) duplicated once in the LSC region. Comparisons of IR boundaries among four asterid species showed that IR/LSC borders were extended into the 5' portion of the psbA gene and IR contraction occurred in Actinidia. The clap gene has been lost from the chloroplast genome in Actinidia, and may have been transferred to the nucleus during chloroplast evolution. Twenty-seven polymorphic simple sequence repeat (SSR) loci were identified in the Actinidia chloroplast genome. Maximum parsimony analyses of a 72-gene, 16 taxa angiosperm dataset strongly support the placement of Actinidiaceae in Ericales within the basal asterids.

  17. Phosphoinositides play differential roles in regulating phototropin1- and phototropin2-mediated chloroplast movements in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Chhavi Aggarwal

    Full Text Available 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(4,5P2]-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 Ca(2+ ((c signaling. Using the transgenic aequorin system, we show that the application of these inhibitors suppresses the blue-light-induced transient Ca(2+ ((c rise. These results demonstrate the importance of PIs in chloroplast movements, with the PI(4,5P2-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 Ca(2+ signaling during movements.

  18. Phosphoinositides play differential roles in regulating phototropin1- and phototropin2-mediated chloroplast movements in Arabidopsis.

    Science.gov (United States)

    Aggarwal, Chhavi; Labuz, 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 Ca(2+) ((c)) signaling. Using the transgenic aequorin system, we show that the application of these inhibitors suppresses the blue-light-induced transient Ca(2+) ((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 Ca(2+) signaling during movements.

  19. The Complete Chloroplast Genome Sequences of the Medicinal Plant Forsythia suspensa (Oleaceae

    Directory of Open Access Journals (Sweden)

    Wenbin Wang

    2017-10-01

    Full Text Available Forsythia suspensa is an important medicinal plant and traditionally applied for the treatment of inflammation, pyrexia, gonorrhea, diabetes, and so on. However, there is limited sequence and genomic information available for F. suspensa. Here, we produced the complete chloroplast genomes of F. suspensa using Illumina sequencing technology. F. suspensa is the first sequenced member within the genus Forsythia (Oleaceae. The gene order and organization of the chloroplast genome of F. suspensa are similar to other Oleaceae chloroplast genomes. The F. suspensa chloroplast genome is 156,404 bp in length, exhibits a conserved quadripartite structure with a large single-copy (LSC; 87,159 bp region, and a small single-copy (SSC; 17,811 bp region interspersed between inverted repeat (IRa/b; 25,717 bp regions. A total of 114 unique genes were annotated, including 80 protein-coding genes, 30 tRNA, and four rRNA. The low GC content (37.8% and codon usage bias for A- or T-ending codons may largely affect gene codon usage. Sequence analysis identified a total of 26 forward repeats, 23 palindrome repeats with lengths >30 bp (identity > 90%, and 54 simple sequence repeats (SSRs with an average rate of 0.35 SSRs/kb. We predicted 52 RNA editing sites in the chloroplast of F. suspensa, all for C-to-U transitions. IR expansion or contraction and the divergent regions were analyzed among several species including the reported F. suspensa in this study. Phylogenetic analysis based on whole-plastome revealed that F. suspensa, as a member of the Oleaceae family, diverged relatively early from Lamiales. This study will contribute to strengthening medicinal resource conservation, molecular phylogenetic, and genetic engineering research investigations of this species.

  20. The Complete Chloroplast Genome of Ye-Xing-Ba (Scrophularia dentata; Scrophulariaceae), an Alpine Tibetan Herb.

    Science.gov (United States)

    Ni, Lianghong; Zhao, Zhili; Dorje, Gaawe; Ma, Mi

    2016-01-01

    Scrophularia dentata is an important Tibetan medicinal plant and traditionally used for the treatment of exanthema and fever in Traditional Tibetan Medicine (TTM). However, there is little sequence and genomic information available for S. dentata. In this paper, we report the complete chloroplast genome sequence of S. dentata and it is the first sequenced member of the Sect. Tomiophyllum within Scrophularia (Scrophulariaceae). The gene order and organization of the chloroplast genome of S. dentata are similar to other Lamiales chloroplast genomes. The plastome is 152,553 bp in length and includes a pair of inverted repeats (IRs) of 25,523 bp that separate a large single copy (LSC) region of 84,058 bp and a small single copy (SSC) region of 17,449 bp. It has 38.0% GC content and includes 114 unique genes, of which 80 are protein-coding, 30 are transfer RNA, and 4 are ribosomal RNA. Also, it contains 21 forward repeats, 19 palindrome repeats and 41 simple sequence repeats (SSRs). The repeats and SSRs within S. dentata were compared with those of S. takesimensis and present certain discrepancies. The chloroplast genome of S. dentata was compared with other five publicly available Lamiales genomes from different families. All the coding regions and non-coding regions (introns and intergenic spacers) within the six chloroplast genomes have been extracted and analysed. Furthermore, the genome divergent hotspot regions were identified. Our studies could provide basic data for the alpine medicinal species conservation and molecular phylogenetic researches of Scrophulariaceae and Lamiales.

  1. The Complete Chloroplast Genome of Ye-Xing-Ba (Scrophularia dentata; Scrophulariaceae, an Alpine Tibetan Herb.

    Directory of Open Access Journals (Sweden)

    Lianghong Ni

    Full Text Available Scrophularia dentata is an important Tibetan medicinal plant and traditionally used for the treatment of exanthema and fever in Traditional Tibetan Medicine (TTM. However, there is little sequence and genomic information available for S. dentata. In this paper, we report the complete chloroplast genome sequence of S. dentata and it is the first sequenced member of the Sect. Tomiophyllum within Scrophularia (Scrophulariaceae. The gene order and organization of the chloroplast genome of S. dentata are similar to other Lamiales chloroplast genomes. The plastome is 152,553 bp in length and includes a pair of inverted repeats (IRs of 25,523 bp that separate a large single copy (LSC region of 84,058 bp and a small single copy (SSC region of 17,449 bp. It has 38.0% GC content and includes 114 unique genes, of which 80 are protein-coding, 30 are transfer RNA, and 4 are ribosomal RNA. Also, it contains 21 forward repeats, 19 palindrome repeats and 41 simple sequence repeats (SSRs. The repeats and SSRs within S. dentata were compared with those of S. takesimensis and present certain discrepancies. The chloroplast genome of S. dentata was compared with other five publicly available Lamiales genomes from different families. All the coding regions and non-coding regions (introns and intergenic spacers within the six chloroplast genomes have been extracted and analysed. Furthermore, the genome divergent hotspot regions were identified. Our studies could provide basic data for the alpine medicinal species conservation and molecular phylogenetic researches of Scrophulariaceae and Lamiales.

  2. High-throughput sequencing of three Lemnoideae (duckweeds chloroplast genomes from total DNA.

    Directory of Open Access Journals (Sweden)

    Wenqin Wang

    Full Text Available BACKGROUND: Chloroplast genomes provide a wealth of information for evolutionary and population genetic studies. Chloroplasts play a particularly important role in the adaption for aquatic plants because they float on water and their major surface is exposed continuously to sunlight. The subfamily of Lemnoideae represents such a collection of aquatic species that because of photosynthesis represents one of the fastest growing plant species on earth. METHODS: We sequenced the chloroplast genomes from three different genera of Lemnoideae, Spirodela polyrhiza, Wolffiella lingulata and Wolffia australiana by high-throughput DNA sequencing of genomic DNA using the SOLiD platform. Unfractionated total DNA contains high copies of plastid DNA so that sequences from the nucleus and mitochondria can easily be filtered computationally. Remaining sequence reads were assembled into contiguous sequences (contigs using SOLiD software tools. Contigs were mapped to a reference genome of Lemna minor and gaps, selected by PCR, were sequenced on the ABI3730xl platform. CONCLUSIONS: This combinatorial approach yielded whole genomic contiguous sequences in a cost-effective manner. Over 1,000-time coverage of chloroplast from total DNA were reached by the SOLiD platform in a single spot on a quadrant slide without purification. Comparative analysis indicated that the chloroplast genome was conserved in gene number and organization with respect to the reference genome of L. minor. However, higher nucleotide substitution, abundant deletions and insertions occurred in non-coding regions of these genomes, indicating a greater genomic dynamics than expected from the comparison of other related species in the Pooideae. Noticeably, there was no transition bias over transversion in Lemnoideae. The data should have immediate applications in evolutionary biology and plant taxonomy with increased resolution and statistical power.

  3. Production of biopharmaceuticals and vaccines in plants via the chloroplast genome.

    Science.gov (United States)

    Daniell, Henry

    2006-10-01

    Transgenic plants offer many advantages, including low cost of production (by elimination of fermenters), storage and transportation; heat stability; and absence of human pathogens. When therapeutic proteins are orally delivered, plant cells protect antigens in the stomach through bioencapsulation and eliminate the need for expensive purification and sterile injections, in addition to development of both systemic and mucosal immunity. Chloroplast genetic engineering offers several advantages, including high levels of transgene expression, transgene containment via maternal inheritance and multi-gene expression in a single transformation event. Hyper-expression of vaccine antigens against cholera, tetanus, anthrax, plague or canine parvovirus (4-31% of total soluble protein, tsp) in transgenic chloroplasts (leaves) or non-green plastids (carrots, tomato), as well as the availability of antibiotic-free selectable markers or the ability to excise selectable marker genes, facilitate oral delivery. Hyper-expression of several therapeutic proteins, including human serum albumin (11.1% tsp), somatotropin (7% tsp), interferon-gamma (6% tsp), anti-microbial peptide (21.5% tsp), facilitates efficient and economic purification. Also, the presence of chaperones and enzymes in chloroplasts facilitate assembly of complex multi-subunit proteins and correct folding of human blood proteins with proper disulfide bonds. Functionality of chloroplast-derived vaccine antigens and therapeutic proteins has been demonstrated by several assays, including the macrophage lysis assay, GM1-ganglioside binding assay, protection of HeLa cells or human lung carcinoma cells against encephalomyocarditis virus, systemic immune response, protection against pathogen challenge, and growth or inhibition of cell cultures. Thus, transgenic chloroplasts are ideal bioreactors for production of functional human and animal therapeutic proteins in an environmentally friendly manner.

  4. A dynamic phase microscopic study of optical characteristics of individual chloroplasts.

    Science.gov (United States)

    Tychinsky, V P; Kretushev, A V; Vyshenskaya, T V; Tikhonov, A N

    2004-10-11

    Dynamic phase microscopy (DPM) allows the monitoring of optical path difference (or phase height), h(x,y,t) approximately integraln(x,y,z,t)dz, an integral refractive index projection of the medium, n(x,y,z,t), in optically transparent biological specimens at high spatial and temporal resolutions. In this study, DPM was used for the analysis of fluctuations in the optical characteristics of individual bean chloroplasts in various metabolic states. A "phase image" of an individual chloroplast, which represents a three-dimensional plot of the "phase height", was obtained for the first time, and the frequency spectra of the fluctuations of h(x,y,t) were investigated. The fluctuation patterns, i.e., the intensity and the frequency spectra of phase height fluctuations in bean chloroplasts (Class B) were found to depend on their metabolic state. Under conditions of noncyclic (or pseudocyclic) electron transport, the fluctuations displayed characteristic frequencies in the range of 0.25-0.6 Hz and were space-time-correlated in the chloroplast domains with the cross sizes of approximately 2 microm. The fluctuation intensity decreased in the presence of uncouplers (nigericin and valinomycin, 20 microM). A stronger (in comparison with 20 microM valinomycin) effect of 20 microM nigericin suggests that the light-induced generation of the transmembrane pH difference (DeltapH) makes the main contribution to the increment of space-correlated fluctuations of h(x,y,t). Studies of chloroplasts incubated in media of various osmolarity (50-500 mM sucrose) have shown that structural changes in thylakoids are among other factors responsible for phase height fluctuations.

  5. A set of primers for analyzing chloroplast DNA diversity in Citrus and related genera.

    Science.gov (United States)

    Cheng, Yunjiang; de Vicente, M Carmen; Meng, Haijun; Guo, Wenwu; Tao, Nengguo; Deng, Xiuxin

    2005-06-01

    Chloroplast simple sequence repeat (cpSSR) markers in Citrus were developed and used to analyze chloroplast diversity of Citrus and closely related genera. Fourteen cpSSR primer pairs from the chloroplast genomes of tobacco (Nicotiana tabacum L.) and Arabidopsis were found useful for analyzing the Citrus chloroplast genome (cpDNA) and recoded with the prefix SPCC (SSR Primers for Citrus Chloroplast). Eleven of the 14 primer pairs revealed some degree of polymorphism among 34 genotypes of Citrus, Fortunella, Poncirus and some of their hybrids, with polymorphism information content (PIC) values ranging from 0.057 to 0.732, and 18 haplotypes were identified. The cpSSR data were analyzed with NTSYS-pc software, and the genetic relationships suggested by the unweighted pair group method based on arithmetic means (UPGMA) dendrogram were congruent with previous taxonomic investigations: the results showed that all samples fell into seven major clusters, i.e., Citrus medica L., Poncirus, Fortunella, C. ichangensis Blanco, C. reticulata Swingle, C. aurantifolia (Christm.) Swingle and C. grandis (L.) Osbeck. The results of previous studies combined with our cpSSR analyses revealed that: (1) Calamondin (C. madurensis Swingle) is the result of hybridization between kumquat (Fortunella) and mandarin (C. reticulata), where kumquat acted as the female parent; (2) Ichang papeda (C. ichangensis) has a unique taxonomic status; and (3) although Bendiguangju mandarin (C. reticulata) and Satsuma mandarin (C. reticulata) are similar in fruit shape and leaf morphology, they have different maternal parents. Bendiguangju mandarin has the same cytoplasm as sweet orange (C. sinensis), whereas Satsuma mandarin has the cytoplasm of C. reticulata. Seventeen PCR products from SPCC1 and 21 from SPCC11 were cloned and sequenced. The results revealed that mononucleotide repeats as well as insertions and deletions of small segments of DNA were associated with SPCC1 polymorphism, whereas polymorphism

  6. Identification of processed Chinese medicinal materials using DNA mini-barcoding.

    Science.gov (United States)

    Song, Ming; Dong, Gang-Qiang; Zhang, Ya-Qin; Liu, Xia; Sun, Wei

    2017-07-01

    Most of Chinese medicinal herbs are subjected to traditional processing procedures, including stir-frying, charring, steaming, boiling, and calcining before they are released into dispensaries. The marketing and identification of processed medicinal materials is a growing issue in the marketplace. However, conventional methods of identification have limitations, while DNA mini-barcoding, based on the sequencing of a short-standardized region, has received considerable attention as a new potential means to identify processed medicinal materials. In the present study, six DNA barcode loci including ITS2, psbA-trnH, rbcL, matK, trnL (UAA) intron and its P6 loop, were employed for the authentication of 45 processed samples belonging to 15 species. We evaluated the amplification efficiency of each locus. We also examined the identification accuracy of the potential mini-barcode locus, of trnL (UAA) intron P6 loop. Our results showed that the five primary barcode loci were successfully amplified in only 8.89%-20% of the processed samples, while the amplification rates of the trnL (UAA) intron P6 loop were higher, at 75.56% successful amplification. We compared the mini-barcode sequences with Genbank using the Blast program. The analysis showed that 45.23% samples could be identified to genus level, while only one sample could be identified to the species level. We conclude that trnL (UAA) p6 loop is a candidate mini-barcode that has shown its potential and may become a universal mini-barcode as complementary barcode for authenticity testing and will play an important role in medicinal materials control. Copyright © 2017 China Pharmaceutical University. Published by Elsevier B.V. All rights reserved.

  7. Chloroplast DNA codon use: evidence for selection at the psb A locus based on tRNA availability.

    Science.gov (United States)

    Morton, B R

    1993-09-01

    Codon use in the three sequenced chloroplast genomes (Marchantia, Oryza, and Nicotiana) is examined. The chloroplast has a bias in that codons NNA and NNT are favored over synonymous NNC and NNG codons. This appears to be a consequence of an overall high A + T content of the genome. This pattern of codon use is not followed by the psb A gene of all three genomes and other psb A sequences examined. In this gene, the codon use favors NNC over NNT for twofold degenerate amino acids. In each case the only tRNA coded by the genome is complementary to the NNC codon. This codon use is similar to the codon use by chloroplast genes examined from Chlamydomonas reinhardtii. Since psb A is the major translation product of the chloroplast, this suggests that selection is acting on the codon use of this gene to adapt codons to tRNA availability, as previously suggested for unicellular organisms.

  8. Vaccination via Chloroplast Genetics: Affordable Protein Drugs for the Prevention and Treatment of Inherited or Infectious Human Diseases.

    Science.gov (United States)

    Daniell, Henry; Chan, Hui-Ting; Pasoreck, Elise K

    2016-11-23

    Plastid-made biopharmaceuticals treat major metabolic or genetic disorders, including Alzheimer's, diabetes, hypertension, hemophilia, and retinopathy. Booster vaccines made in chloroplasts prevent global infectious diseases, such as tuberculosis, malaria, cholera, and polio, and biological threats, such as anthrax and plague. Recent advances in this field include commercial-scale production of human therapeutic proteins in FDA-approved cGMP facilities, development of tags to deliver protein drugs to targeted human cells or tissues, methods to deliver precise doses, and long-term stability of protein drugs at ambient temperature, maintaining their efficacy. Codon optimization utilizing valuable information from sequenced chloroplast genomes enhanced expression of eukaryotic human or viral genes in chloroplasts and offered unique insights into translation in chloroplasts. Support from major biopharmaceutical companies, development of hydroponic production systems, and evaluation by regulatory agencies, including the CDC, FDA, and USDA, augur well for advancing this novel concept to the clinic and revolutionizing affordable healthcare.

  9. Ozone-induced changes in the chloroplast structure of conifer needles, and their use in ozone diagnostics

    International Nuclear Information System (INIS)

    Kivimaeenpaeae, M.; Sellden, G.; Sutinen, S.

    2005-01-01

    Ozone induces characteristic symptoms in the chloroplasts of the needles of several coniferous species. Chloroplasts are (1) reduced in size and (2) the stroma is electron dense. Moreover (3) these chloroplast alterations are more pronounced in the outer mesophyll cell layers and in the upper side of the needle compared to the inner layers and lower side. The syndrome, including the three symptoms (1)-(3), is found in the green needles of Scots pine and Norway spruce not only in the experimental fumigations, but also in mature trees in the field, and has potential for diagnosis of ozone stress. For sound ozone diagnostics all three symptoms must be present in the samples studied. The symptoms in relation to needle anatomy and physiology is discussed, and recommendations for sampling and analysis are given. - Ozone-induced alterations in chloroplast structure of conifer needles are reviewed, and recommendations for field monitoring given

  10. Photosynthetic Characteristics and Chloroplast Ultrastructure of Summer Maize Response to Different Nitrogen Supplies.

    Science.gov (United States)

    Liu, Zheng; Gao, Jia; Gao, Fei; Liu, Peng; Zhao, Bin; Zhang, Jiwang

    2018-01-01

    Maize ( Zea mays L.) is the important crop over the world. Nitrogen (N) as necessary element affects photosynthetic characteristics and grain yield of summer maize. In this study, N0 (0 kg N ha -1 ), N1 (129 kg N ha -1 ), N2 (185 kg N ha -1 ), and N3 (300 kg N ha -1 ) was conducted using hybrid 'ZhengDan958' at Dawenkou research field (36°11'N, 117°06'E, 178 m altitude) in the North China Plain to explore the effects of N rate on photosynthetic characteristics and chloroplast ultrastructure. Gas exchange parameters, chlorophyll fluorescence parameters, leaf area index (LAI), chlorophyll SPAD value, chloroplast ultrastructure, dry matter weight and grain yield were measured. At physiological maturity stage, dry matter weight and grain yield of N2 increased by 33-52% ( P ≤ 0.05) and 6-32% ( P ≤ 0.05), respectively, compared with other treatments. During the growing from silking (R1) to milk (R3) stage, LAI of N0 and N1 were 35-38% ( P ≤ 0.05) and 9-23% ( P ≤ 0.05) less than that of N2, respectively. Chlorophyll SPAD value of N0 and N1 were 13-22% ( P ≤ 0.05) and 5-11% ( P ≤ 0.05) lower than that of N2. There was no significant difference in LAI and chlorophyll SPAD value between N2 and N3 during the period from R1 to R3 ( P > 0.05). The net photosynthetic rate ( P n ), maximal quantum efficiency of PSII ( F v / F m ) and quantum efficiency of PSII (Φ PSII ) were higher with the increase of N rate up to N2 ( P ≤ 0.05), and those of N3 were significantly less than N2 ( P ≤ 0.05). In compared with N2, the chloroplast configuration of N0 and N1 became elliptical, almost circular or irregular. The membrane of chloroplast and thylakoid resolved with growing stage, and the number of chloroplast per cell and lamellae per grana decreased under N0 and N1 treatment ( P ≤ 0.05). Under N0 and N1 treatments, summer maize had more negative photosynthetic characteristics. The more number of osmium granule and vesicle and the larger gap between lamellae were

  11. Photosynthetic Characteristics and Chloroplast Ultrastructure of Summer Maize Response to Different Nitrogen Supplies

    Directory of Open Access Journals (Sweden)

    Zheng Liu

    2018-05-01

    Full Text Available Maize (Zea mays L. is the important crop over the world. Nitrogen (N as necessary element affects photosynthetic characteristics and grain yield of summer maize. In this study, N0 (0 kg N ha-1, N1 (129 kg N ha-1, N2 (185 kg N ha-1, and N3 (300 kg N ha-1 was conducted using hybrid ‘ZhengDan958’ at Dawenkou research field (36°11′N, 117°06′E, 178 m altitude in the North China Plain to explore the effects of N rate on photosynthetic characteristics and chloroplast ultrastructure. Gas exchange parameters, chlorophyll fluorescence parameters, leaf area index (LAI, chlorophyll SPAD value, chloroplast ultrastructure, dry matter weight and grain yield were measured. At physiological maturity stage, dry matter weight and grain yield of N2 increased by 33–52% (P ≤ 0.05 and 6–32% (P ≤ 0.05, respectively, compared with other treatments. During the growing from silking (R1 to milk (R3 stage, LAI of N0 and N1 were 35–38% (P ≤ 0.05 and 9–23% (P ≤ 0.05 less than that of N2, respectively. Chlorophyll SPAD value of N0 and N1 were 13–22% (P ≤ 0.05 and 5–11% (P ≤ 0.05 lower than that of N2. There was no significant difference in LAI and chlorophyll SPAD value between N2 and N3 during the period from R1 to R3 (P > 0.05. The net photosynthetic rate (Pn, maximal quantum efficiency of PSII (Fv/Fm and quantum efficiency of PSII (ΦPSII were higher with the increase of N rate up to N2 (P ≤ 0.05, and those of N3 were significantly less than N2 (P ≤ 0.05. In compared with N2, the chloroplast configuration of N0 and N1 became elliptical, almost circular or irregular. The membrane of chloroplast and thylakoid resolved with growing stage, and the number of chloroplast per cell and lamellae per grana decreased under N0 and N1 treatment (P ≤ 0.05. Under N0 and N1 treatments, summer maize had more negative photosynthetic characteristics. The more number of osmium granule and vesicle and the larger gap between lamellae were shown in N3

  12. Chloroplast genes are expressed during intracellular symbiotic association of Vaucheria litorea plastids with the sea slug Elysia chlorotica.

    OpenAIRE

    Mujer, C V; Andrews, D L; Manhart, J R; Pierce, S K; Rumpho, M E

    1996-01-01

    The marine slug Elysia chlorotica (Gould) forms an intracellular symbiosis with photosynthetically active chloroplasts from the chromophytic alga Vaucheria litorea (C. Agardh). This symbiotic association was characterized over a period of 8 months during which E. chlorotica was deprived of V. litorea but provided with light and CO2. The fine structure of the symbiotic chloroplasts remained intact in E. chlorotica even after 8 months of starvation as revealed by electron microscopy. Southern b...

  13. AT_CHLORO, a comprehensive chloroplast proteome database with subplastidial localization and curated information on envelope proteins.

    Science.gov (United States)

    Ferro, Myriam; Brugière, Sabine; Salvi, Daniel; Seigneurin-Berny, Daphné; Court, Magali; Moyet, Lucas; Ramus, Claire; Miras, Stéphane; Mellal, Mourad; Le Gall, Sophie; Kieffer-Jaquinod, Sylvie; Bruley, Christophe; Garin, Jérôme; Joyard, Jacques; Masselon, Christophe; Rolland, Norbert

    2010-06-01

    Recent advances in the proteomics field have allowed a series of high throughput experiments to be conducted on chloroplast samples, and the data are available in several public databases. However, the accurate localization of many chloroplast proteins often remains hypothetical. This is especially true for envelope proteins. We went a step further into the knowledge of the chloroplast proteome by focusing, in the same set of experiments, on the localization of proteins in the stroma, the thylakoids, and envelope membranes. LC-MS/MS-based analyses first allowed building the AT_CHLORO database (http://www.grenoble.prabi.fr/protehome/grenoble-plant-proteomics/), a comprehensive repertoire of the 1323 proteins, identified by 10,654 unique peptide sequences, present in highly purified chloroplasts and their subfractions prepared from Arabidopsis thaliana leaves. This database also provides extensive proteomics information (peptide sequences and molecular weight, chromatographic retention times, MS/MS spectra, and spectral count) for a unique chloroplast protein accurate mass and time tag database gathering identified peptides with their respective and precise analytical coordinates, molecular weight, and retention time. We assessed the partitioning of each protein in the three chloroplast compartments by using a semiquantitative proteomics approach (spectral count). These data together with an in-depth investigation of the literature were compiled to provide accurate subplastidial localization of previously known and newly identified proteins. A unique knowledge base containing extensive information on the proteins identified in envelope fractions was thus obtained, allowing new insights into this membrane system to be revealed. Altogether, the data we obtained provide unexpected information about plastidial or subplastidial localization of some proteins that were not suspected to be associated to this membrane system. The spectral counting-based strategy was further

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

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    Dheeraj Verma

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

  15. Chloroplast RNA-Binding Protein RBD1 Promotes Chilling Tolerance through 23S rRNA Processing in Arabidopsis.

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    Shuai Wang

    2016-05-01

    Full Text Available Plants have varying abilities to tolerate chilling (low but not freezing temperatures, and it is largely unknown how plants such as Arabidopsis thaliana achieve chilling tolerance. Here, we describe a genome-wide screen for genes important for chilling tolerance by their putative knockout mutants in Arabidopsis thaliana. Out of 11,000 T-DNA insertion mutant lines representing half of the genome, 54 lines associated with disruption of 49 genes had a drastic chilling sensitive phenotype. Sixteen of these genes encode proteins with chloroplast localization, suggesting a critical role of chloroplast function in chilling tolerance. Study of one of these proteins RBD1 with an RNA binding domain further reveals the importance of chloroplast translation in chilling tolerance. RBD1 is expressed in the green tissues and is localized in the chloroplast nucleoid. It binds directly to 23S rRNA and the binding is stronger under chilling than at normal growth temperatures. The rbd1 mutants are defective in generating mature 23S rRNAs and deficient in chloroplast protein synthesis especially under chilling conditions. Together, our study identifies RBD1 as a regulator of 23S rRNA processing and reveals the importance of chloroplast function especially protein translation in chilling tolerance.

  16. Delaying chloroplast turnover increases water-deficit stress tolerance through the enhancement of nitrogen assimilation in rice.

    Science.gov (United States)

    Sade, Nir; Umnajkitikorn, Kamolchanok; Rubio Wilhelmi, Maria Del Mar; Wright, Matthew; Wang, Songhu; Blumwald, Eduardo

    2018-02-12

    Abiotic stress-induced senescence in crops is a process particularly affecting the photosynthetic apparatus, decreasing photosynthetic activity and inducing chloroplast degradation. A pathway for stress-induced chloroplast degradation that involves the CHLOROPLAST VESICULATION (CV) gene was characterized in rice (Oryza sativa) plants. OsCV expression was up-regulated with the age of the plants and when plants were exposed to water-deficit conditions. The down-regulation of OsCV expression contributed to the maintenance of the chloroplast integrity under stress. OsCV-silenced plants displayed enhanced source fitness (i.e. carbon and nitrogen assimilation) and photorespiration, leading to water-deficit stress tolerance. Co-immunoprecipitation, intracellular co-localization, and bimolecular fluorescence demonstrated the in vivo interaction between OsCV and chloroplastic glutamine synthetase (OsGS2), affecting source-sink relationships of the plants under stress. Our results would indicate that the OsCV-mediated chloroplast degradation pathway is involved in the regulation of nitrogen assimilation during stress-induced plant senescence. © The Author(s) 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  17. The complete chloroplast genome of Gracilariopsis lemaneiformis (Rhodophyta) gives new insight into the evolution of family Gracilariaceae.

    Science.gov (United States)

    Du, Qingwei; Bi, Guiqi; Mao, Yunxiang; Sui, Zhenghong

    2016-06-01

    The complete chloroplast genome of Gracilariopsis lemaneiformis was recovered from a Next Generation Sequencing data set. Without quadripartite structure, this chloroplast genome (183,013 bp, 27.40% GC content) contains 202 protein-coding genes, 34 tRNA genes, 3 rRNA genes, and 1 tmRNA gene. Synteny analysis showed plasmid incorporation regions in chloroplast genomes of three species of family Gracilariaceae and in Grateloupia taiwanensis of family Halymeniaceae. Combined with reported red algal plasmid sequences in nuclear and mitochondrial genomes, we postulated that red algal plasmids may have played an important role in ancient horizontal gene transfer among nuclear, chloroplast, and mitochondrial genomes. Substitution rate analysis showed that purifying selective forces maintaining stability of protein-coding genes of nine red algal chloroplast genomes over long periods must be strong and that the forces acting on gene groups and single genes of nine red algal chloroplast genomes were similar and consistent. The divergence of Gp. lemaneiformis occurred ~447.98 million years ago (Mya), close to the divergence time of genus Pyropia and Porphyra (443.62 Mya). © 2016 Phycological Society of America.

  18. Chloroplast-derived vaccine antigens and biopharmaceuticals: protocols for expression, purification, or oral delivery and functional evaluation.

    Science.gov (United States)

    Singh, N Dolendro; Ding, Yi; Daniell, Henry

    2009-01-01

    Many vaccine antigens and biopharmaceutical proteins have been expressed at high levels via the chloroplast genome and their functionality has been evaluated using in vitro assays in cell cultures (i.e., macrophage lysis assay, inhibition of vesicular stomatitis virus-induced cytopathicity in baby hamster kidney cells, or inhibition of human HIV infection in TZM-BL cells) as well as protection after challenge with bacterial or viral pathogens or antitumor assays or delay the onset of insulitis in suitable animal models. Production of therapeutic proteins in chloroplasts eliminates the expensive fermentation technology. Moreover, oral delivery of chloroplast-derived therapeutic proteins eliminates expensive purification steps, cold storage, cold transportation, and delivery via sterile needles, thereby further decreasing their cost. In this chapter, we describe detailed protocols for chloroplast transformation including the construction of chloroplast transformation vectors, delivery of DNA into plant cells using particle bombardment, selection and regeneration of transformants by tissue culture, confirmation of transgene integration into the chloroplast genome and homoplasmy, evaluation of foreign gene expression, purification of foreign protein, or oral delivery via bioencapsulation, functional evaluation using in vitro and in vivo assays, and evaluation of immunity after challenge with pathogens in suitable animal models.

  19. Electromagnetic probes of molecular motors in the electron transport chains of mitochondria and chloroplasts

    Science.gov (United States)

    Miller, J. H., Jr.; Nawarathna, D.; Vajrala, V.; Gardner, J.; Widger, W. R.

    2005-12-01

    We report on measurements of harmonics generated by whole cells, mitochondria, and chloroplasts in response to applied sinusoidal electric fields. The frequency- and amplitude-dependence of the induced harmonics exhibit features that correlate with physiological processes. Budding yeast (S. cerevisiae) cells produce numerous harmonics, the amplitudes of which depend strongly on frequency. When the second or third harmonic amplitude is plotted vs. applied frequency, we observe two peaks, around 3 kHz and 12 kHz, which are suppressed by respiratory inhibitors. We observe similar peaks when measuring the harmonic response of B. indicas, a relative of the mitochondrial ancestor. In uncoupled mitochondria, in which most of the electron transport chain is active but the ATP-synthase molecular turbine is inactive, only one (lower frequency) of the two peaks is present. Finally, we find that harmonics generated by chloroplasts depend dramatically on incident light, and vanish in the absence of light.

  20. DISRUPTION OF ARABIDOPSIS RETICULON GENE RTNLB16 RESULTS IN CHLOROPLAST DYSFUNCTION AND OXIDATIVE STRESS

    Directory of Open Access Journals (Sweden)

    Tarasenko V.I.

    2012-08-01

    Full Text Available Reticulons (RTNs are endoplasmic reticulum (ER-localized proteins that have recently attracted much attention. RTNs are ubiquitous proteins present in all eukaryotic organisms examined so far. In animal and yeast, in which knowledge of this protein family is more advanced, RTNs are involved in numerous cellular processes such as apoptosis, cell division and intracellular trafficking. Up to now, a little attention has been paid to their plant counterparts, RTNLBs. Meanwhile, gene search across sequenced genomes revealed that the RTN gene family is more diverse and numerous in plants than in animals and yeasts, which possibly suggests existence of functions specific for plant RTNs. Recently, the localization in different ER regions was shown for two members of plant reticulon family. The location in close proximity to chloroplast membrane was revealed for one of RTNLBs, which is argument in favor of its role in interorganellar interactions. In spite of growing interest towards to plant RTNs, there are no investigations devoted to insertion mutagenesis of genes encoding these proteins. We have genotyped an Arabidopsis line containing T-DNA insertion in RTNLB16 gene encoding uncharacterized member of RTNLB family. The obtained homozygous plants have marked phenotype expressed in a decreased growth rate and a pale-green leaf color. The leaf total chlorophyll content as well as the chlorophyll a/b ratio was significantly lower in mutant plants. It is interesting to note that the extent of phenotypic expression depended on a light intensity. The growth rate of wild-type and mutant plants was the same in low light conditions. The growth rate was significantly decreased and chlorophyll content was 3-5-fold lower in mutant plants growing under moderate light conditions. The growing of plants under high light conditions led to halted growth and death of mutants on the seedling stage. The demonstrated phenotype probably points out to a chloroplast

  1. The complete chloroplast genome sequence of the medicinal plant Andrographis paniculata.

    Science.gov (United States)

    Ding, Ping; Shao, Yanhua; Li, Qian; Gao, Junli; Zhang, Runjing; Lai, Xiaoping; Wang, Deqin; Zhang, Huiye

    2016-07-01

    The complete chloroplast genome of Andrographis paniculata, an important medicinal plant with great economic value, has been studied in this article. The genome size is 150,249 bp in length, with 38.3% GC content. A pair of inverted repeats (IRs, 25,300 bp) are separated by a large single copy region (LSC, 82,459 bp) and a small single-copy region (SSC, 17,190 bp). The chloroplast genome contains 114 unique genes, 80 protein-coding genes, 30 tRNA genes and 4 rRNA genes. In these genes, 15 genes contained 1 intron and 3 genes comprised of 2 introns.

  2. Comparative effects of glyphosate and atrazine in chloroplast ultrastructure of wheat and downy brome

    International Nuclear Information System (INIS)

    Auge, R.M.; Gealy, D.R.; Ogg, A.G.; Franceschi, V.R.

    1987-01-01

    Developing and mature leaves of winter wheat (Triticum aestivum L. var. Daws) and the weed species downy brome (Bromus tectorum L.) were subjected to 10 mM (foliar application) and 1 mM (root application) herbicide solutions. Glyphosate (N-(phosphonomethyl) glycine) and atrazine (2-chloro-4-(ethyl-amino)-6-(isopropylamino)-s-triazine) were prepared in a carrier composed of 5% soybean oil concentrate, 35% acetone and 60% water. Penetration experiments with 3 H-labelled herbicides assessed what percentage of herbicide entered leaves, and microautoradiography was used to determine qualitatively how much herbicide was present in the sections viewed with TEM. Tissue was excised at 4, 18, 62 and 200 hours, and then either freeze-substituted or fixed chemically. Ultrastructural effects of each herbicide on chloroplasts from leaves of newly-germinated seedlings and of well-tillered plants are depicted and discussed. Temporal differences in response of chloroplasts to each herbicide are noted

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

    DEFF Research Database (Denmark)

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

    Manganese is important for molecular functions in plants, i.e. as a co-factor in enzymes and in the oxygen evolving complex of photosystem II, located like most of the photosynthetic machinery, in the thylakoid membranes of chloroplasts. Soils that lack plant available micronutrients such as mang......Manganese is important for molecular functions in plants, i.e. as a co-factor in enzymes and in the oxygen evolving complex of photosystem II, located like most of the photosynthetic machinery, in the thylakoid membranes of chloroplasts. Soils that lack plant available micronutrients...... involved in manganese use efficiency, focusing on the phosphoproteome from thylakoid preparations from two barley genotypes, manganese efficient (Vanessa) and inefficient (Antonia) genotype. Experimental: By monitoring the photosynthetic efficiency (Fv/Fm) a decline in activity is observed as a consequence...

  4. Influence of nitrogen deficiency on photosynthesis and chloroplast ultrastructure of pepper plants (Research Note

    Directory of Open Access Journals (Sweden)

    S. DONCHEVA

    2008-12-01

    Full Text Available Pepper plants (Capsicum annuum L. cv. Zlaten Medal were grown on nutrient solution without nitrogen, and photosynthetic response of plants was examined by determination of leaf CO2 fixation and chlorophyll and carotenoid contents. The absence of nitrogen in the medium resulted in a decrease of the leaf area and of plant biomass accumulation, and in an increase of the root-shoot dry weight ratio. The photosynthetic activity and chlorophyll and carotenoid contents decreased significantly under nitrogen deprivation. Examination of nitrogen deficient leaves by transmission electron microscopy showed dramatic changes in chloroplast ultrastructure. The proportion of starch granules and plastoglobules in the stroma matrix was increased and internal membrane system was greatly reduced. It seems that nitrogen plays an important role in the formation of chloroplast structure and hence to the photosynthetic intensity and productivity of pepper plants.

  5. A chloroplast pathway for the de novo biosynthesis of triacylglycerol in Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Fan, J.; Xu, C.; Andre, C.

    2011-06-23

    Neutral lipid metabolism has been extensively studied in yeast, plants and mammals. In contrast, little information is available regarding the biochemical pathway, enzymes and regulatory factors involved in the biosynthesis of triacylglycerol (TAG) in microalgae. In the conventional TAG biosynthetic pathway widely accepted for yeast, plants and mammals, TAG is assembled in the endoplasmic reticulum (ER) from its immediate precursor diacylglycerol (DAG) made by ER-specific acyltransferases, and is deposited exclusively in lipid droplets in the cytosol. Here, we demonstrated that the unicellular microalga Chlamydomonas reinhardtii employs a distinct pathway that uses DAG derived almost exclusively from the chloroplast to produce TAG. This unique TAG biosynthesis pathway is largely dependent on de novo fatty acid synthesis, and the TAG formed in this pathway is stored in lipid droplets in both the chloroplast and the cytosol. These findings have wide implications for understanding TAG biosynthesis and storage and other areas of lipid metabolism in microalgae and other organisms.

  6. Changes of chloroplast pigments of maize leaves after space flight in recoverable satellite

    International Nuclear Information System (INIS)

    Li Sherong; Zhu Baoge; Liu Genqi

    2001-01-01

    Dried seeds of maize inbred lines were carried by recoverable satellite flying at an altitude of 175-253 km from sea level. The changes of absorption spectra of acetone extracts and chloroplast pigment contents of maize leaves were studied. It showed that the light-absorption characteristics of space-flight treatment (SP) were quite similar to those of the corresponding ground controls (CK) at the same time of sampling. However, the absorbance of the SP were less than CK at absorption peaks of chlorophyll a and b, respectively. The contents of chlorophyll a and chlorophyll b of SP were significantly reduced, and the reduction of chlorophyll b far exceeded chlorophyll a. The contents of chlorophyll a + b were reduced so much that the total amount of their chloroplast pigments was lowered, but Ca/Cb ratio tended to be higher in comparison with CK

  7. Chloroplast and mitochondrial microsatellites for Millettia pinnata (Fabaceae) and cross-amplification in related species.

    Science.gov (United States)

    Wang, Yanling; Xie, Hongxian; Yang, Yi; Huang, Yelin; Wang, Jianwu; Tan, Fengxiao

    2017-05-01

    Chloroplast and mitochondrial microsatellites were identified to study the population genetics of Millettia pinnata (Fabaceae). Based on publicly available plastid genome sequence data of M. pinnata , 42 primer pairs were developed, of which 17 displayed polymorphisms across 89 individuals from four populations. For chloroplast loci, two to six alleles were recovered and the unbiased haploid diversity per locus ranged from 0.391 to 0.857. For mitochondrial loci, two to four alleles were recovered and the unbiased haploid diversity ranged from 0.264 to 0.740. Sixteen of the 17 screened markers could be successfully amplified in the related species M. pulchra . The 17 microsatellite markers developed here exhibited variation in M. pinnata and 16 presented transferability in the related species M. pulchra , suggesting that these markers will be valuable for genetic studies across M. pinnata and its related species.

  8. Chloroplast and mitochondrial microsatellites for Millettia pinnata (Fabaceae) and cross-amplification in related species1

    Science.gov (United States)

    Wang, Yanling; Xie, Hongxian; Yang, Yi; Huang, Yelin; Wang, Jianwu; Tan, Fengxiao

    2017-01-01

    Premise of the study: Chloroplast and mitochondrial microsatellites were identified to study the population genetics of Millettia pinnata (Fabaceae). Methods and Results: Based on publicly available plastid genome sequence data of M. pinnata, 42 primer pairs were developed, of which 17 displayed polymorphisms across 89 individuals from four populations. For chloroplast loci, two to six alleles were recovered and the unbiased haploid diversity per locus ranged from 0.391 to 0.857. For mitochondrial loci, two to four alleles were recovered and the unbiased haploid diversity ranged from 0.264 to 0.740. Sixteen of the 17 screened markers could be successfully amplified in the related species M. pulchra. Conclusions: The 17 microsatellite markers developed here exhibited variation in M. pinnata and 16 presented transferability in the related species M. pulchra, suggesting that these markers will be valuable for genetic studies across M. pinnata and its related species. PMID:28529836

  9. The complete chloroplast genome of an irreplaceable dietary and model crop, foxtail millet (Setaria italica).

    Science.gov (United States)

    Wang, Shuo; Gao, Li-Zhi

    2016-11-01

    The complete chloroplast genome sequence of foxtail millet (Setaria italica), an important food and fodder crop in the family Poaceae, is first reported in this study. The genome consists of 1 35 516 bp containing a pair of inverted repeats (IRs) of 21 804 bp separated by a large single-copy (LSC) region and a small single-copy (SSC) region of 79 896 bp and 12 012 bp, respectively. Coding sequences constitute 58.8% of the genome harboring 111 unique genes, 71 of which are protein-coding genes, 4 are rRNA genes, and 36 are tRNA genes. Phylogenetic analysis indicated foxtail millet clustered with Panicum virgatum and Echinochloa crus-galli belonging to the tribe Paniceae of the subfamily Panicoideae. This newly determined chloroplast genome will provide valuable information for the future breeding programs of valuable cereal crops in the family Poaceae.

  10. Changes induced by the Pepper mild mottle tobamovirus on the chloroplast proteome of Nicotiana benthamiana.

    Science.gov (United States)

    Pineda, M; Sajnani, C; Barón, M

    2010-01-01

    We have analyzed the chloroplast proteome of Nicotiana benthamiana using two-dimensional gel electrophoresis and mass spectrometry followed by a database search. In order to improve the resolution of the two-dimensional electrophoresis gels, we have made separate maps for the low and the high pH range. At least 200 spots were detected. We identified 72 polypeptides, some being isoforms of different multiprotein families. In addition, changes in this chloroplast proteome induced by the infection with the Spanish strain of the Pepper mild mottle virus were investigated. Viral infection induced the down-regulation of several chloroplastidic proteins involved in both the photosynthetic electron-transport chain and the Benson-Calvin cycle.

  11. Complete sequence and comparative analysis of the chloroplast genome of coconut palm (Cocos nucifera).

    Science.gov (United States)

    Huang, Ya-Yi; Matzke, Antonius J M; Matzke, Marjori

    2013-01-01

    Coconut, a member of the palm family (Arecaceae), is one of the most economically important trees used by mankind. Despite its diverse morphology, coconut is recognized taxonomically as only a single species (Cocos nucifera L.). There are two major coconut varieties, tall and dwarf, the latter of which displays traits resulting from selection by humans. We report here the complete chloroplast (cp) genome of a dwarf coconut plant, and describe the gene content and organization, inverted repeat fluctuations, repeated sequence structure, and occurrence of RNA editing. Phylogenetic relationships of monocots were inferred based on 47 chloroplast protein-coding genes. Potential nodes for events of gene duplication and pseudogenization related to inverted repeat fluctuation were mapped onto the tree using parsimony criteria. We compare our findings with those from other palm species for which complete cp genome sequences are available.

  12. Efficient in vitro import of a cytosolic heat shock protein into pea chloroplasts

    OpenAIRE

    Lubben, Thomas H.; Keegstra, Kenneth

    1986-01-01

    In order to further our understanding of the targeting of nuclear-encoded proteins into intracellular organelles, we have investigated the import of chimeric precursor proteins into pea chloroplasts. Two different chimeric precursor proteins were produced by in vitro expression of chimeric genes. One chimeric precursor contained the transit peptide of the small subunit of soybean ribulose 1,5-bisphosphate carboxylase and the mature peptide of the same protein from pea. The second contained th...

  13. Characterisation by nuclear magnetic resonance of the β catalytic subunit of the chloroplastic coupling factor

    International Nuclear Information System (INIS)

    Andre, Francois

    1986-09-01

    This academic work addressed the use of nuclear magnetic resonance (NMR) for the structural and dynamic study of the catalytic sub-unit of the extrinsic section of a membrane complex, the chloroplastic H+-ATPase. This work included the development of a protocol of preparation and quantitative purification of β subunits isolated from the CF1 for the elaboration of a concentrated sample for NMR, and then the study of the β subunit by using proton NMR

  14. A family of selfish minicircular chromosomes with jumbled chloroplast gene fragments from a dinoflagellate.

    Science.gov (United States)

    Zhang, Z; Cavalier-Smith, T; Green, B R

    2001-08-01

    Chloroplast genes of several dinoflagellate species are located on unigenic DNA minicircular chromosomes. We have now completely sequenced five aberrant minicircular chromosomes from the dinoflagellate Heterocapsa triquetra. These probably nonfunctional DNA circles lack complete genes, with each being composed of several short fragments of two or three different chloroplast genes and a common conserved region with a tripartite 9G-9A-9G core like the putative replicon origin of functional single-gene circular chloroplast chromosomes. Their sequences imply that all five circles evolved by differential deletions and duplications from common ancestral circles bearing fragments of four genes: psbA, psbC, 16S rRNA, and 23S rRNA. It appears that recombination between separate unigenic chromosomes initially gave intermediate heterodimers, which were subsequently stabilized by deletions that included part or all of one putative replicon origin. We suggest that homologous recombination at the 9G-9A-9G core regions produced a psbA/psbC heterodimer which generated two distinct chimeric circles by differential deletions and duplications. A 23S/16S rRNA heterodimer more likely formed by illegitimate recombination between 16S and 23S rRNA genes. Homologous recombination between the 9G-9A-9G core regions of both heterodimers and additional differential deletions and duplications could then have yielded the other three circles. Near identity of the gene fragments and 9G-9A-9G cores, despite diverging adjacent regions, may be maintained by gene conversion. The conserved organization of the 9G-9A-9G cores alone favors the idea that they are replicon origins and suggests that they may enable the aberrant minicircles to parasitize the chloroplast's replication machinery as selfish circles.

  15. The Role of Heterologous Chloroplast Sequence Elements in Transgene Integration and Expression1[W][OA

    Science.gov (United States)

    Ruhlman, Tracey; Verma, Dheeraj; Samson, Nalapalli; Daniell, Henry

    2010-01-01

    Heterologous regulatory elements and flanking sequences have been used in chloroplast transformation of several crop species, but their roles and mechanisms have not yet been investigated. Nucleotide sequence identity in the photosystem II protein D1 (psbA) upstream region is 59% across all taxa; similar variation was consistent across all genes and taxa examined. Secondary structure and predicted Gibbs free energy values of the psbA 5′ untranslated region (UTR) among different families reflected this variation. Therefore, chloroplast transformation vectors were made for tobacco (Nicotiana tabacum) and lettuce (Lactuca sativa), with endogenous (Nt-Nt, Ls-Ls) or heterologous (Nt-Ls, Ls-Nt) psbA promoter, 5′ UTR and 3′ UTR, regulating expression of the anthrax protective antigen (PA) or human proinsulin (Pins) fused with the cholera toxin B-subunit (CTB). Unique lettuce flanking sequences were completely eliminated during homologous recombination in the transplastomic tobacco genomes but not unique tobacco sequences. Nt-Ls or Ls-Nt transplastomic lines showed reduction of 80% PA and 97% CTB-Pins expression when compared with endogenous psbA regulatory elements, which accumulated up to 29.6% total soluble protein PA and 72.0% total leaf protein CTB-Pins, 2-fold higher than Rubisco. Transgene transcripts were reduced by 84% in Ls-Nt-CTB-Pins and by 72% in Nt-Ls-PA lines. Transcripts containing endogenous 5′ UTR were stabilized in nonpolysomal fractions. Stromal RNA-binding proteins were preferentially associated with endogenous psbA 5′ UTR. A rapid and reproducible regeneration system was developed for lettuce commercial cultivars by optimizing plant growth regulators. These findings underscore the need for sequencing complete crop chloroplast genomes, utilization of endogenous regulatory elements and flanking sequences, as well as optimization of plant growth regulators for efficient chloroplast transformation. PMID:20130101

  16. The role of heterologous chloroplast sequence elements in transgene integration and expression.

    Science.gov (United States)

    Ruhlman, Tracey; Verma, Dheeraj; Samson, Nalapalli; Daniell, Henry

    2010-04-01

    Heterologous regulatory elements and flanking sequences have been used in chloroplast transformation of several crop species, but their roles and mechanisms have not yet been investigated. Nucleotide sequence identity in the photosystem II protein D1 (psbA) upstream region is 59% across all taxa; similar variation was consistent across all genes and taxa examined. Secondary structure and predicted Gibbs free energy values of the psbA 5' untranslated region (UTR) among different families reflected this variation. Therefore, chloroplast transformation vectors were made for tobacco (Nicotiana tabacum) and lettuce (Lactuca sativa), with endogenous (Nt-Nt, Ls-Ls) or heterologous (Nt-Ls, Ls-Nt) psbA promoter, 5' UTR and 3' UTR, regulating expression of the anthrax protective antigen (PA) or human proinsulin (Pins) fused with the cholera toxin B-subunit (CTB). Unique lettuce flanking sequences were completely eliminated during homologous recombination in the transplastomic tobacco genomes but not unique tobacco sequences. Nt-Ls or Ls-Nt transplastomic lines showed reduction of 80% PA and 97% CTB-Pins expression when compared with endogenous psbA regulatory elements, which accumulated up to 29.6% total soluble protein PA and 72.0% total leaf protein CTB-Pins, 2-fold higher than Rubisco. Transgene transcripts were reduced by 84% in Ls-Nt-CTB-Pins and by 72% in Nt-Ls-PA lines. Transcripts containing endogenous 5' UTR were stabilized in nonpolysomal fractions. Stromal RNA-binding proteins were preferentially associated with endogenous psbA 5' UTR. A rapid and reproducible regeneration system was developed for lettuce commercial cultivars by optimizing plant growth regulators. These findings underscore the need for sequencing complete crop chloroplast genomes, utilization of endogenous regulatory elements and flanking sequences, as well as optimization of plant growth regulators for efficient chloroplast transformation.

  17. Thioredoxin Selectivity for Thiol-based Redox Regulation of Target Proteins in Chloroplasts*

    Science.gov (United States)

    Yoshida, Keisuke; Hara, Satoshi; Hisabori, Toru

    2015-01-01

    Redox regulation based on the thioredoxin (Trx) system is believed to ensure light-responsive control of various functions in chloroplasts. Five Trx subtypes have been reported to reside in chloroplasts, but their functional diversity in the redox regulation of Trx target proteins remains poorly clarified. To directly address this issue, we studied the Trx-dependent redox shifts of several chloroplast thiol-modulated enzymes in vitro and in vivo. In vitro assays using a series of Arabidopsis recombinant proteins provided new insights into Trx selectivity for the redox regulation as well as the underpinning for previous suggestions. Most notably, by combining the discrimination of thiol status with mass spectrometry and activity measurement, we identified an uncharacterized aspect of the reductive activation of NADP-malate dehydrogenase; two redox-active Cys pairs harbored in this enzyme were reduced via distinct utilization of Trxs even within a single polypeptide. In our in vitro assays, Trx-f was effective in reducing all thiol-modulated enzymes analyzed here. We then investigated the in vivo physiological relevance of these in vitro findings, using Arabidopsis wild-type and Trx-f-deficient plants. Photoreduction of fructose-1,6-bisphosphatase was partially impaired in Trx-f-deficient plants, but the global impact of Trx-f deficiency on the redox behaviors of thiol-modulated enzymes was not as striking as expected from the in vitro data. Our results provide support for the in vivo functionality of the Trx system and also highlight the complexity and plasticity of the chloroplast redox network. PMID:25878252

  18. Overexpression of yeast ArDH gene in chloroplasts confers salinity tolerance in plants (abstract)

    International Nuclear Information System (INIS)

    Khan, M.S.; Kanwal, B.; Khalid, A.M.; Zafar, Y.; Malik, K.A.

    2005-01-01

    Water stress due to salinity and drought is the main limiting factor for plant growth, productivity and quality. A common response to water deficit is the accumulation of osmoprotectants such as sugars and amino acids. In yeast, arabitol dehydrogenase is found responsible for the production of arabitol from ribulose-5-phosphate. All plants synthesize ribulose-5-phosphate via pentose pathway in chloroplasts.. Therefore, osmotolerance of the plants could be enhanced through metabolic engineering of chloroplasts by introducing ArDH gene into the plastome, which is responsible for the conversion of ribulose-5- phosphate to arabitol. Here we report high-level expression of arabitol dehydrogenase (ArDH) in chloroplasts. Homoplasmic transgenic plants were recovered on spectinomycin-containing regeneration medium. Transformed tobacco plants survived whereas non-transformed were severely stressed or killed when two weeks old seedlings were exposed to NaCl (up to 400 mM), suggesting a role for arabitol in salt tolerance. Seedlings survived up to five weeks on medium containing high salt concentrations (350-400 mM). Nevertheless, seedlings remained green and grew normal on concentrations up to 350 mM NaCl for several weeks. Hypothesis that membranes are protected under stress conditions due to the arabitol accumulation in chloroplasts, seedlings were grown in liquid medium containing polyethylene glycol (PEG, up to 6%). Seedlings were tolerant to 6% PEG, suggesting that ArDH enzyme protects membranes integrity under stress. Therefore, it is concluded that ArDH gene could be expressed in crop plants to withstand abiotic stresses. (author)

  19. The chloroplast min system functions differentially in two specific nongreen plastids in Arabidopsis thaliana.

    Science.gov (United States)

    Wang, Peng; Zhang, Jie; Su, Jianbin; Wang, Peng; Liu, Jun; Liu, Bing; Feng, Dongru; Wang, Jinfa; Wang, Hongbin

    2013-01-01

    The nongreen plastids, such as etioplasts, chromoplasts, etc., as well as chloroplasts, are all derived from proplastids in the meristem. To date, the Min system members in plants have been identified as regulators of FtsZ-ring placement, which are essential for the symmetrical division of chloroplasts. However, the regulation of FtsZ-ring placement in nongreen plastids is poorly understood. In this study, we investigated the division site placement of nongreen plastids by examining the etioplasts as representative in Arabidopsis Min system mutants. Surprisingly, the shape and number of etioplasts in cotyledons of arc3, arc11 and mcd1 mutants were similar to that observed in wild-type plants, whereas arc12 and parc6 mutants exhibited enlarged etioplasts that were reduced in number. In order to examine nongreen plastids in true leaves, we silenced the ALB3 gene in these Min system mutant backgrounds to produce immature chloroplasts without the thylakoidal network using virus induced gene silencing (VIGS). Interestingly, consistent with our observations in etioplasts, enlarged and fewer nongreen plastids were only detected in leaves of parc6 (VIGS-ALB3) and arc12 (VIGS-ALB3) plants. Further, the FtsZ-ring assembled properly at the midpoint in nongreen plastids of arc3, arc11 and mcd1 (VIGS-ALB3) plants, but organized into multiple rings in parc6 (VIGS-ALB3) and presented fragmented filaments in arc12 (VIGS-ALB3) plants, suggesting that division site placement in nongreen plastids requires fewer components of the plant Min system. Taken together, these results suggest that division site placement in nongreen plastids is different from that in chloroplasts.

  20. The chloroplast min system functions differentially in two specific nongreen plastids in Arabidopsis thaliana.

    Directory of Open Access Journals (Sweden)

    Peng Wang

    Full Text Available The nongreen plastids, such as etioplasts, chromoplasts, etc., as well as chloroplasts, are all derived from proplastids in the meristem. To date, the Min system members in plants have been identified as regulators of FtsZ-ring placement, which are essential for the symmetrical division of chloroplasts. However, the regulation of FtsZ-ring placement in nongreen plastids is poorly understood. In this study, we investigated the division site placement of nongreen plastids by examining the etioplasts as representative in Arabidopsis Min system mutants. Surprisingly, the shape and number of etioplasts in cotyledons of arc3, arc11 and mcd1 mutants were similar to that observed in wild-type plants, whereas arc12 and parc6 mutants exhibited enlarged etioplasts that were reduced in number. In order to examine nongreen plastids in true leaves, we silenced the ALB3 gene in these Min system mutant backgrounds to produce immature chloroplasts without the thylakoidal network using virus induced gene silencing (VIGS. Interestingly, consistent with our observations in etioplasts, enlarged and fewer nongreen plastids were only detected in leaves of parc6 (VIGS-ALB3 and arc12 (VIGS-ALB3 plants. Further, the FtsZ-ring assembled properly at the midpoint in nongreen plastids of arc3, arc11 and mcd1 (VIGS-ALB3 plants, but organized into multiple rings in parc6 (VIGS-ALB3 and presented fragmented filaments in arc12 (VIGS-ALB3 plants, suggesting that division site placement in nongreen plastids requires fewer components of the plant Min system. Taken together, these results suggest that division site placement in nongreen plastids is different from that in chloroplasts.

  1. [Isolation and partial characterization of DNA topoisomerase I from the nucleoids of white mustard chloroplasts].

    Science.gov (United States)

    Belkina, G G; Pogul'skaia, E V; Iurina, N P

    2004-01-01

    DNA topoisomerase was isolated for the first time from nucleoids of white mustard (Sinapis alba L.) chloroplasts. The enzyme had a molecular weight of 70 kDa; it was ATP-independent, required the presence of mono- (K+) and bivalent (Mg2+) cations, and was capable of relaxing both negatively and positively supercoiled DNA. These results suggest that the enzyme isolated belongs to type IB DNA topoisomerases.

  2. Field production and functional evaluation of chloroplast-derived interferon-alpha2b.

    Science.gov (United States)

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

    2007-07-01

    Type I interferons (IFNs) inhibit viral replication and cell growth and enhance the immune response, and therefore have many clinical applications. IFN-alpha2b ranks third in world market use for a biopharmaceutical, behind only insulin and erythropoietin. The average annual cost of IFN-alpha2b 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-alpha2b 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-alpha2b levels reached up to 20% of total soluble protein, or 3 mg per gram of leaf (fresh weight). Transgenic IFN-alpha2b had similar in vitro biological activity to commercially produced PEG-Introntrade mark 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-alpha2b were also seen in vivo. Chloroplast-derived IFN-alpha2b increased the expression of major histocompatibility complex class I (MHC I) on splenocytes and the total number of natural killer (NK) cells. Finally, IFN-alpha2b purified from chloroplast transgenic lines (cpIFN-alpha2b) protected mice from a highly metastatic tumour line. This demonstration of high levels of expression of IFN-alpha2b, transgene containment and biological activity akin to that of commercial preparations of IFN-alpha2b facilitated the first field production of a plant-derived human blood protein, a critical step towards human clinical trials and commercialization.

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

    Science.gov (United States)

    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

  4. Accumulation of chloroplast DNA sequences on the Y chromosome of Silene latifolia

    Czech Academy of Sciences Publication Activity Database

    Kejnovský, Eduard; Kubát, Zdeněk; Hobza, Roman; Lengerová, Martina; Sato, S.; Tabata, S.; Fukui, K.; Matsunaga, S.; Vyskot, Boris

    2006-01-01

    Roč. 128, 1-3 (2006), s. 167-175 ISSN 0016-6707 R&D Projects: GA ČR(CZ) GA204/05/2097; GA ČR(CZ) GD204/05/H505; GA AV ČR(CZ) 1QS500040507 Institutional research plan: CEZ:AV0Z50040507 Keywords : accumulation * chloroplast DNA * Y chromosome Subject RIV: BO - Biophysics Impact factor: 1.492, year: 2006

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

    Czech Academy of Sciences Publication Activity Database

    Polanská, Lenka; Vičánková, Anna; Dobrev, Petre; Macháčková, Ivana; Vaňková, Radomíra

    2004-01-01

    Roč. 26, č. 20 (2004), s. 1549-1555 ISSN 0141-5492 R&D Projects: GA MŠk OC 840.20; GA MŠk LN00A081; GA ČR GA206/03/0369 Institutional research plan: CEZ:AV0Z5038910 Keywords : calcium alginate * chloroplast ultrastructure * cytokinin metabolism Subject RIV: ED - Physiology Impact factor: 0.849, year: 2004

  6. The first complete chloroplast genome sequence of a lycophyte,Huperzia lucidula (Lycopodiaceae)

    Energy Technology Data Exchange (ETDEWEB)

    Wolf, Paul G.; Karol, Kenneth G.; Mandoli, Dina F.; Kuehl,Jennifer V.; Arumuganathan, K.; Ellis, Mark W.; Mishler, Brent D.; Kelch,Dean G.; Olmstead, Richard G.; Boore, Jeffrey L.

    2005-02-01

    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 154,373bp, containing inverted repeats of 15,314 bp each, a large single-copy region of 104,088 bp, and a small single-copy region of 19,671 bp. Gene order is more similar to those of mosses, liverworts, and hornworts than to gene order for other vascular plants. For example, the Huperziachloroplast 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 lycophytechloroplast genome data also enable a better reconstruction of the basaltracheophyte 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, 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.

  7. PHYLOGENETIC RELATIONSHIPS AMONG VIETNAMESE COCOA ACCESSIONS USING A NON-CODING REGION OF THE CHLOROPLAST DNA

    OpenAIRE

    Lam Thi, Viet Ha; D.T., Khang; Everaert, Helena; T.N, Dung; P.H.D, Phuoc; H.T., Toan; Dewettinck, Koen; Messens, Kathy

    2017-01-01

    Cocoa (Theobroma cacao L.) cultivation has increased in tropical areas around the world, including Vietnam, due to the high demand of cocoa beans for chocolate production. The genetic diversity of cocoa genotypes is recognized to be complex, however, their phylogenetic relationships need to be clarified. The present study aimed to classify the cocoa genotypes that are imported and cultivated in Vietnam based on a chloroplast DNA region. Sixty-three Vietnamese Cocoa accessions were collected f...

  8. Physiological and Proteomic Analysis in Chloroplasts of Solanum lycopersicum L. under Silicon Efficiency and Salinity Stress

    Directory of Open Access Journals (Sweden)

    Sowbiya Muneer

    2014-11-01

    Full Text Available Tomato plants often grow in saline environments in Mediterranean countries where salt accumulation in the soil is a major abiotic stress that limits its productivity. However, silicon (Si supplementation has been reported to improve tolerance against several forms of abiotic stress. The primary aim of our study was to investigate, using comparative physiological and proteomic approaches, salinity stress in chloroplasts of tomato under silicon supplementation. Tomato seedlings (Solanum lycopersicum L. were grown in nutrient media in the presence or absence of NaCl and supplemented with silicon for 5 days. Salinity stress caused oxidative damage, followed by a decrease in silicon concentrations in the leaves of the tomato plants. However, supplementation with silicon had an overall protective effect against this stress. The major physiological parameters measured in our studies including total chlorophyll and carotenoid content were largely decreased under salinity stress, but were recovered in the presence of silicon. Insufficient levels of net-photosynthesis, transpiration and stomatal conductance were also largely improved by silicon supplementation. Proteomics analysis of chloroplasts analyzed by 2D-BN-PAGE (second-dimensional blue native polyacrylamide-gel electrophoresis revealed a high sensitivity of multiprotein complex proteins (MCPs such as photosystems I (PSI and II (PSII to the presence of saline. A significant reduction in cytochrome b6/f and the ATP-synthase complex was also alleviated by silicon during salinity stress, while the complex forms of light harvesting complex trimers and monomers (LHCs were rapidly up-regulated. Our results suggest that silicon plays an important role in moderating damage to chloroplasts and their metabolism in saline environments. We therefore hypothesize that tomato plants have a greater capacity for tolerating saline stress through the improvement of photosynthetic metabolism and chloroplast proteome

  9. Photosynthesis in a different light: Spectro-microscopy for in vivo characterisation of chloroplasts

    Directory of Open Access Journals (Sweden)

    Sébastien ePeter

    2014-06-01

    Full Text Available During photosynthesis, energy conversion at the two photosystems is controlled by highly complex and dynamic adaptation processes triggered by external factors such as light quality, intensity, and duration, or internal cues such as carbon availability. These dynamics have remained largely concealed so far, because current analytical techniques are based on the investigation of isolated chloroplasts lacking full adaptation ability and are performed at non-physiologically low temperatures. Here, we use non-invasive in planta spectro-microscopic approaches to investigate living chloroplasts in their native environment at ambient temperatures. This is a valuable approach to study the complex function of these systems, because an intrinsic property – the fluorescence emission – is exploited and no additional external perturbations are introduced. Our analysis demonstrates a dynamic adjustment of not only the photosystemI/photosystemII (PSI/PSII intensity ratio in the chloroplasts but also of the capacity of the LHCs for energy transfer in response to environmental and internal cues.

  10. Chloroplast DNA Structural Variation, Phylogeny, and Age of Divergence among Diploid Cotton Species

    Science.gov (United States)

    Li, Pengbo; Liu, Fang; Wang, Yumei; Xu, Qin; Shang, Mingzhao; Zhou, Zhongli; Cai, Xiaoyan; Wang, Xingxing; Wendel, Jonathan F.; Wang, Kunbo

    2016-01-01

    The cotton genus (Gossypium spp.) contains 8 monophyletic diploid genome groups (A, B, C, D, E, F, G, K) and a single allotetraploid clade (AD). To gain insight into the phylogeny of Gossypium and molecular evolution of the chloroplast genome in this group, we performed a comparative analysis of 19 Gossypium chloroplast genomes, six reported here for the first time. Nucleotide distance in non-coding regions was about three times that of coding regions. As expected, distances were smaller within than among genome groups. Phylogenetic topologies based on nucleotide and indel data support for the resolution of the 8 genome groups into 6 clades. Phylogenetic analysis of indel distribution among the 19 genomes demonstrates contrasting evolutionary dynamics in different clades, with a parallel genome downsizing in two genome groups and a biased accumulation of insertions in the clade containing the cultivated cottons leading to large (for Gossypium) chloroplast genomes. Divergence time estimates derived from the cpDNA sequence suggest that the major diploid clades had diverged approximately 10 to 11 million years ago. The complete nucleotide sequences of 6 cpDNA genomes are provided, offering a resource for cytonuclear studies in Gossypium. PMID:27309527

  11. Engineering chloroplasts to improve Rubisco catalysis: prospects for translating improvements into food and fiber crops.

    Science.gov (United States)

    Sharwood, Robert E

    2017-01-01

    494 I. 495 II. 496 III. 496 IV. 499 V. 499 VI. 501 VII. 501 VIII. 502 IX. 505 X. 506 507 References 507 SUMMARY: The uncertainty of future climate change is placing pressure on cropping systems to continue to provide stable increases in productive yields. To mitigate future climates and the increasing threats against global food security, new solutions to manipulate photosynthesis are required. This review explores the current efforts available to improve carbon assimilation within plant chloroplasts by engineering Rubisco, which catalyzes the rate-limiting step of CO 2 fixation. Fixation of CO 2 and subsequent cycling of 3-phosphoglycerate through the Calvin cycle provides the necessary carbohydrate building blocks for maintaining plant growth and yield, but has to compete with Rubisco oxygenation, which results in photorespiration that is energetically wasteful for plants. Engineering improvements in Rubisco is a complex challenge and requires an understanding of chloroplast gene regulatory pathways, and the intricate nature of Rubisco catalysis and biogenesis, to transplant more efficient forms of Rubisco into crops. In recent times, major advances in Rubisco engineering have been achieved through improvement of our knowledge of Rubisco synthesis and assembly, and identifying amino acid catalytic switches in the L-subunit responsible for improvements in catalysis. Improving the capacity of CO 2 fixation in crops such as rice will require further advances in chloroplast bioengineering and Rubisco biogenesis. © 2016 The Author. New Phytologist © 2016 New Phytologist Trust.

  12. Membrane composition and physiological activity of plastids from an oenothera plastome mutator-induced chloroplast mutant.

    Science.gov (United States)

    Johnson, E M; Sears, B B

    1990-01-01

    Plastids were isolated from a plastome mutator-induced mutant (pm7) of Oenothera hookeri and were analyzed for various physiological and biochemical attributes. No photosynthetic electron transport activity was detected in the mutant plastids. This is consistent with previous ultrastructural analysis showing the absence of thylakoid membranes in the pm7 plastids and with the observation of aberrant processing and accumulation of chloroplast proteins in the mutant. In comparison to wild type, the mutant tissue lacks chlorophyll, and has significant differences in levels of four fatty acids. The analyses did not reveal any differences in carotenoid levels nor in the synthesis of several chloroplast lipids. The consequences of the altered composition of the chloroplast membrane are discussed in terms of their relation to the aberrant protein processing of the pm7 plastids. The pigment, fatty acid, and lipid measurements were also performed on two distinct nuclear genotypes (A/A and A/C) which differ in their compatibility with the plastid genome (type I) contained in these lines. In these cases, only chlorophyll concentrations differed significantly.

  13. Chloroplast Preproteins Bind to the Dimer Interface of the Toc159 Receptor during Import1[OPEN

    Science.gov (United States)

    Chen, Lih-Jen; Yeh, Yi-Hung; Hsiao, Chwan-Deng

    2017-01-01

    Most chloroplast proteins are synthesized in the cytosol as higher molecular weight preproteins and imported via the translocons in the outer (TOC) and inner (TIC) envelope membranes of chloroplasts. Toc159 functions as a primary receptor and directly binds preproteins through its dimeric GTPase domain. As a first step toward a molecular understanding of how Toc159 mediates preprotein import, we mapped the preprotein-binding regions on the Toc159 GTPase domain (Toc159G) of pea (Pisum sativum) using cleavage by bound preproteins conjugated with the artificial protease FeBABE and cysteine-cysteine cross-linking. Our results show that residues at the dimer interface and the switch II region of Toc159G are in close proximity to preproteins. The mature portion of preproteins was observed preferentially at the dimer interface, whereas the transit peptide was found at both regions equally. Chloroplasts from transgenic plants expressing engineered Toc159 with a cysteine placed at the dimer interface showed increased cross-linking to bound preproteins. Our data suggest that, during preprotein import, the Toc159G dimer disengages and the dimer interface contacts translocating preproteins, which is consistent with a model in which conformational changes induced by dimer-monomer conversion in Toc159 play a direct role in facilitating preprotein import. PMID:28250068

  14. Evidence for an operative glutamine translocator in chloroplasts from maritime pine (Pinus pinaster Ait.) cotyledons.

    Science.gov (United States)

    Claros, M G; Aguilar, M L; Cánovas, F M

    2010-09-01

    In higher plants, ammonium is assimilated into amino acids through the glutamine synthetase (GS)/glutamate synthase (GOGAT) cycle. This metabolic cycle is distributed in different cellular compartments in conifer seedlings: glutamine synthesis occurs in the cytosol and glutamate synthesis within the chloroplast. A method for preparing intact chloroplasts of pine cotyledons is presented with the aim of identifying a glutamine-glutamate translocator. Glutamine-glutamate exchange has been studied using the double silicone layer system, suggesting the existence of a translocator that imports glutamine into the chloroplast and exports glutamate to the cytoplasm. The translocator identified is specific for glutamine and glutamate, and the kinetic constants for both substrates indicate that it is unsaturated at intracellular concentrations. Thus, the experimental evidence obtained supports the model of the GS/GOGAT cycle in developing pine seedlings that accounts for the stoichiometric balance of metabolites. As a result, the efficient assimilation of free ammonia produced by photorespiration, nitrate reduction, storage protein mobilisation, phenylpropanoid pathway or S-adenosylmethionine synthesis is guaranteed.

  15. Proton gradients and proton-dependent transport processes in the chloroplast

    Directory of Open Access Journals (Sweden)

    Ricarda eHöhner

    2016-02-01

    Full Text Available Proton gradients are fundamental to chloroplast function. Across thylakoid membranes, the light induced proton gradient is essential for ATP synthesis. As a result of proton pumping into the thylakoid lumen, an alkaline stromal pH develops, which is required for full activation of pH-dependent Calvin Benson cycle enzymes. This implies that a pH gradient between the cytosol (pH 7 and the stroma (pH 8 is established upon illumination. To maintain this pH gradient chloroplasts actively extrude protons. More than 30 years ago it was already established that these proton fluxes are electrically counterbalanced by Mg2+, K+ or Cl- fluxes, but only recently the first transport systems that regulate the pH gradient were identified. Notably several (Na+,K+/H+ antiporter systems where identified, that play a role in pH gradient regulation, ion homeostasis, osmoregulation, or coupling of secondary active transport. The established pH gradients are important to drive uptake of essential ions and solutes, but not many transporters involved have been identified to date. In this mini review we summarize the current status in the field and the open questions that need to be addressed in order to understand how pH gradients are maintained, how this is interconnected with other transport processes and what this means for chloroplast function.

  16. Chloroplast osmotic adjustment allows for acclimation of photosynthesis to low water potentials

    International Nuclear Information System (INIS)

    Gupta, A.S.; Berkowitz, G.

    1987-01-01

    Previously in this laboratory, studies indicated that photosynthesis (PS) of chloroplasts isolated from spinach plants which underwent osmotic adjustment during in situ water deficits was inhibited less at low osmotic potentials (Psi/sub s/) in vitro than PS of plastids isolated from well watered plants. In this study, an attempt was made to determine if chloroplast acclimation to low Psi/sub s/ was associated with in situ stromal solute accumulation. During a 14d stress cycle, in situ stromal volume was estimated by measuring (using the 3 H 2 O, 14 C-sorbitol silicon oil centrifugation technique) the stromal space of plastids in solutions which had the Psi/sub s/ adjusted to the leaf Psi/sub s/. During the first lid of the cycle, stromal volume did not decline, despite a decrease of over 20% in the leaf RWC. After this time, stromal volume dropped rapidly. In situ stromal Psi/sub s/ was also estimated during a stress cycle. These studies indicated that stromal Psi/sub s/ was lowered by net solute accumulation. The data presented in this report suggest that chloroplast acclimation to low Psi/sub s/ may involve stromal solute accumulation and volume maintenance during cell water loss

  17. Restriction endonuclease analysis of chloroplast DNA in interspecies somatic Hybrids of Petunia.

    Science.gov (United States)

    Kumar, A; Cocking, E C; Bovenberg, W A; Kool, A J

    1982-12-01

    Restriction endonuclease cleavage pattern analysis of chloroplast DNA (cpDNA) of three different interspecific somatic hybrid plants revealed that the cytoplasms of the hybrids contained only cpDNA of P. parodii. The somatic hybrid plants analysed were those between P. parodii (wild type) + P. hybrida (wild type); P. parodii (wild type)+P. inflata (cytoplasmic albino mutant); P. parodii (wild type) + P. parviflora (nuclear albino mutant). The presence of only P. parodii chloroplasts in the somatic hybrid of P. parodii + P. inflata is possibly due to the stringent selection used for somatic hybrid production. However, in the case of the two other somatic hybrids P. parodii + P. hybrida and P. parodii + P. parviflora it was not possible to determine whether the presence of only P. parodii chloroplasts in these somatic hybrid plants was due to the nature of the selection schemes used or simply occurred by chance. The relevance of such somatic hybrid material for the study of genomic-cytoplasmic interaction is discussed, as well as the use of restriction endonuclease fragment patterns for the analysis of taxonomic and evolutionary inter-relationships in the genus Petunia.

  18. Transcriptional regulation and DNA methylation in plastids during transitional conversion of chloroplasts to chromoplasts.

    Science.gov (United States)

    Kobayashi, H; Ngernprasirtsiri, J; Akazawa, T

    1990-01-01

    During transitional conversion of chloroplasts to chromoplasts in ripening tomato (Lycopersicon esculentum) fruits, transcripts for several plastid genes for photosynthesis decreased to undetectable levels. Run-on transcription of plastids indicated that transcriptional regulation operated as a predominant factor. We found that most of the genes in chloroplasts were actively transcribed in vitro by Escherichia coli and soluble plastid RNA polymerases, but some genes in chromoplasts seemed to be silent when assayed by the in vitro systems. The regulatory step, therefore, was ascribed to DNA templates. The analysis of modified base composition revealed the presence of methylated bases in chromoplast DNA, in which 5-methylcytosine was most abundant. The presence of 5-methylcytosine detected by isoschizomeric endonucleases and Southern hybridization was correlated with the undetectable transcription activity of each gene in the run-on assay and in vitro transcription experiments. It is thus concluded that the suppression of transcription mediated by DNA methylation is one of the mechanisms governing gene expression in plastids converting from chloroplasts to chromoplasts. Images Fig. 1 Fig. 2 Fig. 3. Fig. 4. Fig. 5. PMID:2303026

  19. Complete Chloroplast Genome Sequence of Coptis chinensis Franch. and Its Evolutionary History

    Science.gov (United States)

    He, Yang; Deng, Cao; Fan, Gang; Qin, Shishang

    2017-01-01

    The Coptis chinensis Franch. is an important medicinal plant from the Ranunculales. We used next generation sequencing technology to determine the complete chloroplast genome of C. chinensis. This genome is 155,484 bp long with 38.17% GC content. Two 26,758 bp long inverted repeats separated the genome into a typical quadripartite structure. The C. chinensis chloroplast genome consists of 128 gene loci, including eight rRNA gene loci, 28 tRNA gene loci, and 92 protein-coding gene loci. Most of the SSRs in C. chinensis are poly-A/T. The numbers of mononucleotide SSRs in C. chinensis and other Ranunculaceae species are fewer than those in Berberidaceae species, while the number of dinucleotide SSRs is greater than that in the Berberidaceae. C. chinensis diverged from other Ranunculaceae species an estimated 81 million years ago (Mya). The divergence between Ranunculaceae and Berberidaceae was ~111 Mya, while the Ranunculales and Magnoliaceae shared a common ancestor during the Jurassic, ~153 Mya. Position 104 of the C. chinensis ndhG protein was identified as a positively selected site, indicating possible selection for the photosystem-chlororespiration system in C. chinensis. In summary, the complete sequencing and annotation of the C. chinensis chloroplast genome will facilitate future studies on this important medicinal species. PMID:28698879

  20. Complete Chloroplast Genome Sequence of Coptis chinensis Franch. and Its Evolutionary History

    Directory of Open Access Journals (Sweden)

    Yang He

    2017-01-01

    Full Text Available The Coptis chinensis Franch. is an important medicinal plant from the Ranunculales. We used next generation sequencing technology to determine the complete chloroplast genome of C. chinensis. This genome is 155,484 bp long with 38.17% GC content. Two 26,758 bp long inverted repeats separated the genome into a typical quadripartite structure. The C. chinensis chloroplast genome consists of 128 gene loci, including eight rRNA gene loci, 28 tRNA gene loci, and 92 protein-coding gene loci. Most of the SSRs in C. chinensis are poly-A/T. The numbers of mononucleotide SSRs in C. chinensis and other Ranunculaceae species are fewer than those in Berberidaceae species, while the number of dinucleotide SSRs is greater than that in the Berberidaceae. C. chinensis diverged from other Ranunculaceae species an estimated 81 million years ago (Mya. The divergence between Ranunculaceae and Berberidaceae was ~111 Mya, while the Ranunculales and Magnoliaceae shared a common ancestor during the Jurassic, ~153 Mya. Position 104 of the C. chinensis ndhG protein was identified as a positively selected site, indicating possible selection for the photosystem-chlororespiration system in C. chinensis. In summary, the complete sequencing and annotation of the C. chinensis chloroplast genome will facilitate future studies on this important medicinal species.

  1. The complete chloroplast genome sequence of Dodonaea viscosa: comparative and phylogenetic analyses.

    Science.gov (United States)

    Saina, Josphat K; Gichira, Andrew W; Li, Zhi-Zhong; Hu, Guang-Wan; Wang, Qing-Feng; Liao, Kuo

    2018-02-01

    The plant chloroplast (cp) genome is a highly conserved structure which is beneficial for evolution and systematic research. Currently, numerous complete cp genome sequences have been reported due to high throughput sequencing technology. However, there is no complete chloroplast genome of genus Dodonaea that has been reported before. To better understand the molecular basis of Dodonaea viscosa chloroplast, we used Illumina sequencing technology to sequence its complete genome. The whole length of the cp genome is 159,375 base pairs (bp), with a pair of inverted repeats (IRs) of 27,099 bp separated by a large single copy (LSC) 87,204 bp, and small single copy (SSC) 17,972 bp. The annotation analysis revealed a total of 115 unique genes of which 81 were protein coding, 30 tRNA, and four ribosomal RNA genes. Comparative genome analysis with other closely related Sapindaceae members showed conserved gene order in the inverted and single copy regions. Phylogenetic analysis clustered D. viscosa with other species of Sapindaceae with strong bootstrap support. Finally, a total of 249 SSRs were detected. Moreover, a comparison of the synonymous (Ks) and nonsynonymous (Ka) substitution rates in D. viscosa showed very low values. The availability of cp genome reported here provides a valuable genetic resource for comprehensive further studies in genetic variation, taxonomy and phylogenetic evolution of Sapindaceae family. In addition, SSR markers detected will be used in further phylogeographic and population structure studies of the species in this genus.

  2. IRscope: An online program to visualize the junction sites of chloroplast genomes.

    Science.gov (United States)

    Amiryousefi, Ali; Hyvönen, Jaakko; Poczai, Peter

    2018-04-05

    Genome plotting is performed using a wide range of visualizations tools each with emphasis on a different informative dimension of the genome. These tools can provide a deeper insight into the genomic structure of the organism. Here we announce a new visualization tool that is specifically designed for chloroplast genomes. It allows the users to depict the genetic architecture of up to ten chloroplast genomes in the vicinity of the sites connecting the inverted repeats to the short and long single copy regions. The software and its dependent libraries are fully coded in R and the reflected plot is scaled up to realistic size of nucleotide base pairs in the vicinity of the junction sites. We introduce a website for easier use of the program as well as R source code of the software to be used in case of preferences to be changed and integrated into personal pipelines. The input of the program is an annotation GenBank (.gb) file, the accession or GI number of the sequence or a DOGMA output file. The software was tested using over a hundred embryophyte chloroplast genomes and in all cases a reliable output was obtained. Source codes and the online suit available @ https://irscope.shinyapps.io/irapp/ or @ https://github.com/Limpfrog/irscope. ali.amiryousefi@helsinki.fi.

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

    Directory of Open Access Journals (Sweden)

    Catalina Perello

    Full Text Available 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.

  4. Complete chloroplast genome of Trachelium caeruleum: extensiverearrangements are associated with repeats and tRNAs

    Energy Technology Data Exchange (ETDEWEB)

    Haberle, Rosemarie C.; Fourcade, Matthew L.; Boore, Jeffrey L.; Jansen, Robert K.

    2006-01-09

    Chloroplast genome structure, gene order and content arehighly conserved in land plants. We sequenced the complete chloroplastgenome sequence of Trachelium caeruleum (Campanulaceae) a member of anangiosperm family known for highly rearranged chloroplast genomes. Thetotal genome size is 162,321 bp with an IR of 27,273 bp, LSC of 100,113bp and SSC of 7,661 bp. The genome encodes 115 unique genes, with 19duplicated in the IR, a tRNA (trnI-CAU) duplicated once in the LSC and aprotein coding gene (psbJ) duplicated twice, for a total of 137 genes.Four genes (ycf15, rpl23, infA and accD) are truncated and likelynonfunctional; three others (clpP, ycf1 and ycf2) are so highly divergedthat they may now be pseudogenes. The most conspicuous feature of theTrachelium genome is the presence of eighteen internally unrearrangedblocks of genes that have been inverted or relocated within the genome,relative to the typical gene order of most angiosperm chloroplastgenomes. Recombination between repeats or tRNAs has been suggested as twomeans of chloroplast genome rearrangements. We compared the relativenumber of repeats in Trachelium to eight other angiosperm chloroplastgenomes, and evaluated the location of repeats and tRNAs in relation torearrangements. Trachelium has the highest number and largest repeats,which are concentrated near inversion endpoints or other rearrangements.tRNAs occur at many but not all inversion endpoints. There is likely nosingle mechanism responsible for the remarkable number of alterations inthis genome, but both repeats and tRNAs are clearly associated with theserearrangements. Land plant chloroplast genomes are highly conserved instructure, gene order and content. The chloroplast genomes of ferns, thegymnosperm Ginkgo, and most angiosperms are nearly collinear, reflectingthe gene order in lineages that diverged from lycopsids and the ancestralchloroplast gene order over 350 million years ago (Raubeson and Jansen,1992). Although earlier mapping studies

  5. Complete Chloroplast Genome of the Wollemi Pine (Wollemia nobilis): Structure and Evolution.

    Science.gov (United States)

    Yap, Jia-Yee S; Rohner, Thore; Greenfield, Abigail; Van Der Merwe, Marlien; McPherson, Hannah; Glenn, Wendy; Kornfeld, Geoff; Marendy, Elessa; Pan, Annie Y H; Wilton, Alan; Wilkins, Marc R; Rossetto, Maurizio; Delaney, Sven K

    2015-01-01

    The Wollemi pine (Wollemia nobilis) is a rare Southern conifer with striking morphological similarity to fossil pines. A small population of W. nobilis was discovered in 1994 in a remote canyon system in the Wollemi National Park (near Sydney, Australia). This population contains fewer than 100 individuals and is critically endangered. Previous genetic studies of the Wollemi pine have investigated its evolutionary relationship with other pines in the family Araucariaceae, and have suggested that the Wollemi pine genome contains little or no variation. However, these studies were performed prior to the widespread use of genome sequencing, and their conclusions were based on a limited fraction of the Wollemi pine genome. In this study, we address this problem by determining the entire sequence of the W. nobilis chloroplast genome. A detailed analysis of the structure of the genome is presented, and the evolution of the genome is inferred by comparison with the chloroplast sequences of other members of the Araucariaceae and the related family Podocarpaceae. Pairwise alignments of whole genome sequences, and the presence of unique pseudogenes, gene duplications and insertions in W. nobilis and Araucariaceae, indicate that the W. nobilis chloroplast genome is most similar to that of its sister taxon Agathis. However, the W. nobilis genome contains an unusually high number of repetitive sequences, and these could be used in future studies to investigate and conserve any remnant genetic diversity in the Wollemi pine.

  6. [Identification of medicinal plant Dendrobium based on the chloroplast psbK-psbI intergenic spacer].

    Science.gov (United States)

    Yao, Hui; Yang, Pei; Zhou, Hong; Ma, Shuang-jiao; Song, Jing-yuan; Chen, Shi-lin

    2015-06-01

    In this paper, the chloroplast psbK-psbI intergenic spacers of 18 species of Dendrobium and their adulterants were amplified and sequenced, and then the sequence characteristics were analyzed. The sequence lengths of chloroplast psbK-psbI regions of Dendrobium ranged from 474 to 513 bp and the GC contents were 25.4%-27.6%. The variable sites were 71 while the informative sites were 46. The inter-specific genetic distances calculated by Kimura 2-parameter (K2P) of Dendrobium were 0.006 1-0.058 1, with an average of 0.028 4. The K2P genetic distances between Dendrobium species and Bulbophyllum odoratissimum were 0.093 2-0.120 4. The NJ tree showed that the Dendrobium species can be easily differentiated from each other and 6 samples of the inspected Dendrobium species were identified successfully through sequencing the psbK-psbI intergenic spacer. Therefore, the chloroplast psbK-psbI intergenic spacer can be used as a candidate marker to identify Dendrobium species and its adulterants.

  7. Developmental changes in aspartate-family amino acid biosynthesis in pea chloroplasts

    International Nuclear Information System (INIS)

    Mills, W.R.; Cato, L.W.; Stephens, B.W.; Reeves, M.

    1990-01-01

    Isolated chloroplasts are known to synthesize the asp-derived amino acids (ile, hse, lys and thr) from [ 14 C]asp (Mills et al, 1980, Plant Physiol. 65, 1166). Now, we have studied the influence of tissue age on essential amino acid biosynthesis in pea (Pisum sativum) plastids. Chloroplasts from the younger (third and fourth) leaves of 12 day old plants, were 2-3 times more active in synthesizing lys and thr from [ 14 C]asp than those from older (first or second) leaves. We also examined two key pathway enzymes (aspartate kinase and homoserine dehydrogenase); with each enzyme,a activity in younger leaves was about 2 times that in plastids from older tissue. Both lys- and thr-sensitive forms of aspartate kinase are known in plants; in agreement with earlier work, we found that lys-sensitive activity was about 4 times higher in the younger tissues, while the thr-sensitive activity changed little during development (Davies and Miflin, 1977, Plant Sci. Lett. 9, 323). Recently the role of aspartate kinase and homoserine dehydrogenase in controlling asp-family amino acid synthesis has been questioned (Giovanelli et al, 1989, Plant Physiol. 90, 1584); we hope that measurements of amino acid levels in chloroplasts as well as further enzyme studies will help us to better understand the regulation of asp-family amino acid synthesis

  8. 2010 GORDON RESEARCH CONFERENCE ON MITOCHONDRIA & CHLOROPLASTS, LUCCA, ITALY, JULY 11-16, 2010

    Energy Technology Data Exchange (ETDEWEB)

    Alice Barkan

    2010-07-16

    The 2010 GRC on Mitochondria & Chloroplasts will assemble an international group of molecular, structural and cellular biologists, biochemists and geneticists investigating a broad spectrum of fundamental problems related to the biology of these organelles in animal, plant and fungal cells. This field has witnessed an extraordinary expansion in recent years, fueled by the discovery of the role of mitochondria in human disease and ageing, and of the synergy of chloroplasts and mitochondria in energetic output, the identification of novel factors involved in organelle division, movement, signaling and acclimation to changing environmental conditions, and by the powerful tools of organelle proteomics. The 2010 GRC will highlight advances in the elucidation of molecular mechanisms of organelle biogenesis including regulation of genome structure, evolution and expression, organellar protein import, assembly and turnover of respiratory and photosynthetic complexes, bidirectional signaling between organelles and nucleus, organelle morphology and dynamics, and the integration of cellular metabolism. We will also explore progress in mechanisms of disease and ageing/ senescence in animals and plants. The organellar field has forged new fronts toward a global and comprehensive understanding of mitochondrial and chloroplast biology at the molecular level. Many of the molecules under study in model organisms are responsible for human diseases, providing significant impetus for a meeting that encourages interactions between mammalian, fungal and plant organellar biologists.

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

    Science.gov (United States)

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

    2016-05-01

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

  10. Light affects the chloroplast ultrastructure and post-storage photosynthetic performance of watermelon (Citrullus lanatus) plug seedlings.

    Science.gov (United States)

    Duan, Qingqing; Jiang, Wu; Ding, Ming; Lin, Ye; Huang, Danfeng

    2014-01-01

    Watermelon [Citrullus lanatus (Thunb.) Matsum. and Nakai] plug seedlings were stored at 15°C in the light at a photosynthetic photon flux density of 15 µmol·m(-2)·s(-1) or in darkness for 6 days, to evaluate their chloroplast ultrastructure, and associated photosynthetic characteristics. Storage in the dark caused swelling, disordered granal arrangement, and starch grain disappearance in the chloroplasts. In contrast, the chloroplasts stored in the light were relatively normal. As a result, the light-stored seedlings had a significantly higher chlorophyll content, Fv/Fm, and Pn than did dark-stored seedlings. Regardless of whether the seedlings were stored in light or darkness, the Gs and Ls of the seedlings significantly decreased, while the Ci obviously increased when the Pn decreased after 6 days of storage. This result suggests that the decreased Pn is not solely a stomatal effect, as the effects on the chloroplasts contributed to this photosynthetic inhibition. Six days after transplanting, seedlings that were stored in the light or darkness for 2 or 4 days showed complete recovery of chloroplast ultrastructure, chlorophyll content, Fv/Fm, Gs and Pn. When the storage period increased to 6 days, the dark-stored seedlings had a significantly lower Fv/Fm and Pn than the light-stored and control seedlings 6 days after transplanting, which was mainly ascribed to incomplete recovery of chloroplast ultrastructure. Furthermore, the light-stored seedlings exhibited a significantly higher shoot dry weight during storage and a higher percentage dry weight increase after transplanting than the dark-stored seedlings. These effects were enhanced by prolonged storage (4 to 6 days). This study demonstrated that dim light during storage is beneficial for maintaining chloroplast ultrastructure as well as photosynthetic efficiency in watermelon seedlings, thus contributing to the rapid recovery of post-storage photosynthetic performance, which ensures the transplant quality

  11. Arabidopsis EMB1990 Encoding a Plastid-Targeted YlmG Protein Is Required for Chloroplast Biogenesis and Embryo Development

    Directory of Open Access Journals (Sweden)

    Hongyu Chen

    2018-02-01

    Full Text Available In higher plants, embryo development originated from fertilized egg cell is the first step of the life cycle. The chloroplast participates in many essential metabolic pathways, and its function is highly associated with embryo development. However, the mechanisms and relevant genetic components by which the chloroplast functions in embryogenesis are largely uncharacterized. In this paper, we describe the Arabidopsis EMB1990 gene, encoding a plastid-targeted YlmG protein which is required for chloroplast biogenesis and embryo development. Loss of the EMB1990/YLMG1-1 resulted in albino seeds containing abortive embryos, and the morphological development of homozygous emb1990 embryos was disrupted after the globular stage. Our results showed that EMB1990/YLMG1-1 was expressed in the primordia and adaxial region of cotyledon during embryogenesis, and the encoded protein was targeted to the chloroplast. TEM observation of cellular ultrastructure showed that chloroplast biogenesis was impaired in emb1990 embryo cells. Expression of certain plastid genes was also affected in the loss-of-function mutants, including genes encoding core protein complex subunits located in the thylakoid membrane. Moreover, the tissue-specific genes of embryo development were misexpressed in emb1990 mutant, including genes known to delineate cell fate decisions in the SAM (shoot apical meristem, cotyledon and hypophysis. Taken together, we propose that the nuclear-encoded YLMG1-1 is targeted to the chloroplast and required for normal plastid gene expression. Hence, YLMG1-1 plays a critical role in Arabidopsis embryogenesis through participating in chloroplast biogenesis.

  12. Complete chloroplast genome sequence of MD-2 pineapple and its comparative analysis among nine other plants from the subclass Commelinidae.

    Science.gov (United States)

    Redwan, R M; Saidin, A; Kumar, S V

    2015-08-12

    Pineapple (Ananas comosus var. comosus) is known as the king of fruits for its crown and is the third most important tropical fruit after banana and citrus. The plant, which is indigenous to South America, is the most important species in the Bromeliaceae family and is largely traded for fresh fruit consumption. Here, we report the complete chloroplast sequence of the MD-2 pineapple that was sequenced using the PacBio sequencing technology. In this study, the high error rate of PacBio long sequence reads of A. comosus's total genomic DNA were improved by leveraging on the high accuracy but short Illumina reads for error-correction via the latest error correction module from Novocraft. Error corrected long PacBio reads were assembled by using a single tool to produce a contig representing the pineapple chloroplast genome. The genome of 159,636 bp in length is featured with the conserved quadripartite structure of chloroplast containing a large single copy region (LSC) with a size of 87,482 bp, a small single copy region (SSC) with a size of 18,622 bp and two inverted repeat regions (IRA and IRB) each with the size of 26,766 bp. Overall, the genome contained 117 unique coding regions and 30 were repeated in the IR region with its genes contents, structure and arrangement similar to its sister taxon, Typha latifolia. A total of 35 repeats structure were detected in both the coding and non-coding regions with a majority being tandem repeats. In addition, 205 SSRs were detected in the genome with six protein-coding genes contained more than two SSRs. Comparative chloroplast genomes from the subclass Commelinidae revealed a conservative protein coding gene albeit located in a highly divergence region. Analysis of selection pressure on protein-coding genes using Ka/Ks ratio showed significant positive selection exerted on the rps7 gene of the pineapple chloroplast with P less than 0.05. Phylogenetic analysis confirmed the recent taxonomical relation among the member of

  13. Reduced starch granule number per chloroplast in the dpe2/phs1 mutant is dependent on initiation of starch degradation.

    Science.gov (United States)

    Malinova, Irina; Fettke, Joerg

    2017-01-01

    An Arabidopsis double knock-out mutant lacking cytosolic disproportionating enzyme 2 (DPE2) and the plastidial phosphorylase (PHS1) revealed a dwarf-growth phenotype, reduced starch content, an uneven distribution of starch within the plant rosette, and a reduced number of starch granules per chloroplast under standard growth conditions. In contrast, the wild type contained 5-7 starch granules per chloroplast. Mature and old leaves of the double mutant were essentially starch free and showed plastidial disintegration. Several analyses revealed that the number of starch granules per chloroplast was affected by the dark phase. So far, it was unclear if it was the dark phase per se or starch degradation in the dark that was connected to the observed decrease in the number of starch granules per chloroplast. Therefore, in the background of the double mutant dpe2/phs1, a triple mutant was generated lacking the initial starch degrading enzyme glucan, water dikinase (GWD). The triple mutant showed improved plant growth, a starch-excess phenotype, and a homogeneous starch distribution. Furthermore, the number of starch granules per chloroplast was increased and was similar to wild type. However, starch granule morphology was only slightly affected by the lack of GWD as in the triple mutant and, like in dpe2/phs1, more spherical starch granules were observed. The characterized triple mutant was discussed in the context of the generation of starch granules and the formation of starch granule morphology.

  14. The Arabidopsis ppi1 Mutant Is Specifically Defective in the Expression, Chloroplast Import, and Accumulation of Photosynthetic ProteinsW⃞

    Science.gov (United States)

    Kubis, Sybille; Baldwin, Amy; Patel, Ramesh; Razzaq, Azam; Dupree, Paul; Lilley, Kathryn; Kurth, Joachim; Leister, Dario; Jarvis, Paul

    2003-01-01

    The import of nucleus-encoded proteins into chloroplasts is mediated by translocon complexes in the envelope membranes. A component of the translocon in the outer envelope membrane, Toc34, is encoded in Arabidopsis by two homologous genes, atTOC33 and atTOC34. Whereas atTOC34 displays relatively uniform expression throughout development, atTOC33 is strongly upregulated in rapidly growing, photosynthetic tissues. To understand the reason for the existence of these two related genes, we characterized the atTOC33 knockout mutant ppi1. Immunoblotting and proteomics revealed that components of the photosynthetic apparatus are deficient in ppi1 chloroplasts and that nonphotosynthetic chloroplast proteins are unchanged or enriched slightly. Furthermore, DNA array analysis of 3292 transcripts revealed that photosynthetic genes are moderately, but specifically, downregulated in ppi1. Proteome differences in ppi1 could be correlated with protein import rates: ppi1 chloroplasts imported the ribulose-1,5-bisphosphate carboxylase/oxygenase small subunit and 33-kD oxygen-evolving complex precursors at significantly reduced rates, but the import of a 50S ribosomal subunit precursor was largely unaffected. The ppi1 import defect occurred at the level of preprotein binding, which is consistent with a role for atToc33 during preprotein recognition. The data suggest that atToc33 is involved preferentially in the import of photosynthetic proteins and, by extension, that atToc34 is involved in the import of nonphotosynthetic chloroplast proteins. PMID:12897258

  15. Chloroplast digestion and the development of functional kleptoplasty in juvenile Elysia timida (Risso, 1818 as compared to short-term and non-chloroplast-retaining sacoglossan slugs.

    Directory of Open Access Journals (Sweden)

    Elise Marie Jerschabek Laetz

    Full Text Available Sacoglossan sea slugs are the only metazoans known to perform functional kleptoplasty, the sequestration and retention of functional chloroplasts within their digestive gland cells. Remarkably, a few species with this ability can survive starvation periods of 3-12 months likely due to their stolen chloroplasts. There are no reports of kleptoplast transfer from mother slug to either eggs or juveniles, demonstrating that each animal must independently acquire its kleptoplasts and develop the ability to maintain them within its digestive gland. We present here an investigation into the development of functional kleptoplasty in a long-term kleptoplast retaining species, Elysia timida. Laboratory-reared juvenile slugs of different post-metamorphic ages were placed in starvation and compared to 5 known short-term retaining slug species and 5 non-retaining slug species. The subsequent results indicate that functional kleptoplasty is not performed by E. timida until after 15 days post-metamorphosis and that by 25 days, these animals outlive many of the short-term retention species. Digestive activity was also monitored using lysosomal abundance as an indicator, revealing different patterns in starving juveniles versus adults. Starved juveniles were reintroduced to food to determine any differences in digestive activity when starvation ends, resulting in an increase in the number of kleptoplasts, but no overall change in lysosomal activity. By revealing some of the changes that occur during early development in these animals, which begin as non-kleptoplast-retaining and grow into long-term retaining slugs, this investigation provides a basis for future inquiries into the origin and development of this remarkable ability.

  16. The complete chloroplast genome sequence of Pelargonium xhortorum: Or ganization and evolution of the largest and most highlyrearranged chloroplast genome of land plants

    Energy Technology Data Exchange (ETDEWEB)

    Chumley, Timothy W.; Palmer, Jeffrey D.; Mower, Jeffrey P.; Fourcade, H. Matthew; Calie, Patrick J.; Boore, Jeffrey L.; Jansen,Robert K.

    2006-01-20

    The chloroplast genome of Pelargonium e hortorum has beencompletely sequenced. It maps as a circular molecule of 217,942 bp, andis both the largest and most rearranged land plant chloroplast genome yetsequenced. It features two copies of a greatly expanded inverted repeat(IR) of 75,741 bp each, and consequently diminished single copy regionsof 59,710 bp and 6,750 bp. It also contains two different associations ofrepeated elements that contribute about 10 percent to the overall sizeand account for the majority of repeats found in the genome. Theyrepresent hotspots for rearrangements and gene duplications and include alarge number of pseudogenes. We propose simple models that account forthe major rearrangements with a minimum of eight IR boundary changes and12 inversions in addition to a several insertions of duplicated sequence.The major processes at work (duplication, IR expansion, and inversion)have disrupted at least one and possibly two or three transcriptionaloperons, and the genes involved in these disruptions form the core of thetwo major repeat associations. Despite the vast increase in size andcomplexity of the genome, the gene content is similar to that of otherangiosperms, with the exceptions of a large number of pseudogenes as partof the repeat associations, the recognition of two open reading frames(ORF56 and ORF42) in the trnA intron with similarities to previouslyidentified mitochondrial products (ACRS and pvs-trnA), the loss of accDand trnT-GGU, and in particular, the lack of a recognizably functionalrpoA. One or all of three similar open reading frames may possibly encodethe latter, however.

  17. Insights into Alternanthera mosaic virus TGB3 functions: Interactions with Nicotiana benthamiana PsbO correlate with chloroplast vesiculation and veinal necrosis caused by TGB3 overexpression

    Science.gov (United States)

    Alternanthera mosaic virus (AltMV) triple gene block 3 (TGB3) protein is involved in viral movement. AltMV TGB3 subcellular localization was previously shown to be distinct from that of Potato virus X (PVX) TGB3, and a chloroplast binding domain identified; veinal necrosis and chloroplast vesiculati...

  18. A nuclear-encoded chloroplast protein harboring a single CRM domain plays an important role in the Arabidopsis growth and stress response.

    Science.gov (United States)

    Lee, Kwanuk; Lee, Hwa Jung; Kim, Dong Hyun; Jeon, Young; Pai, Hyun-Sook; Kang, Hunseung

    2014-04-16

    Although several chloroplast RNA splicing and ribosome maturation (CRM) domain-containing proteins have been characterized for intron splicing and rRNA processing during chloroplast gene expression, the functional role of a majority of CRM domain proteins in plant growth and development as well as chloroplast RNA metabolism remains largely unknown. Here, we characterized the developmental and stress response roles of a nuclear-encoded chloroplast protein harboring a single CRM domain (At4g39040), designated CFM4, in Arabidopsis thaliana. Analysis of CFM4-GFP fusion proteins revealed that CFM4 is localized to chloroplasts. The loss-of-function T-DNA insertion mutants for CFM4 (cfm4) displayed retarded growth and delayed senescence, suggesting that CFM4 plays a role in growth and development of plants under normal growth conditions. In addition, cfm4 mutants showed retarded seed germination and seedling growth under stress conditions. No alteration in the splicing patterns of intron-containing chloroplast genes was observed in the mutant plants, but the processing of 16S and 4.5S rRNAs was abnormal in the mutant plants. Importantly, CFM4 was determined to possess RNA chaperone activity. These results suggest that the chloroplast-targeted CFM4, one of two Arabidopsis genes encoding a single CRM domain-containing protein, harbors RNA chaperone activity and plays a role in the Arabidopsis growth and stress response by affecting rRNA processing in chloroplasts.

  19. Characterization of Chloroplastic Fructose 1,6-Bisphosphate Aldolases as Lysine-methylated Proteins in Plants*

    Science.gov (United States)

    Mininno, Morgane; Brugière, Sabine; Pautre, Virginie; Gilgen, Annabelle; Ma, Sheng; Ferro, Myriam; Tardif, Marianne; Alban, Claude; Ravanel, Stéphane

    2012-01-01

    In pea (Pisum sativum), the protein-lysine methyltransferase (PsLSMT) catalyzes the trimethylation of Lys-14 in the large subunit (LS) of ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco), the enzyme catalyzing the CO2 fixation step during photosynthesis. Homologs of PsLSMT, herein referred to as LSMT-like enzymes, are found in all plant genomes, but methylation of LS Rubisco is not universal in the plant kingdom, suggesting a species-specific protein substrate specificity of the methyltransferase. In this study, we report the biochemical characterization of the LSMT-like enzyme from Arabidopsis thaliana (AtLSMT-L), with a focus on its substrate specificity. We show that, in Arabidopsis, LS Rubisco is not naturally methylated and that the physiological substrates of AtLSMT-L are chloroplastic fructose 1,6-bisphosphate aldolase isoforms. These enzymes, which are involved in the assimilation of CO2 through the Calvin cycle and in chloroplastic glycolysis, are trimethylated at a conserved lysyl residue located close to the C terminus. Both AtLSMT-L and PsLSMT are able to methylate aldolases with similar kinetic parameters and product specificity. Thus, the divergent substrate specificity of LSMT-like enzymes from pea and Arabidopsis concerns only Rubisco. AtLSMT-L is able to interact with unmethylated Rubisco, but the complex is catalytically unproductive. Trimethylation does not modify the kinetic properties and tetrameric organization of aldolases in vitro. The identification of aldolases as methyl proteins in Arabidopsis and other species like pea suggests a role of protein lysine methylation in carbon metabolism in chloroplasts. PMID:22547063

  20. Molecular Evolution of the Substrate Specificity of Chloroplastic Aldolases/Rubisco Lysine Methyltransferases in Plants.

    Science.gov (United States)

    Ma, Sheng; Martin-Laffon, Jacqueline; Mininno, Morgane; Gigarel, Océane; Brugière, Sabine; Bastien, Olivier; Tardif, Marianne; Ravanel, Stéphane; Alban, Claude

    2016-04-04

    Rubisco and fructose-1,6-bisphosphate aldolases (FBAs) are involved in CO2 fixation in chloroplasts. Both enzymes are trimethylated at a specific lysine residue by the chloroplastic protein methyltransferase LSMT. Genes coding LSMT are present in all plant genomes but the methylation status of the substrates varies in a species-specific manner. For example, chloroplastic FBAs are naturally trimethylated in both Pisum sativum and Arabidopsis thaliana, whereas the Rubisco large subunit is trimethylated only in the former species. The in vivo methylation status of aldolases and Rubisco matches the catalytic properties of AtLSMT and PsLSMT, which are able to trimethylate FBAs or FBAs and Rubisco, respectively. Here, we created chimera and site-directed mutants of monofunctional AtLSMT and bifunctional PsLSMT to identify the molecular determinants responsible for substrate specificity. Our results indicate that the His-Ala/Pro-Trp triad located in the central part of LSMT enzymes is the key motif to confer the capacity to trimethylate Rubisco. Two of the critical residues are located on a surface loop outside the methyltransferase catalytic site. We observed a strict correlation between the presence of the triad motif and the in vivo methylation status of Rubisco. The distribution of the motif into a phylogenetic tree further suggests that the ancestral function of LSMT was FBA trimethylation. In a recent event during higher plant evolution, this function evolved in ancestors of Fabaceae, Cucurbitaceae, and Rosaceae to include Rubisco as an additional substrate to the archetypal enzyme. Our study provides insight into mechanisms by which SET-domain protein methyltransferases evolve new substrate specificity. Copyright © 2016 The Author. Published by Elsevier Inc. All rights reserved.

  1. Dynamic Evolution of the Chloroplast Genome in the Green Algal Classes Pedinophyceae and Trebouxiophyceae.

    Science.gov (United States)

    Turmel, Monique; Otis, Christian; Lemieux, Claude

    2015-07-01

    Previous studies of trebouxiophycean chloroplast genomes revealed little information regarding the evolutionary dynamics of this genome because taxon sampling was too sparse and the relationships between the sampled taxa were unknown. We recently sequenced the chloroplast genomes of 27 trebouxiophycean and 2 pedinophycean green algae to resolve the relationships among the main lineages recognized for the Trebouxiophyceae. These taxa and the previously sampled members of the Pedinophyceae and Trebouxiophyceae are included in the comparative chloroplast genome analysis we report here. The 38 genomes examined display considerable variability at all levels, except gene content. Our results highlight the high propensity of the rDNA-containing large inverted repeat (IR) to vary in size, gene content and gene order as well as the repeated losses it experienced during trebouxiophycean evolution. Of the seven predicted IR losses, one event demarcates a superclade of 11 taxa representing 5 late-diverging lineages. IR expansions/contractions account not only for changes in gene content in this region but also for changes in gene order and gene duplications. Inversions also led to gene rearrangements within the IR, including the reversal or disruption of the rDNA operon in some lineages. Most of the 20 IR-less genomes are more rearranged compared with their IR-containing homologs and tend to show an accelerated rate of sequence evolution. In the IR-less superclade, several ancestral operons were disrupted, a few genes were fragmented, and a subgroup of taxa features a G+C-biased nucleotide composition. Our analyses also unveiled putative cases of gene acquisitions through horizontal transfer. © The Author(s) 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  2. Molecular characterization and expression analysis of chloroplast protein import components in tomato (Solanum lycopersicum.

    Directory of Open Access Journals (Sweden)

    Jianmin Yan

    Full Text Available The translocon at the outer envelope membrane of chloroplasts (Toc mediates the recognition and initial import into the organelle of thousands of nucleus-encoded proteins. These proteins are translated in the cytosol as precursor proteins with cleavable amino-terminal targeting sequences called transit peptides. The majority of the known Toc components that mediate chloroplast protein import were originally identified in pea, and more recently have been studied most extensively in Arabidopsis. With the completion of the tomato genome sequencing project, it is now possible to identify putative homologues of the chloroplast import components in tomato. In the work reported here, the Toc GTPase cDNAs from tomato were identified, cloned and analyzed. The analysis revealed that there are four Toc159 homologues (slToc159-1, -2, -3 and -4 and two Toc34 homologues (slToc34-1 and -2 in tomato, and it was shown that tomato Toc159 and Toc34 homologues share high sequence similarity with the comparable import apparatus components from Arabidopsis and pea. Thus, tomato is a valid model for further study of this system. The expression level of Toc complex components was also investigated in different tissues during tomato development. The two tomato Toc34 homologues are expressed at higher levels in non-photosynthetic tissues, whereas, the expression of two tomato Toc159 homologues, slToc159-1 and slToc159-4, were higher in photosynthetic tissues, and the expression patterns of slToc159-2 was not significantly different in photosynthetic and non-photosynthetic tissues, and slToc159-3 expression was limited to a few select tissues.

  3. Physiology of pepper fruit and the metabolism of antioxidants: chloroplasts, mitochondria and peroxisomes

    Science.gov (United States)

    Palma, José M.; Sevilla, Francisca; Jiménez, Ana; del Río, Luis A.; Corpas, Francisco J.; Álvarez de Morales, Paz; Camejo, Daymi M.

    2015-01-01

    Background and Aims Pepper (Capsicum annuum) contains high levels of antioxidants, such as vitamins A and C and flavonoids. However, information on the role of these beneficial compounds in the physiology of pepper fruit remains scarce. Recent studies have shown that antioxidants in ripe pepper fruit play a key role in responses to temperature changes, and the redox state at the time of harvest affects the nutritional value for human consumption. In this paper, the role of antioxidant metabolism of pepper fruit during ripening and in the response to low temperature is addressed, paying particular attention to ascorbate, NADPH and the superoxide dismutase enzymatic system. The participation of chloroplasts, mitochondria and peroxisomes in the ripening process is also investigated. Scope and Results Important changes occur at a subcellular level during ripening of pepper fruit. Chloroplasts turn into chromoplasts, with drastic conversion of their metabolism, and the role of the ascorbate–glutathione cycle is essential. In mitochondria from red fruits, higher ascorbate peroxidase (APX) and Mn-SOD activities are involved in avoiding the accumulation of reactive oxygen species in these organelles during ripening. Peroxisomes, whose antioxidant capacity at fruit ripening is substantially affected, display an atypical metabolic pattern during this physiological stage. In spite of these differences observed in the antioxidative metabolism of mitochondria and peroxisomes, proteomic analysis of these organelles, carried out by 2-D electrophoresis and MALDI-TOF/TOF and provided here for the first time, reveals no changes between the antioxidant metabolism from immature (green) and ripe (red) fruits. Conclusions Taken together, the results show that investigation of molecular and enzymatic antioxidants from cell compartments, especially chloroplasts, mitochondria and peroxisomes, is a useful tool to study the physiology of pepper fruit, particularly in the context of

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

    Science.gov (United States)

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

    2003-06-01

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

  5. Selectable tolerance to herbicides by mutated acetolactate synthase genes integrated into the chloroplast genome of tobacco.

    Science.gov (United States)

    Shimizu, Masanori; Goto, Maki; Hanai, Moeko; Shimizu, Tsutomu; Izawa, Norihiko; Kanamoto, Hirosuke; Tomizawa, Ken-Ichi; Yokota, Akiho; Kobayashi, Hirokazu

    2008-08-01

    Strategies employed for the production of genetically modified (GM) crops are premised on (1) the avoidance of gene transfer in the field; (2) the use of genes derived from edible organisms such as plants; (3) preventing the appearance of herbicide-resistant weeds; and (4) maintaining transgenes without obstructing plant cell propagation. To this end, we developed a novel vector system for chloroplast transformation with acetolactate synthase (ALS). ALS catalyzes the first step in the biosynthesis of the branched amino acids, and its enzymatic activity is inhibited by certain classes of herbicides. We generated a series of Arabidopsis (Arabidopsis thaliana) mutated ALS (mALS) genes and introduced constructs with mALS and the aminoglycoside 3'-adenyltransferase gene (aadA) into the tobacco (Nicotiana tabacum) chloroplast genome by particle bombardment. Transplastomic plants were selected using their resistance to spectinomycin. The effects of herbicides on transplastomic mALS activity were examined by a colorimetric assay using the leaves of transplastomic plants. We found that transplastomic G121A, A122V, and P197S plants were specifically tolerant to pyrimidinylcarboxylate, imidazolinon, and sulfonylurea/pyrimidinylcarboxylate herbicides, respectively. Transplastomic plants possessing mALSs were able to grow in the presence of various herbicides, thus affirming the relationship between mALSs and the associated resistance to herbicides. Our results show that mALS genes integrated into the chloroplast genome are useful sustainable markers that function to exclude plants other than those that are GM while maintaining transplastomic crops. This investigation suggests that the resistance management of weeds in the field amid growing GM crops is possible using (1) a series of mALSs that confer specific resistance to herbicides and (2) a strategy that employs herbicide rotation.

  6. Mitochondrial DNA, chloroplast DNA and the origins of development in eukaryotic organisms

    Directory of Open Access Journals (Sweden)

    Bendich Arnold J

    2010-06-01

    Full Text Available Abstract Background Several proposals have been made to explain the rise of multicellular life forms. An internal environment can be created and controlled, germ cells can be protected in novel structures, and increased organismal size allows a "division of labor" among cell types. These proposals describe advantages of multicellular versus unicellular organisms at levels of organization at or above the individual cell. I focus on a subsequent phase of evolution, when multicellular organisms initiated the process of development that later became the more complex embryonic development found in animals and plants. The advantage here is realized at the level of the mitochondrion and chloroplast. Hypothesis The extreme instability of DNA in mitochondria and chloroplasts has not been widely appreciated even though it was first reported four decades ago. Here, I show that the evolutionary success of multicellular animals and plants can be traced to the protection of organellar DNA. Three stages are envisioned. Sequestration allowed mitochondria and chloroplasts to be placed in "quiet" germ line cells so that their DNA is not exposed to the oxidative stress produced by these organelles in "active" somatic cells. This advantage then provided Opportunity, a period of time during which novel processes arose for signaling within and between cells and (in animals for cell-cell recognition molecules to evolve. Development then led to the enormous diversity of animals and plants. Implications The potency of a somatic stem cell is its potential to generate cell types other than itself, and this is a systems property. One of the biochemical properties required for stemness to emerge from a population of cells might be the metabolic quiescence that protects organellar DNA from oxidative stress. Reviewers This article was reviewed by John Logsdon, Arcady Mushegian, and Patrick Forterre.

  7. [In vivo and in vitro actions of biscarbamates on the photosynthetic activity of chloroplasts].

    Science.gov (United States)

    Chueca, A; Barón, M; López-Gorgé, J

    1982-01-01

    The "photosynthetic inhibition" component in the whole context of plant toxicity, when different concentrations of the bis-carbamate phenmedipham are supplied through the roots or foliar application to spinach plants grown in hydroponic media have been determined. Chloroplasts were isolated after eight days of the herbicide addition, and then determined: electron transport H2O leads to NADP+, H2O leads to ferrycyanide and ascorbate/DPIP leads to NADP+, cyclic and non cyclic photophosphorilation, CO2 assimilation rate and intermediate patterns of CO2 fixation. We have also determined in foliar disks the O2 evolving and the CO2 assimilation capabilities. Type A and type B chloroplasts showed increased inhibition, respectively, of the Phot. II dependent electron transport chains H2O leads to NADP+ and H2O leads to ferricyanide, to the extent that the phenmedipham concentration increased in the hydroponic media and the spraying solution, so that a 50% inhibition of both processes was obtained at 100 microM and 10 microM, respectively, against 0.2 microM in the in vitro experiments. Non cyclic photophosphorylation showed a stronger inhibition than the cyclic one. Concerning the Phot. I dependent electron transport ascorbate/DPIP leads to NADP+, the chloroplast preparations showed a negligible inhibition. We have found a synergistic effect of the above two factors on the CO2 assimilation. The intermediate patterns of CO2 assimilation showed a decrease of the 3C-compounds P-glycerate and trioses-P, with a parallel increase of the sugar mono and diphosphates as well as disaccharides and amino acids.

  8. Salinity inhibits post transcriptional processing of chloroplast 16S rRNA in shoot cultures of jojoba (Simmondsia chinesis).

    Science.gov (United States)

    Mizrahi-Aviv, Ela; Mills, David; Benzioni, Aliza; Bar-Zvi, Dudy

    2005-03-01

    Chloroplast metabolism is rapidly affected by salt stress. Photosynthesis is one of the first processes known to be affected by salinity. Here, we report that salinity inhibits chloroplast post-transcriptional RNA processing. A differentially expressed 680-bp cDNA, containing the 3' sequence of 16S rRNA, transcribed intergenic spacer, exon 1 and intron of tRNA(Ile), was isolated by differential display reverse transcriptase PCR from salt-grown jojoba (Simmondsia chinesis) shoot cultures. Northern blot analysis indicated that although most rRNA appears to be fully processed, partially processed chloroplast 16S rRNA accumulates in salt-grown cultures. Thus, salinity appears to decrease the processing of the rrn transcript. The possible effect of this decreased processing on physiological processes is, as yet, unknown.

  9. Arabidopsis thaliana AMY3 Is a Unique Redox-regulated Chloroplastic α-Amylase

    DEFF Research Database (Denmark)

    Seung, David; Thalmann, Matthias; Sparla, Francesca

    2013-01-01

    α-Amylases are glucan hydrolases that cleave α-1,4-glucosidic bonds in starch. In vascular plants, α-amylases can be classified into three subfamilies. Arabidopsis has one member of each subfamily. Among them, only AtAMY3 is localized in the chloroplast. We expressed and purified AtAMY3 from...... to be dependent on a conserved aspartic acid residue (Asp666), identified as the catalytic nucleophile in other plant α-amylases such as the barley AMY1. AtAMY3 released small linear and branched glucans from Arabidopsis starch granules, and the proportion of branched glucans increased after the predigestion...

  10. Strong Accumulation of Chloroplast DNA in the Y Chromosomes of Rumex acetosa and Silene latifolia

    Czech Academy of Sciences Publication Activity Database

    Šteflová, Pavlína; Hobza, Roman; Vyskot, Boris; Kejnovský, Eduard

    2014-01-01

    Roč. 142, č. 1 (2014), s. 59-65 ISSN 1424-8581 R&D Projects: GA ČR(CZ) GAP305/10/0930; GA ČR(CZ) GAP501/10/0102; GA ČR(CZ) GBP501/12/G090; GA ČR GAP501/12/2220; GA ČR(CZ) GA522/09/0083; GA MŠk(CZ) LO1204 Institutional research plan: CEZ:AV0Z50040702 Institutional support: RVO:68081707 Keywords : Chloroplast DNA * Rumex acetosa * Sex chromosomes Subject RIV: BO - Biophysics Impact factor: 1.561, year: 2014

  11. The chloroplasts membrane phospholipids of Chlamydomonas reinhardii mutant not forming the Photosystem 2

    International Nuclear Information System (INIS)

    Trusova, V.M.; Ladygin, V.G.; Mezentsev, V.V.; Molchanov, M.I.

    1987-01-01

    Study on a component composition and physical state of photosynthetic membranes of Chlamydomonas chloroplasts of the wild type and mutant A-110 with disturbance of electron transfer chain in the photosystem 2 region permitted to conclude that 170 A diameter particles localized on the internal hydrophobic surface of membrane chips are deleted with respect to phosphatidylglycerin. The results obtained permit to suggest that the formation of protein-lipid complexes containing phosphatidylglycerins is suppressed in mutant A-110 which is not capable of the lamellar system differentation in

  12. Origins of the plant chloroplasts and mitochondria based on comparisons of 5S ribosomal RNAs

    Science.gov (United States)

    Delihas, N.; Fox, G. E.

    1987-01-01

    In this paper, we provide macromolecular comparisons utilizing the 5S ribosomal RNA structure to suggest extant bacteria that are the likely descendants of chloroplast and mitochondria endosymbionts. The genetic stability and near universality of the 5S ribosomal gene allows for a useful means to study ancient evolutionary changes by macromolecular comparisons. The value in current and future ribosomal RNA comparisons is in fine tuning the assignment of ancestors to the organelles and in establishing extant species likely to be descendants of bacteria involved in presumed multiple endosymbiotic events.

  13. Consequences of C4 differentiation for chloroplast membrane proteomes in maize mesophyll and bundle sheath cells.

    Science.gov (United States)

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

    2008-09-01

    Chloroplasts of maize leaves differentiate into specific bundle sheath (BS) and mesophyll (M) types to accommodate C(4) 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 C(4) 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 b(6)f, 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/P(i) 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

  14. The architecture of the chloroplast trnH-psbA non-coding region in angiosperms

    Czech Academy of Sciences Publication Activity Database

    Štorchová, Helena; Olson, M.S.

    2007-01-01

    Roč. 268, 1-4 (2007), s. 235-256 ISSN 0378-2697 R&D Projects: GA MŠk(CZ) LC06004 Grant - others:ESPSCor Visiting Scholar Research Grant(US) NSF DEB 0317115 Institutional research plan: CEZ:AV0Z50380511 Source of funding: V - iné verejné zdroje ; V - iné verejné zdroje Keywords : Chloroplast DNA * psbA-trnH intergenic region * Silene * deletions * insertions and inversions in stem-loop region * psbA 3´untranslated region * RNA secondary structure Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.492, year: 2007

  15. The complete chloroplast genome sequence of an endemic monotypic genus Hagenia (Rosaceae: structural comparative analysis, gene content and microsatellite detection

    Directory of Open Access Journals (Sweden)

    Andrew W. Gichira

    2017-01-01

    Full Text Available Hagenia is an endangered monotypic genus endemic to the topical mountains of Africa. The only species, Hagenia abyssinica (Bruce J.F. Gmel, is an important medicinal plant producing bioactive compounds that have been traditionally used by African communities as a remedy for gastrointestinal ailments in both humans and animals. Complete chloroplast genomes have been applied in resolving phylogenetic relationships within plant families. We employed high-throughput sequencing technologies to determine the complete chloroplast genome sequence of H. abyssinica. The genome is a circular molecule of 154,961 base pairs (bp, with a pair of Inverted Repeats (IR 25,971 bp each, separated by two single copies; a large (LSC, 84,320 bp and a small single copy (SSC, 18,696. H. abyssinica’s chloroplast genome has a 37.1% GC content and encodes 112 unique genes, 78 of which code for proteins, 30 are tRNA genes and four are rRNA genes. A comparative analysis with twenty other species, sequenced to-date from the family Rosaceae, revealed similarities in structural organization, gene content and arrangement. The observed size differences are attributed to the contraction/expansion of the inverted repeats. The translational initiation factor gene (infA which had been previously reported in other chloroplast genomes was conspicuously missing in H. abyssinica. A total of 172 microsatellites and 49 large repeat sequences were detected in the chloroplast genome. A Maximum Likelihood analyses of 71 protein-coding genes placed Hagenia in Rosoideae. The availability of a complete chloroplast genome, the first in the Sanguisorbeae tribe, is beneficial for further molecular studies on taxonomic and phylogenomic resolution within the Rosaceae family.

  16. The complete chloroplast genome sequence of an endemic monotypic genus Hagenia (Rosaceae): structural comparative analysis, gene content and microsatellite detection.

    Science.gov (United States)

    Gichira, Andrew W; Li, Zhizhong; Saina, Josphat K; Long, Zhicheng; Hu, Guangwan; Gituru, Robert W; Wang, Qingfeng; Chen, Jinming

    2017-01-01

    Hagenia is an endangered monotypic genus endemic to the topical mountains of Africa. The only species, Hagenia abyssinica (Bruce) J.F. Gmel, is an important medicinal plant producing bioactive compounds that have been traditionally used by African communities as a remedy for gastrointestinal ailments in both humans and animals. Complete chloroplast genomes have been applied in resolving phylogenetic relationships within plant families. We employed high-throughput sequencing technologies to determine the complete chloroplast genome sequence of H. abyssinica. The genome is a circular molecule of 154,961 base pairs (bp), with a pair of Inverted Repeats (IR) 25,971 bp each, separated by two single copies; a large (LSC, 84,320 bp) and a small single copy (SSC, 18,696). H. abyssinica 's chloroplast genome has a 37.1% GC content and encodes 112 unique genes, 78 of which code for proteins, 30 are tRNA genes and four are rRNA genes. A comparative analysis with twenty other species, sequenced to-date from the family Rosaceae, revealed similarities in structural organization, gene content and arrangement. The observed size differences are attributed to the contraction/expansion of the inverted repeats. The translational initiation factor gene ( infA ) which had been previously reported in other chloroplast genomes was conspicuously missing in H. abyssinica . A total of 172 microsatellites and 49 large repeat sequences were detected in the chloroplast genome. A Maximum Likelihood analyses of 71 protein-coding genes placed Hagenia in Rosoideae. The availability of a complete chloroplast genome, the first in the Sanguisorbeae tribe, is beneficial for further molecular studies on taxonomic and phylogenomic resolution within the Rosaceae family.

  17. Importance of the green color, absorption gradient, and spectral absorption of chloroplasts for the radiative energy balance of leaves.

    Science.gov (United States)

    Kume, Atsushi

    2017-05-01

    Terrestrial green plants absorb photosynthetically active radiation (PAR; 400-700 nm) but do not absorb photons evenly across the PAR waveband. The spectral absorbance of photosystems and chloroplasts is lowest for green light, which occurs within the highest irradiance waveband of direct solar radiation. We demonstrate a close relationship between this phenomenon and the safe and efficient utilization of direct solar radiation in simple biophysiological models. The effects of spectral absorptance on the photon and irradiance absorption processes are evaluated using the spectra of direct and diffuse solar radiation. The radiation absorption of a leaf arises as a consequence of the absorption of chloroplasts. The photon absorption of chloroplasts is strongly dependent on the distribution of pigment concentrations and their absorbance spectra. While chloroplast movements in response to light are important mechanisms controlling PAR absorption, they are not effective for green light because chloroplasts have the lowest spectral absorptance in the waveband. With the development of palisade tissue, the incident photons per total palisade cell surface area and the absorbed photons per chloroplast decrease. The spectral absorbance of carotenoids is effective in eliminating shortwave PAR (solar radiation. However, most of the near infrared radiation is unabsorbed and heat stress is greatly reduced. The incident solar radiation is too strong to be utilized for photosynthesis under the current CO 2 concentration in the terrestrial environment. Therefore, the photon absorption of a whole leaf is efficiently regulated by photosynthetic pigments with low spectral absorptance in the highest irradiance waveband and through a combination of pigment density distribution and leaf anatomical structures.

  18. New chloroplast microsatellite markers suitable for assessing genetic diversity of Lolium perenne and other related grass species.

    Science.gov (United States)

    Diekmann, Kerstin; Hodkinson, Trevor R; Barth, Susanne

    2012-11-01

    Lolium perenne (perennial ryegrass) is the most important forage grass species of temperate regions. We have previously released the chloroplast genome sequence of L. perenne 'Cashel'. Here nine chloroplast microsatellite markers are published, which were designed based on knowledge about genetically variable regions within the L. perenne chloroplast genome. These markers were successfully used for characterizing the genetic diversity in Lolium and different grass species. Chloroplast genomes of 14 Poaceae taxa were screened for mononucleotide microsatellite repeat regions and primers designed for their amplification from nine loci. The potential of these markers to assess genetic diversity was evaluated on a set of 16 Irish and 15 European L. perenne ecotypes, nine L. perenne cultivars, other Lolium taxa and other grass species. All analysed Poaceae chloroplast genomes contained more than 200 mononucleotide repeats (chloroplast simple sequence repeats, cpSSRs) of at least 7 bp in length, concentrated mainly in the large single copy region of the genome. Nucleotide composition varied considerably among subfamilies (with Pooideae biased towards poly A repeats). The nine new markers distinguish L. perenne from all non-Lolium taxa. TeaCpSSR28 was able to distinguish between all Lolium species and Lolium multiflorum due to an elongation of an A(8) mononucleotide repeat in L. multiflorum. TeaCpSSR31 detected a considerable degree of microsatellite length variation and single nucleotide polymorphism. TeaCpSSR27 revealed variation within some L. perenne accessions due to a 44-bp indel and was hence readily detected by simple agarose gel electrophoresis. Smaller insertion/deletion events or single nucleotide polymorphisms detected by these new markers could be visualized by polyacrylamide gel electrophoresis or DNA sequencing, respectively. The new markers are a valuable tool for plant breeding companies, seed testing agencies and the wider scientific community due to

  19. Expression of the recombinant bacterial outer surface protein A in tobacco chloroplasts leads to thylakoid localization and loss of photosynthesis.

    Science.gov (United States)

    Hennig, Anna; Bonfig, Katharina; Roitsch, Thomas; Warzecha, Heribert

    2007-11-01

    Bacterial lipoproteins play crucial roles in host-pathogen interactions and pathogenesis and are important targets for the immune system. A prominent example is the outer surface protein A (OspA) of Borrelia burgdorferi, which has been efficiently used as a vaccine for the prevention of Lyme disease. In a previous study, OspA could be produced in tobacco chloroplasts in a lipidated and immunogenic form. To further explore the potential of chloroplasts for the production of bacterial lipoproteins, the role of the N-terminal leader sequence was investigated. The amount of recombinant OspA could be increased up to ten-fold by the variation of the insertion site in the chloroplast genome. Analysis of OspA mutants revealed that replacement of the invariant cysteine residue as well as deletion of the leader sequence abolishes palmitolyation of OspA. Also, decoration of OspA with an N-terminal eukaryotic lipidation motif does not lead to palmitoylation in chloroplasts. Strikingly, the bacterial signal peptide of OspA efficiently targets the protein to thylakoids, and causes a mutant phenotype. Plants accumulating OspA at 10% total soluble protein could not grow without exogenously supplied sugars and rapidly died after transfer to soil under greenhouse conditions. The plants were found to be strongly affected in photosystem II, as revealed by the analyses of temporal and spatial dynamics of photosynthetic activity by chlorophyll fluorescence imaging. Thus, overexpression of OspA in chloroplasts is limited by its concentration-dependent interference with essential functions of chloroplastic membranes required for primary metabolism.

  20. Expression and characterization of antimicrobial peptides Retrocyclin-101 and Protegrin-1 in chloroplasts to control viral and bacterial infections.

    Science.gov (United States)

    Lee, Seung-Bum; Li, Baichuan; Jin, Shuangxia; Daniell, Henry

    2011-01-01

    Retrocyclin-101 (RC101) and Protegrin-1 (PG1) are two important antimicrobial peptides that can be used as therapeutic agents against bacterial and/or viral infections, especially those caused by the HIV-1 or sexually transmitted bacteria. Because of their antimicrobial activity and complex secondary structures, they have not yet been produced in microbial systems and their chemical synthesis is prohibitively expensive. Therefore, we created chloroplast transformation vectors with the RC101 or PG1 coding sequence, fused with GFP to confer stability, furin or Factor Xa cleavage site to liberate the mature peptide from their fusion proteins and a His-tag to aid in their purification. Stable integration of RC101 into the tobacco chloroplast genome and homoplasmy were confirmed by Southern blots. RC101 and PG1 accumulated up to 32%-38% and 17%∼26% of the total soluble protein. Both RC101 and PG1 were cleaved from GFP by corresponding proteases in vitro, and Factor Xa-like protease activity was observed within chloroplasts. Confocal microscopy studies showed location of GFP fluorescence within chloroplasts. Organic extraction resulted in 10.6-fold higher yield of RC101 than purification by affinity chromatography using His-tag. In planta bioassays with Erwinia carotovora confirmed the antibacterial activity of RC101 and PG1 expressed in chloroplasts. RC101 transplastomic plants were resistant to tobacco mosaic virus infections, confirming antiviral activity. Because RC101 and PG1 have not yet been produced in other cell culture or microbial systems, chloroplasts can be used as bioreactors for producing these proteins. Adequate yield of purified antimicrobial peptides from transplastomic plants should facilitate further preclinical studies. © 2010 The Authors. Plant Biotechnology Journal © 2010 Society for Experimental Biology and Blackwell Publishing Ltd.

  1. Construction of a restriction map and gene map of the lettuce chloroplast small single-copy region using Southern cross-hybridization.

    Science.gov (United States)

    Mitchelson, K R

    1996-01-01

    The small single-copy region (SSCR) of the chloroplast genome of many higher plants typically contain ndh genes encoding proteins that share homology with subunits of the respiratory-chain reduced nicotinamide adenine dinucleotide (NADH) dehydrogenase complex of mitochondria. A map of the lettuce chloroplast SSCR has been determined by Southern cross-hybridization, taking advantage of the high degree of homology between a tobacco small single-copy fragment and a corresponding lettuce chloroplast fragment. The gene order of the SSCR of lettuce and tobacco chloroplasts is similar. The cross-hybridization method can rapidly create a primary gene map of unknown chloroplast fragments, thus providing detailed information of the localization and arrangement of genes and conserved open reading frame regions.

  2. Downregulation of chloroplast-targeted beta-amylase leads to a starch-excess phenotype in leaves

    DEFF Research Database (Denmark)

    Scheidig, A.; Fröhlich, A.; Schulze, S.

    2002-01-01

    showed that the protein product was a functional beta-amylase that could degrade both starch granules and solubilized amylopectin, while import experiments demonstrated that the beta-amylase was imported and processed into pea chloroplasts. To study the function of the protein in transitory starch......A functional screen in Escherichia coli was established to identify potato genes coding for proteins involved in transitory starch degradation. One clone isolated had a sequence very similar to a recently described chloroplast-targeted 5-amylase of Arabidopsis. Expression of the gene in E. coli...

  3. Arabidopsis VARIEGATED 3 encodes a chloroplasttargeted, zinc-finger protein required for chloroplast and palisade cell development

    DEFF Research Database (Denmark)

    Næsted, Henrik; Holm, A.; Jenkins, T.

    2004-01-01

    The stable, recessive Arabidopsis variegated 3 (var3) mutant exhibits a variegated phenotype due to somatic areas lacking or containing developmentally retarded chloroplasts and greatly reduced numbers of palisade cells. The VAR3 gene, isolated by transposon tagging, encodes the 85.9 kDa VAR3...... that pigment profiles are qualitatively similar in wild type and var3, although var3 accumulates lower levels of chlorophylls and carotenoids. These results indicate that VAR3 is a part of a protein complex required for normal chloroplast and palisade cell development....

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

    DEFF Research Database (Denmark)

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

    2010-01-01

    of Dinophysis acuminata was established by feeding it the phototrophic ciliate Mesodinium rubrum (= Myrionecta rubra), which again was fed the cryptophyte Teleaulax amphioxeia. Molecular analysis comprising the nucleomorph LSU and two chloroplast markers (tufA gene and a fragment from the end of 16S r......DNA to the beginning of 23S rDNA) resulted in identical sequences for the three organisms. Yet, transmission electron microscopy of the three organisms revealed that several chloroplast features separated D. acuminata from both T. amphioxeia and M. rubrum. The thylakoid arrangement, the number of membranes around...

  5. A pea chloroplast translation elongation factor that is regulated by abiotic factors

    International Nuclear Information System (INIS)

    Singh, B.N.; Mishra, R.N.; Agarwal, Pradeep K.; Goswami, Mamta; Nair, Suresh; Sopory, S.K.; Reddy, M.K.

    2004-01-01

    We report the cloning and characterization of both the cDNA (tufA) and genomic clones encoding for a chloroplast translation elongation factor (EF-Tu) from pea. The analysis of the deduced amino acids of the cDNA clone reveals the presence of putative transit peptide sequence and four GTP binding domains and two EF-Tu signature motifs in the mature polypeptide region. Using in vivo immunostaining followed by confocal microscopy pea EF-Tu was localized to chloroplast. The steady state transcript level of pea tufA was high in leaves and not detectable in roots. The expression of this gene is stimulated by light. The differential expression of this gene in response to various abiotic stresses showed that it is down-regulated in response to salinity and ABA and up-regulated in response to low temperature and salicylic acid treatment. These results indicate that regulation of pea tufA may have an important role in plant adaptation to environmental stresses

  6. Role of plastidic pyruvate dehydrogenase complex (pl. PDC) in chloroplast metabolism of spinach

    International Nuclear Information System (INIS)

    Schulze-Siebert, D.; Homeyer, U.; Schultz, G.

    1986-01-01

    Labeling experiments of chloroplasts in the light ( 14 CO 2 , 2- 14 C-pyruvate etc.) revealed that pl. PDC is predominantly involved in the synthesis of branched chain amino acids and pl. isoprenoids (carotenes, PQ, α-T). In this context, pl. phosphoglycerate mutase as missing link in the C 3 → C 2 metabolism of chloroplasts was identified by latency experiments. This indicates a direct pathway from Calvin cycle to pl. PDC. Using protoplasts, maximal rates in pl. PDC metabolism were obtained. On the other hand, mitochondrial PDC in protoplasts is mainly involved in fatty acid synthesis by known mechanism. Additionally, cytosolic-ER-isoprenoids were formed (e.g. sterols). When 14 CO 2 was simultaneously applied with unlabeled acetate to protoplasts in the light an isotopic dilution of fatty acids were found but not of pl. isoprenoids. This may indicate an partially channeling of pl. PDC and mevalonate pathway for pl. isoprenoid synthesis. Inhibitory studies with DCMU point in the same direction

  7. 2012 Gordon Research Conference, Mitochondria and Chloroplasts, July 29 - Aug 3 2012

    Energy Technology Data Exchange (ETDEWEB)

    Barkan, Alice [Univ. of Oregon, Eugene, OR (United States)

    2012-08-03

    The 2012 Gordon Research Conference on Mitochondria and Chloroplasts will assemble an international group of scientists investigating fundamental properties of these organelles, and their integration into broader physiological processes. The conference will emphasize the many commonalities between mitochondria and chloroplasts: their evolution from bacterial endosymbionts, their genomes and gene expression systems, their energy transducing membranes whose proteins derive from both nuclear and organellar genes, the challenge of maintaining organelle integrity in the presence of the reactive oxygen species that are generated during energy transduction, their incorporation into organismal signaling pathways, and more. The conference will bring together investigators working in animal, plant, fungal and protozoan systems who specialize in cell biology, genetics, biochemistry, physiology, proteomics, genomics, and structural biology. As such, this conference will provide a unique forum that engenders cross-disciplinary discussions concerning the biogenesis, dynamics, and regulation of these key cellular structures. By fostering interactions among mammalian, fungal and plant organellar biologists, this conference also provides a conduit for the transmission of mechanistic insights obtained in model organisms to applications in medicine and agriculture. The 2012 conference will highlight areas that are moving rapidly and emerging themes. These include new insights into the ultrastructure and organization of the energy transducing membranes, the coupling of organellar gene expression with the assembly of photosynthetic and respiratory complexes, the regulatory networks that couple organelle biogenesis with developmental and physiological signals, the signaling events through which organellar physiology influences nuclear gene expression, and the roles of organelles in disease and development.

  8. The Cytochrome b 6 f Complex: Biophysical Aspects of Its Functioning in Chloroplasts.

    Science.gov (United States)

    Tikhonov, Alexander N

    2018-01-01

    This chapter presents an overview of structural properties of the cytochrome (Cyt) b 6 f complex and its functioning in chloroplasts. The Cyt b 6 f complex stands at the crossroad of photosynthetic electron transport pathways, providing connectivity between Photosystem (PSI) and Photosysten II (PSII) and pumping protons across the membrane into the thylakoid lumen. After a brief review of the chloroplast electron transport chain, the consideration is focused on the structural organization of the Cyt b 6 f complex and its interaction with plastoquinol (PQH 2 , reduced form of plastoquinone), a mediator of electron transfer from PSII to the Cyt b 6 f complex. The processes of PQH 2 oxidation by the Cyt b 6 f complex have been considered within the framework of the Mitchell's Q-cycle. The overall rate of the intersystem electron transport is determined by PQH 2 turnover at the quinone-binding site Q o of the Cyt b 6 f complex. The rate of PQH 2 oxidation is controlled by the intrathylakoid pH in , which value determines the protonation/deprotonation events in the Q o -center. Two other regulatory mechanisms associated with the Cyt b 6 f complex are briefly overviewed: (i) redistribution of electron fluxes between alternative (linear and cyclic) pathways, and (ii) "state transitions" related to redistribution of solar energy between PSI and PSII.

  9. Outlook in the application of Chlamydomonas reinhardtii chloroplast as a platform for recombinant protein production.

    Science.gov (United States)

    Shamriz, Shabnam; Ofoghi, Hamideh

    Microalgae, also called microphytes, are a vast group of microscopic photosynthetic organisms living in aquatic ecosystems. Microalgae have attracted the attention of biotechnology industry as a platform for extracting natural products with high commercial value. During last decades, microalgae have been also used as cost-effective and easily scalable platform for the production of recombinant proteins with medical and industrial applications. Most progress in this field has been made with Chlamydomonas reinhardtii as a model organism mainly because of its simple life cycle, well-established genetics and ease of cultivation. However, due to the scarcity of existing infrastructure for commercial production and processing together with relatively low product yields, no recombinant products from C. reinhardtii have gained approval for commercial production and most of them are still in research and development. In this review, we focus on the chloroplast of C. reinhardtii as an algal recombinant expression platform and compare its advantages and disadvantages to other currently used expression systems. We then discuss the strategies for engineering the chloroplast of C. reinhardtii to produce recombinant cells and present a comprehensive overview of works that have used this platform for the expression of high-value products.

  10. Binding of 14C-5-aminolevulinic acid to a stromal protein from developing pea chloroplasts

    International Nuclear Information System (INIS)

    Thayer, S.S.; Castelfranco, P.A.; Wilkinson, J.; Benson, G.

    1987-01-01

    14 -5-Aminolevulinic acid ( 14 C-ALA) binds to a stromal protein with an apparent molecular weight of 42-43 KD on LDS and non-denaturing gels. The reaction is rapid. Binding is inhibited by sulfhydryl reagents, mM concentrations of levulinic, dihydroxy heptanoic acids and gabaculine, 10 μM N-methylprotoporphyrin. Dicarboxilic acids, such as δKG, Glu, OAA, do not inhibit. Chloramphenicol, ATP, protoporphyrin, anoxia, light, darkness have no effect. The product, once formed, is stable to treatment with 5% conc. HCl in cold acetone. It can be chased in a second incubation with unlabeled ALA, but not with levulinic acid. No activity was detected in the subplastidic membrane fractions. Western blot analysis failed to reveal any homology between the labeled protein and either cytochrome for ALA dehydratase. This ALA-binding protein was not formed in chloroplasts isolated from fully expanded pea leaves. Therefore, it is deemed likely to participate in ALA metabolism during chloroplast development

  11. Impact of the ion transportome of chloroplasts on the optimization of photosynthesis.

    Science.gov (United States)

    Szabò, Ildikò; Spetea, Cornelia

    2017-06-01

    Ions play fundamental roles in all living cells, and their gradients are often essential to fuel transport, regulate enzyme activities, and transduce energy within cells. Regulation of their homeostasis is essential for cell metabolism. Recent results indicate that modulation of ion fluxes might also represent a useful strategy to regulate one of the most important physiological processes taking place in chloroplasts, photosynthesis. Photosynthesis is highly regulated, due to its unique role as a cellular engine for growth in the light. Controlling the balance between ATP and NADPH synthesis is a critical task, and availability of these molecules can limit the overall photosynthetic yield. Photosynthetic organisms optimize photosynthesis in low light, where excitation energy limits CO2 fixation, and minimize photo-oxidative damage in high light by dissipating excess photons. Despite extensive studies of these phenomena, the mechanism governing light utilization in plants is still poorly understood. In this review, we provide an update of the recently identified chloroplast-located ion channels and transporters whose function impacts photosynthetic efficiency in plants. © The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  12. Chloroplast Dysfunction Causes Multiple Defects in Cell Cycle Progression in the Arabidopsis crumpled leaf Mutant

    KAUST Repository

    Hudik, Elodie

    2014-07-18

    The majority of research on cell cycle regulation is focused on the nuclear events that govern the replication and segregation of the genome between the two daughter cells. However, eukaryotic cells contain several compartmentalized organelles with specialized functions, and coordination among these organelles is required for proper cell cycle progression, as evidenced by the isolation of several mutants in which both organelle function and overall plant development were affected. To investigate how chloroplast dysfunction affects the cell cycle, we analyzed the crumpled leaf (crl) mutant of Arabidopsis (Arabidopsis thaliana), which is deficient for a chloroplastic protein and displays particularly severe developmental defects. In the crl mutant, we reveal that cell cycle regulation is altered drastically and that meristematic cells prematurely enter differentiation, leading to reduced plant stature and early endoreduplication in the leaves. This response is due to the repression of several key cell cycle regulators as well as constitutive activation of stress-response genes, among them the cell cycle inhibitor SIAMESE-RELATED5. One unique feature of the crl mutant is that it produces aplastidic cells in several organs, including the root tip. By investigating the consequence of the absence of plastids on cell cycle progression, we showed that nuclear DNA replication occurs in aplastidic cells in the root tip, which opens future research prospects regarding the dialogue between plastids and the nucleus during cell cycle regulation in higher plants.

  13. Effects of gold nanoparticles on the photophysical and photosynthetic parameters of leaves and chloroplasts.

    Science.gov (United States)

    Torres, Rocio; Diz, Virginia E; Lagorio, M Gabriela

    2018-04-18

    Effects of gold nanoparticles (average diameter: 10-14 nm) on leaves and chloroplasts have been studied. Gold nanoparticles (AuNPs) quenched significantly chlorophyll fluorescence when introduced both in intact leaves and isolated chloroplasts. Additionally, the fluorescence spectra corrected for light re-absorption processes showed a net decrease in the fluorescence ratio calculated as the quotient between the maximum fluorescence at 680 and 735 nm. This fact gave evidence for a reduction in the fluorescence emission of the PSII relative to that of the PSI. Strikingly, the photosynthetic parameters derived from the analysis of the slow phase of Kautsky's kinetics, the rate of oxygen evolution and the rate of photo-reduction of 2,6-dichlorophenolindophenol were increased in the presence of AuNPs indicating an apparent greater photosynthetic capacity. The observed results were consistent with an electron transfer process from the excited PSII, which was thermodynamically possible, and which competed with both the electron transport process that initiated photosynthesis and the deactivation of the excited PSII by fluorescence emission. Additionally, it is here explained, in terms of a completely rational kinetic scheme and their corresponding algebraic expressions, why the photosynthetic parameters and the variable and non-variable fluorescence of chlorophyll are modified in a photosynthetic tissue containing gold nanoparticles.

  14. Two CRM protein subfamilies cooperate in the splicing of group IIB introns in chloroplasts.

    Science.gov (United States)

    Asakura, Yukari; Bayraktar, Omer Ali; Barkan, Alice

    2008-11-01

    Chloroplast genomes in angiosperms encode approximately 20 group II introns, approximately half of which are classified as subgroup IIB. The splicing of all but one of the subgroup IIB introns requires a heterodimer containing the peptidyl-tRNA hydrolase homolog CRS2 and one of two closely related proteins, CAF1 or CAF2, that harbor a recently recognized RNA binding domain called the CRM domain. Two CRS2/CAF-dependent introns require, in addition, a CRM domain protein called CFM2 that is only distantly related to CAF1 and CAF2. Here, we show that CFM3, a close relative of CFM2, associates in vivo with those CRS2/CAF-dependent introns that are not CFM2 ligands. Mutant phenotypes in rice and Arabidopsis support a role for CFM3 in the splicing of most of the introns with which it associates. These results show that either CAF1 or CAF2 and either CFM2 or CFM3 simultaneously bind most chloroplast subgroup IIB introns in vivo, and that the CAF and CFM subunits play nonredundant roles in splicing. These results suggest that the expansion of the CRM protein family in plants resulted in two subfamilies that play different roles in group II intron splicing, with further diversification within a subfamily to accommodate multiple intron ligands.

  15. A plastome mutation affects processing of both chloroplast and nuclear DNA-encoded plastid proteins.

    Science.gov (United States)

    Johnson, E M; Schnabelrauch, L S; Sears, B B

    1991-01-01

    Immunoblotting of a chloroplast mutant (pm7) of Oenothera showed that three proteins, cytochrome f and the 23 kDa and 16 kDa subunits of the oxygen-evolving subcomplex of photosystem II, were larger than the corresponding mature proteins of the wild type and, thus, appear to be improperly processed in pm7. The mutant is also chlorotic and has little or no internal membrane development in the plastids. The improperly processed proteins, and other proteins that are completely missing, represent products of both the plastid and nuclear genomes. To test for linkage of these defects, a green revertant of pm7 was isolated from cultures in which the mutant plastids were maintained in a nuclear background homozygous for the plastome mutator (pm) gene. In this revertant, all proteins analyzed co-reverted to the wild-type condition, indicating that a single mutation in a plastome gene is responsible for the complex phenotype of pm7. These results suggest that the defect in pm7 lies in a gene that affects a processing protease encoded in the chloroplast genome.

  16. The complete chloroplast genome sequence of Aster spathulifolius (Asteraceae); genomic features and relationship with Asteraceae.

    Science.gov (United States)

    Choi, Kyoung Su; Park, SeonJoo

    2015-11-10

    Aster spathulifolius, a member of the Asteraceae family, is distributed along the coast of Japan and Korea. This plant is used for medicinal and ornamental purposes. The complete chloroplast (cp) genome of A. sphathulifolius consists of 149,473 bp that include a pair of inverted repeats of 24,751 bp separated by a large single copy region of 81,998 bp and a small single copy region of 17,973 bp. The chloroplast genome contains 78 coding genes, four rRNA genes and 29 tRNA genes. When compared to other cpDNA sequences of Asteraceae, A. spathulifolius showed the closest relationship with Jacobaea vulgaris, and its atpB gene was found to be a pseudogene, unlike J. vulgaris. Furthermore, evaluation of the gene compositions of J. vulgaris, Helianthus annuus, Guizotia abyssinica and A. spathulifolius revealed that 13.6-kb showed inversion from ndhF to rps15, unlike Lactuca of Asteraceae. Comparison of the synonymous (Ks) and nonsynonymous (Ka) substitution rates with J. vulgaris revealed that synonymous genes related to a small subunit of the ribosome showed the highest value (0.1558), while nonsynonymous rates of genes related to ATP synthase genes were highest (0.0118). These findings revealed that substitution has occurred at similar rates in most genes, and the substitution rates suggested that most genes is a purified selection. Copyright © 2015 Elsevier B.V. All rights reserved.

  17. Optimization of ATP synthase function in mitochondria and chloroplasts via the adenylate kinase equilibrium

    Directory of Open Access Journals (Sweden)

    Abir U Igamberdiev

    2015-01-01

    Full Text Available The bulk of ATP synthesis in plants is performed by ATP synthase, the main bioenergetics engine of cells, operating both in mitochondria and in chloroplasts. The reaction mechanism of ATP synthase has been studied in detail for over half a century; however, its optimal performance depends also on the steady delivery of ATP synthase substrates and the removal of its products. For mitochondrial ATP synthase, we analyze here the provision of stable conditions for (i the supply of ADP and Mg2+, supported by adenylate kinase (AK equilibrium in the intermembrane space, (ii the supply of phosphate via membrane transporter in symport with H+, and (iii the conditions of outflow of ATP by adenylate transporter carrying out the exchange of free adenylates. We also show that, in chloroplasts, AK equilibrates adenylates and governs Mg2+ contents in the stroma, optimizing ATP synthase and Calvin cycle operation, and affecting the import of inorganic phosphate in exchange with triose phosphates. It is argued that chemiosmosis is not the sole component of ATP synthase performance, which also depends on AK-mediated equilibrium of adenylates and Mg2+, adenylate transport and phosphate release and supply.

  18. Chloroplastic protein NRIP1 mediates innate immune receptor recognition of a viral effector

    Science.gov (United States)

    Caplan, Jeffrey L.; Mamillapalli, Padmavathi; Burch-Smith, Tessa M.; Czymmek, Kirk; Dinesh-Kumar, S.P.

    2008-01-01

    Summary Plant innate immunity relies on the recognition of pathogen effector molecules by nucleotide-binding-leucine-rich repeat (NB-LRR) immune receptor families. Previously we have shown the N immune receptor, a member of TIR-NB-LRR family, indirectly recognizes the 50-kDa helicase (p50) domain of Tobacco mosaic virus (TMV) through its TIR domain. We have identified an N receptor-interacting protein, NRIP1, that directly interacts with both N's TIR domain and p50. NRIP1 is a functional rhodanese sulfurtransferase and is required for N to provide complete resistance to TMV. Interestingly, NRIP1 that normally localizes to the chloroplasts is recruited to the cytoplasm and nucleus by the p50 effector. As a consequence, NRIP1 interacts with N only in the presence of the p50 effector. Our findings show that a chloroplastic protein is intimately involved in pathogen recognition. We propose that N's activation requires a pre-recognition complex containing the p50 effector and NRIP1. PMID:18267075

  19. The different fates of mitochondria and chloroplasts during dark-induced senescence in Arabidopsis leaves.

    Science.gov (United States)

    Keech, Olivier; Pesquet, Edouard; Ahad, Abdul; Askne, Anna; Nordvall, Dag; Vodnala, Sharvani Munender; Tuominen, Hannele; Hurry, Vaughan; Dizengremel, Pierre; Gardeström, Per

    2007-12-01

    Senescence is an active process allowing the reallocation of valuable nutrients from the senescing organ towards storage and/or growing tissues. Using Arabidopsis thaliana leaves from both whole darkened plants (DPs) and individually darkened leaves (IDLs), we investigated the fate of mitochondria and chloroplasts during dark-induced leaf senescence. Combining in vivo visualization of fates of the two organelles by three-dimensional reconstructions of abaxial parts of leaves with functional measurements of photosynthesis and respiration, we showed that the two experimental systems displayed major differences during 6 d of dark treatment. In whole DPs, organelles were largely retained in both epidermal and mesophyll cells. However, while the photosynthetic capacity was maintained, the capacity of mitochondrial respiration decreased. In contrast, IDLs showed a rapid decline in photosynthetic capacity while maintaining a high capacity for mitochondrial respiration throughout the treatment. In addition, we noticed an unequal degradation of organelles in the different cell types of the senescing leaf. From these data, we suggest that metabolism in leaves of the whole DPs enters a 'stand-by mode' to preserve the photosynthetic machinery for as long as possible. However, in IDLs, mitochondria actively provide energy and carbon skeletons for the degradation of cell constituents, facilitating the retrieval of nutrients. Finally, the heterogeneity of the degradation processes involved during senescence is discussed with regard to the fate of mitochondria and chloroplasts in the different cell types.

  20. Unraveling the nuclear and chloroplast genomes of an agar producing red macroalga, Gracilaria changii (Rhodophyta, Gracilariales).

    Science.gov (United States)

    Ho, Chai-Ling; Lee, Wei-Kang; Lim, Ee-Leen

    2018-03-01

    Agar and agarose have wide applications in food and pharmaceutical industries. Knowledge on the genome of red seaweeds that produce them is still lacking. To fill the gap in genome analyses of these red algae, we have sequenced the nuclear and organellar genomes of an agarophyte, Gracilaria changii. The partial nuclear genome sequence of G. changii has a total length of 35.8Mb with 10,912 predicted protein coding sequences. Only 39.4% predicted proteins were found to have significant matches to protein sequences in SwissProt. The chloroplast genome of G. changii is 183,855bp with a total of 201 open reading frames (ORFs), 29 tRNAs and 3 rRNAs predicted. Five genes: ssrA, leuC and leuD CP76_p173 (orf139) and pbsA were absent in the chloroplast genome of G. changii. The genome information is valuable in accelerating functional studies of individual genes and resolving evolutionary relationship of red seaweeds. Copyright © 2017 Elsevier Inc. All rights reserved.

  1. Combining chloroplast and nuclear microsatellites to investigate origin and dispersal of New World sweet potato landraces.

    Science.gov (United States)

    Roullier, C; Rossel, G; Tay, D; McKey, D; Lebot, V

    2011-10-01

    We analysed a representative collection of New World sweet potato landraces (329 accessions from Mexico to Peru) with both chloroplast and nuclear microsatellite markers. Both kinds of markers supported the existence of two geographically restricted genepools, corresponding to accessions from the north-western part of South America and accessions from the Caribbean and Central America region. Our conservative cpSSRs markers revealed that the divergence between the two haplotype groups is associated with numerous mutation events concerning various markers, supporting the idea that this divergence may be ancient, predating domestication. For both kinds of markers, we found no significant difference in diversity between the two genepools and detected region-specific alleles in both groups. Previous studies have favoured the hypothesis of a single domestication of this crop. Our analysis suggests at least two independent domestications, in Central/Caribbean America and in the north-western part of South America. Sweet potato was then dispersed from these centres throughout tropical America. Comparison of nuclear and chloroplast data suggests that exchanges of clones and sexual reproduction were both important processes in landrace diversification in this clonally propagated crop. Our analysis provides useful tools for rationalizing the conservation and use of sweet potato germplasm collections. © 2011 Blackwell Publishing Ltd.

  2. Organization and post-transcriptional processing of the psb B operon from chloroplasts of Populus deltoides.

    Science.gov (United States)

    Dixit, R; Trivedi, P K; Nath, P; Sane, P V

    1999-09-01

    Chloroplast genes are typically organized into polycistronic transcription units that give rise to complex sets of mono- and oligo-cistronic overlapping RNAs through a series of processing steps. The psbB operon contains genes for the PSII (psbB, psbT, psbH) and cytochrome b(6)f (petB and petD) complexes which are needed in different amounts during chloroplast biogenesis. The functional significance of gene organization in this polycistronic unit, containing information for two different complexes, is not known and is of interest. To determine the organization and expression of these complexes, studies have been carried out on crop plants by different groups, but not much information is known about trees. We present the nucleotide sequences of PSII genes and RNA profiles of the genes located in the psbB operon from Populus deltoides, a tree species. Although the gene organization of this operon in P. deltoides is similar to that in other species, a few variations have been observed in the processing scheme.

  3. Seawater spray injury to Quercus acutissima leaves: crystal deposition, stomatal clogging, and chloroplast degeneration.

    Science.gov (United States)

    Kim, Ki Woo; Koo, Kyosang; Kim, Pan-Gi

    2011-05-01

    Effects of seawater spray on leaf structure were investigated in Quercus acutissima by electron microscopy and X-ray microanalysis. Two-year-old seedlings of Q. acutissima were sprayed with seawater and kept in a greenhouse maintained at 25°C. The most recognizable symptoms of seawater-sprayed seedlings included leaf necrosis, crystal deposition, stomatal clogging, and chloroplast degeneration. Field emission scanning electron microscopy revealed that the leaf surface was covered with additional layers of remnants of seawater spray. Composed of sodium and chloride, cube-shaped crystals (halite) were prevalently found on trichomes and epidermis, and formed aggregates. Meanwhile, wedge-shaped crystals were deposited on epidermis and consisted of calcium and sulfur. As a result of stomatal clogging by crystal deposition on the abaxial surface, it was conceivable that plant respiration became severely hampered. Transmission electron microscopy showed degenerated cytoplasm of seawater-sprayed leaves. It was common to observe severe plasmolysis and disrupted chloroplasts with a reduced number of thylakoids in grana. These results indicate that foliar applications of seawater were sufficient to induce necrosis of Q. acutissima seedlings as an abiotic disturbance factor. Copyright © 2010 Wiley-Liss, Inc.

  4. Reductive evolution of chloroplasts in non-photosynthetic plants, algae and protists.

    Science.gov (United States)

    Hadariová, Lucia; Vesteg, Matej; Hampl, Vladimír; Krajčovič, Juraj

    2018-04-01

    Chloroplasts are generally known as eukaryotic organelles whose main function is photosynthesis. They perform other functions, however, such as synthesizing isoprenoids, fatty acids, heme, iron sulphur clusters and other essential compounds. In non-photosynthetic lineages that possess plastids, the chloroplast genomes have been reduced and most (or all) photosynthetic genes have been lost. Consequently, non-photosynthetic plastids have also been reduced structurally. Some of these non-photosynthetic or "cryptic" plastids were overlooked or unrecognized for decades. The number of complete plastid genome sequences and/or transcriptomes from non-photosynthetic taxa possessing plastids is rapidly increasing, thus allowing prediction of the functions of non-photosynthetic plastids in various eukaryotic lineages. In some non-photosynthetic eukaryotes with photosynthetic ancestors, no traces of plastid genomes or of plastids have been found, suggesting that they have lost the genomes or plastids completely. This review summarizes current knowledge of non-photosynthetic plastids, their genomes, structures and potential functions in free-living and parasitic plants, algae and protists. We introduce a model for the order of plastid gene losses which combines models proposed earlier for land plants with the patterns of gene retention and loss observed in protists. The rare cases of plastid genome loss and complete plastid loss are also discussed.

  5. Chloroplast mutations induced by 9-aminoacridine hydrochloride are independent of the plastome mutator in Oenothera.

    Science.gov (United States)

    GuhaMajumdar, M; Baldwin, S; Sears, B B

    2004-02-01

    Oenothera plants homozygous for the recessive plastome mutator allele ( pm) show chloroplast DNA (cpDNA) mutation frequencies that are about 1,000-fold higher than spontaneous levels. The pm-encoded gene product has been hypothesized to have a function in cpDNA replication, repair and/or mutation avoidance. Previous chemical mutagenesis experiments with the alkylating agent nitroso-methyl urea (NMU) showed a synergistic effect of NMU on the induction of mutations in the pm line, suggesting an interaction between the pm-encoded gene product and one of the repair systems that corrects alkylation damage. The goal of the experiments described here was to examine whether the pm activity extends to the repair of damage caused by non-alkylating mutagens. To this end, the intercalating mutagen, 9-aminoacridine hydrochloride (9AA) was tested for synergism with the plastome mutator. A statistical analysis of the data reported here indicates that the pm-encoded gene product is not involved in the repair of the 9AA-induced mutations. However, the recovery of chlorotic sectors in plants derived from the mutagenized seeds shows that 9AA can act as a mutagen of the chloroplast genome.

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

    Directory of Open Access Journals (Sweden)

    Hanul eKim

    2015-02-01

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

  7. A chloroplast lipoxygenase is required for wound-induced jasmonic acid accumulation in Arabidopsis.

    Science.gov (United States)

    Bell, E; Creelman, R A; Mullet, J E

    1995-09-12

    Plant lipoxygenases are thought to be involved in the biosynthesis of lipid-derived signaling molecules. The potential involvement of a specific Arabidopsis thaliana lipoxygenase isozyme, LOX2, in the biosynthesis of the plant growth regulators jasmonic acid (JA) and abscisic acid was investigated. Our characterization of LOX2 indicates that the protein is targeted to chloroplasts. The physiological role of this chloroplast lipoxygenase was analyzed in transgenic plants where cosuppression reduced LOX2 accumulation. The reduction in LOX2 levels caused no obvious changes in plant growth or in the accumulation of abscisic acid. However, the wound-induced accumulation of JA observed in control plants was absent in leaves of transgenic plants that lacked LOX2. Thus, LOX2 is required for the wound-induced synthesis of the plant growth regulator JA in leaves. We also examined the expression of a wound- and JA-inducible Arabidopsis gene, vsp, in transgenic and control plants. Leaves of transgenic plants lacking LOX2 accumulated less vsp mRNA than did control leaves in response to wounding. This result suggests that wound-induced JA (or some other LOX2-requiring component of the wound response pathway) is involved in the wound-induced regulation of this gene.

  8. Expression and assembly of largest foreign protein in chloroplasts: oral delivery of human FVIII made in lettuce chloroplasts robustly suppresses inhibitor formation in haemophilia A mice.

    Science.gov (United States)

    Kwon, Kwang-Chul; Sherman, Alexandra; Chang, Wan-Jung; Kamesh, Aditya; Biswas, Moanaro; Herzog, Roland W; Daniell, Henry

    2017-11-06

    Inhibitor formation is a serious complication of factor VIII (FVIII) replacement therapy for the X-linked bleeding disorder haemophilia A and occurs in 20%-30% of patients. No prophylactic tolerance protocol currently exists. Although we reported oral tolerance induction using FVIII domains expressed in tobacco chloroplasts, significant challenges in clinical advancement include expression of the full-length CTB-FVIII sequence to cover the entire patient population, regardless of individual CD4 + T-cell epitope responses. Codon optimization of FVIII heavy chain (HC) and light chain (LC) increased expression 15- to 42-fold higher than the native human genes. Homoplasmic lettuce lines expressed CTB fusion proteins of FVIII-HC (99.3 kDa), LC (91.8 kDa), C2 (31 kDa) or single chain (SC, 178.2 kDa) up to 3622, 263, 3321 and 852 μg/g in lyophilized plant cells, when grown in a cGMP hydroponic facility (Fraunhofer). CTB-FVIII-SC is the largest foreign protein expressed in chloroplasts; despite a large pentamer size (891 kDa), assembly, folding and disulphide bonds were maintained upon lyophilization and long-term storage as revealed by GM1-ganglioside receptor binding assays. Repeated oral gavages (twice/week for 2 months) of CTB-FVIII-HC/CTB-FVIII-LC reduced inhibitor titres ~10-fold (average 44 BU/mL to 4.7 BU/mL) in haemophilia A mice. Most importantly, increase in the frequency of circulating LAP-expressing CD4 + CD25 + FoxP3 + Treg in tolerized mice could be used as an important cellular biomarker in human clinical trials for plant-based oral tolerance induction. In conclusion, this study reports the first clinical candidate for oral tolerance induction that is urgently needed to protect haemophilia A patients receiving FVIII injections. © 2017 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

  9. Balanced gene losses, duplications and intensive rearrangements led to an unusual regularly sized genome in Arbutus unedo chloroplasts.

    Science.gov (United States)

    Martínez-Alberola, Fernando; Del Campo, Eva M; Lázaro-Gimeno, David; Mezquita-Claramonte, Sergio; Molins, Arantxa; Mateu-Andrés, Isabel; Pedrola-Monfort, Joan; Casano, Leonardo M; Barreno, Eva

    2013-01-01

    Completely sequenced plastomes provide a valuable source of information about the duplication, loss, and transfer events of chloroplast genes and phylogenetic data for resolving relationships among major groups of plants. Moreover, they can also be useful for exploiting chloroplast genetic engineering technology. Ericales account for approximately six per cent of eudicot diversity with 11,545 species from which only three complete plastome sequences are currently available. With the aim of increasing the number of ericalean complete plastome sequences, and to open new perspectives in understanding Mediterranean plant adaptations, a genomic study on the basis of the complete chloroplast genome sequencing of Arbutus unedo and an updated phylogenomic analysis of Asteridae was implemented. The chloroplast genome of A. unedo shows extensive rearrangements but a medium size (150,897 nt) in comparison to most of angiosperms. A number of remarkable distinct features characterize the plastome of A. unedo: five-fold dismissing of the SSC region in relation to most angiosperms; complete loss or pseudogenization of a number of essential genes; duplication of the ndhH-D operon and its location within the two IRs; presence of large tandem repeats located near highly re-arranged regions and pseudogenes. All these features outline the primary evolutionary split between Ericaceae and other ericalean families. The newly sequenced plastome of A. unedo with the available asterid sequences allowed the resolution of some uncertainties in previous phylogenies of Asteridae.

  10. Balanced gene losses, duplications and intensive rearrangements led to an unusual regularly sized genome in Arbutus unedo chloroplasts.

    Directory of Open Access Journals (Sweden)

    Fernando Martínez-Alberola

    Full Text Available Completely sequenced plastomes provide a valuable source of information about the duplication, loss, and transfer events of chloroplast genes and phylogenetic data for resolving relationships among major groups of plants. Moreover, they can also be useful for exploiting chloroplast genetic engineering technology. Ericales account for approximately six per cent of eudicot diversity with 11,545 species from which only three complete plastome sequences are currently available. With the aim of increasing the number of ericalean complete plastome sequences, and to open new perspectives in understanding Mediterranean plant adaptations, a genomic study on the basis of the complete chloroplast genome sequencing of Arbutus unedo and an updated phylogenomic analysis of Asteridae was implemented. The chloroplast genome of A. unedo shows extensive rearrangements but a medium size (150,897 nt in comparison to most of angiosperms. A number of remarkable distinct features characterize the plastome of A. unedo: five-fold dismissing of the SSC region in relation to most angiosperms; complete loss or pseudogenization of a number of essential genes; duplication of the ndhH-D operon and its location within the two IRs; presence of large tandem repeats located near highly re-arranged regions and pseudogenes. All these features outline the primary evolutionary split between Ericaceae and other ericalean families. The newly sequenced plastome of A. unedo with the available asterid sequences allowed the resolution of some uncertainties in previous phylogenies of Asteridae.

  11. Chloroplast DNA variation in European white oaks phylogeography and patterns of diversity based on data from over 2600 populations

    NARCIS (Netherlands)

    Petit, R.J.; Csaikl, U.M.; Bordács, S.; Burg, K.; Coart, E.; Cottrell, J.; Dam, van B.C.; Deans, J.D.; Dumolin-LapOgue, S.; Fineschi, S.; Finkeldey, R.; Gillies, A.; Glaz, I.; Goicoechea, P.G.; Jensen, J.S.; König, A.O.; Lowe, A.J.; Madsen, S.F.; Mátyás, G.; Munro, R.C.; Olalde, M.; Pemonge, M.H.; Popescu, F.; Slade, D.; Tabbener, H.; Taurchini, D.; Vries, de S.G.M.; Ziegenhagen, B.; Kremer, A.

    2002-01-01

    A consortium of 16 laboratories have studied chloroplast DNA (cpDNA) variation in European white oaks. A common strategy for molecular screening, based on restriction analysis of four PCR-amplified cpDNA fragments, was used to allow comparison among the different laboratories. A total of 2613 oak

  12. The electrical potential as a gauge of photosynthetic performance in plant chloroplasts : a patch-clamp study

    NARCIS (Netherlands)

    Voorthuysen, van T.

    1997-01-01

    The earliest events in the energisation of the photosynthetic membrane upon light capture are the formation of a transmembrane electrical potential (AV) and a transmembrane proton gradient (ΔpH). In this thesis ΔΨis employed for the study of the bioenergetics of chloroplast photosynthesis

  13. Evolutionary transition towards permanent chloroplasts? - Division of kleptochloroplasts in starved cells of two species of Dinophysis (Dinophyceae.

    Directory of Open Access Journals (Sweden)

    Pernille Møller Rusterholz

    Full Text Available Species within the marine toxic dinoflagellate genus Dinophysis are phagotrophic organisms that exploit chloroplasts (kleptochloroplasts from other protists to perform photosynthesis. Dinophysis spp. acquire the kleptochloroplasts from the ciliate Mesodinium rubrum, which in turn acquires the chloroplasts from a unique clade of cryptophytes. Dinophysis spp. digest the prey nuclei and all other cell organelles upon ingestion (except the kleptochloroplasts and they are therefore believed to constantly acquire new chloroplasts as the populations grow. Previous studies have, however, indicated that Dinophysis can keep the kleptochloroplasts active during long term starvation and are able to produce photosynthetic pigments when exposed to prey starvation. This indicates a considerable control over the kleptochloroplasts and the ability of Dinophysis to replicate its kleptochloroplasts was therefore re-investigated in detail in this study. The kleptochloroplasts of Dinophysis acuta and Dinophysis acuminata were analyzed using confocal microscopy and 3D bioimaging software during long term starvation experiments. The cell concentrations were monitored to confirm cell divisions and samples were withdrawn each time a doubling had occurred. The results show direct evidence of kleptochloroplastidic division and that the decreases in total kleptochloroplast volume, number of kleptochloroplasts and number of kleptochloroplast centers were not caused by dilution due to cell divisions. This is the first report of division of kleptochloroplasts in any protist without the associated prey nuclei. This indicates that Dinophysis spp. may be in a transitional phase towards possessing permanent chloroplasts, which thereby potentially makes it a key organism to understand the evolution of phototrophic protists.

  14. Characterization and Comparative Analysis of the Complete Chloroplast Genome of the Critically Endangered Species Streptocarpus teitensis (Gesneriaceae

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    Cornelius M. Kyalo

    2018-01-01

    Full Text Available Streptocarpus teitensis (Gesneriaceae is an endemic species listed as critically endangered in the International Union for Conservation of Nature (IUCN red list of threatened species. However, the sequence and genome information of this species remains to be limited. In this article, we present the complete chloroplast genome structure of Streptocarpus teitensis and its evolution inferred through comparative studies with other related species. S. teitensis displayed a chloroplast genome size of 153,207 bp, sheltering a pair of inverted repeats (IR of 25,402 bp each split by small and large single-copy (SSC and LSC regions of 18,300 and 84,103 bp, respectively. The chloroplast genome was observed to contain 116 unique genes, of which 80 are protein-coding, 32 are transfer RNAs, and four are ribosomal RNAs. In addition, a total of 196 SSR markers were detected in the chloroplast genome of Streptocarpus teitensis with mononucleotides (57.1% being the majority, followed by trinucleotides (33.2% and dinucleotides and tetranucleotides (both 4.1%, and pentanucleotides being the least (1.5%. Genome alignment indicated that this genome was comparable to other sequenced members of order Lamiales. The phylogenetic analysis suggested that Streptocarpus teitensis is closely related to Lysionotus pauciflorus and Dorcoceras hygrometricum.

  15. The complete chloroplast genome sequences of Lychnis wilfordii and Silene capitata and comparative analyses with other Caryophyllaceae genomes.

    Science.gov (United States)

    Kang, Jong-Soo; Lee, Byoung Yoon; Kwak, Myounghai

    2017-01-01

    The complete chloroplast genomes of Lychnis wilfordii and Silene capitata were determined and compared with ten previously reported Caryophyllaceae chloroplast genomes. The chloroplast genome sequences of L. wilfordii and S. capitata contain 152,320 bp and 150,224 bp, respectively. The gene contents and orders among 12 Caryophyllaceae species are consistent, but several microstructural changes have occurred. Expansion of the inverted repeat (IR) regions at the large single copy (LSC)/IRb and small single copy (SSC)/IR boundaries led to partial or entire gene duplications. Additionally, rearrangements of the LSC region were caused by gene inversions and/or transpositions. The 18 kb inversions, which occurred three times in different lineages of tribe Sileneae, were thought to be facilitated by the intermolecular duplicated sequences. Sequence analyses of the L. wilfordii and S. capitata genomes revealed 39 and 43 repeats, respectively, including forward, palindromic, and reverse repeats. In addition, a total of 67 and 56 simple sequence repeats were discovered in the L. wilfordii and S. capitata chloroplast genomes, respectively. Finally, we constructed phylogenetic trees of the 12 Caryophyllaceae species and two Amaranthaceae species based on 73 protein-coding genes using both maximum parsimony and likelihood methods.

  16. Assembly of the Complete Sitka Spruce Chloroplast Genome Using 10X Genomics' GemCode Sequencing Data.

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    Lauren Coombe

    Full Text Available The linked read sequencing library preparation platform by 10X Genomics produces barcoded sequencing libraries, which are subsequently sequenced using the Illumina short read sequencing technology. In this new approach, long fragments of DNA are partitioned into separate micro-reactions, where the same index sequence is incorporated into each of the sequencing fragment inserts derived from a given long fragment. In this study, we exploited this property by using reads from index sequences associated with a large number of reads, to assemble the chloroplast genome of the Sitka spruce tree (Picea sitchensis. Here we report on the first Sitka spruce chloroplast genome assembled exclusively from P. sitchensis genomic libraries prepared using the 10X Genomics protocol. We show that the resulting 124,049 base pair long genome shares high sequence similarity with the related white spruce and Norway spruce chloroplast genomes, but diverges substantially from a previously published P. sitchensis- P. thunbergii chimeric genome. The use of reads from high-frequency indices enabled separation of the nuclear genome reads from that of the chloroplast, which resulted in the simplification of the de Bruijn graphs used at the various stages of assembly.

  17. Large Diversity of Nonstandard Genes and Dynamic Evolution of Chloroplast Genomes in Siphonous Green Algae (Bryopsidales, Chlorophyta).

    Science.gov (United States)

    Cremen, Ma Chiela M; Leliaert, Frederik; Marcelino, Vanessa R; Verbruggen, Heroen

    2018-04-01

    Chloroplast genomes have undergone tremendous alterations through the evolutionary history of the green algae (Chloroplastida). This study focuses on the evolution of chloroplast genomes in the siphonous green algae (order Bryopsidales). We present five new chloroplast genomes, which along with existing sequences, yield a data set representing all but one families of the order. Using comparative phylogenetic methods, we investigated the evolutionary dynamics of genomic features in the order. Our results show extensive variation in chloroplast genome architecture and intron content. Variation in genome size is accounted for by the amount of intergenic space and freestanding open reading frames that do not show significant homology to standard plastid genes. We show the diversity of these nonstandard genes based on their conserved protein domains, which are often associated with mobile functions (reverse transcriptase/intron maturase, integrases, phage- or plasmid-DNA primases, transposases, integrases, ligases). Investigation of the introns showed proliferation of group II introns in the early evolution of the order and their subsequent loss in the core Halimedineae, possibly through RT-mediated intron loss.

  18. Brassinosteroid-induced CO{sub 2} assimilation is associated with increased stability of redox-sensitive photosynthetic enzymes in the chloroplasts in cucumber plants

    Energy Technology Data Exchange (ETDEWEB)

    Jiang, Yu Ping; Cheng, Fei; Zhou, Yan Hong; Xia, Xiao Jian; Mao, Wei Hua; Shi, Kai [Department of Horticulture, Zijingang Campus, Zhejiang University, Yuhangtang Road 866, Hangzhou 310058 (China); Chen, Zhi Xiang [Department of Horticulture, Zijingang Campus, Zhejiang University, Yuhangtang Road 866, Hangzhou 310058 (China); Department of Botany and Plant Pathology, Purdue University, West Lafayette, IN 47907-2054 (United States); Yu, Jing Quan, E-mail: jqyu@zju.edu.cn [Department of Horticulture, Zijingang Campus, Zhejiang University, Yuhangtang Road 866, Hangzhou 310058 (China); Key Laboratory of Horticultural Plants Growth, Development and Quality Improvement, Ministry of Agriculture of China, Yuhangtang Road 866, Hangzhou 310058 (China)

    2012-09-28

    Highlights: Black-Right-Pointing-Pointer Activity of certain Calvin cycle enzymes and CO{sub 2} assimilation are induced by BRs. Black-Right-Pointing-Pointer BRs upregulate the activity of the ascorbate-glutathione cycle in the chloroplasts. Black-Right-Pointing-Pointer BRs increase the chloroplast thiol reduction state. Black-Right-Pointing-Pointer A BR-induced reducing environment increases the stability of photosynthetic enzymes. -- Abstract: Brassinosteroids (BRs) play important roles in plant growth, development, photosynthesis and stress tolerance; however, the mechanism underlying BR-enhanced photosynthesis is currently unclear. Here, we provide evidence that an increase in the BR level increased the quantum yield of PSII, activities of Rubisco activase (RCA) and fructose-1,6-bisphosphatase (FBPase), and CO{sub 2} assimilation. BRs upregulated the transcript levels of genes and activity of enzymes involved in the ascorbate-glutathione cycle in the chloroplasts, leading to an increased ratio of reduced (GSH) to oxidized (GSSG) glutathione in the chloroplasts. An increased GSH/GSSG ratio protected RCA from proteolytic digestion and increased the stability of redox-sensitive enzymes in the chloroplasts. These results strongly suggest that BRs are capable of regulating the glutathione redox state in the chloroplasts through the activation of the ascorbate-glutathione cycle. The resulting increase in the chloroplast thiol reduction state promotes CO{sub 2} assimilation, at least in part, by enhancing the stability and activity of redox-sensitive photosynthetic enzymes through post-translational modifications.

  19. Brassinosteroid-induced CO2 assimilation is associated with increased stability of redox-sensitive photosynthetic enzymes in the chloroplasts in cucumber plants

    International Nuclear Information System (INIS)

    Jiang, Yu Ping; Cheng, Fei; Zhou, Yan Hong; Xia, Xiao Jian; Mao, Wei Hua; Shi, Kai; Chen, Zhi Xiang; Yu, Jing Quan

    2012-01-01

    Highlights: ► Activity of certain Calvin cycle enzymes and CO 2 assimilation are induced by BRs. ► BRs upregulate the activity of the ascorbate–glutathione cycle in the chloroplasts. ► BRs increase the chloroplast thiol reduction state. ► A BR-induced reducing environment increases the stability of photosynthetic enzymes. -- Abstract: Brassinosteroids (BRs) play important roles in plant growth, development, photosynthesis and stress tolerance; however, the mechanism underlying BR-enhanced photosynthesis is currently unclear. Here, we provide evidence that an increase in the BR level increased the quantum yield of PSII, activities of Rubisco activase (RCA) and fructose-1,6-bisphosphatase (FBPase), and CO 2 assimilation. BRs upregulated the transcript levels of genes and activity of enzymes involved in the ascorbate–glutathione cycle in the chloroplasts, leading to an increased ratio of reduced (GSH) to oxidized (GSSG) glutathione in the chloroplasts. An increased GSH/GSSG ratio protected RCA from proteolytic digestion and increased the stability of redox-sensitive enzymes in the chloroplasts. These results strongly suggest that BRs are capable of regulating the glutathione redox state in the chloroplasts through the activation of the ascorbate–glutathione cycle. The resulting increase in the chloroplast thiol reduction state promotes CO 2 assimilation, at least in part, by enhancing the stability and activity of redox-sensitive photosynthetic enzymes through post-translational modifications.

  20. WHITE PANICLE3, a Novel Nucleus-Encoded Mitochondrial Protein, Is Essential for Proper Development and Maintenance of Chloroplasts and Mitochondria in Rice

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    Hongchang Li

    2018-06-01

    Full Text Available Mitochondria and chloroplasts are interacting organelles that play important roles in plant development. In addition to a small number proteins encoded by their own genomes, the majority of mitochondrial and chloroplast proteins are encoded in the cell nucleus and imported into the organelle. As a consequence, coordination between mitochondria, chloroplasts, and the nucleus is of crucial importance to plant cells. Variegated mutants are chloroplast-defective mutants and are considered to be ideal models for studying the intercommunication between these organelles. Here, we report the isolation of WHITE PANICLE3 (WP3, a nuclear gene involved in variegation, from a naturally occurring white panicle rice mutant. Disrupted expression of WP3 in the mutant leads to severe developmental defects in both chloroplasts and mitochondria, and consequently causes the appearance of white-striped leaves and white panicles in the mutant plants. Further investigation showed that WP3 encodes a protein most likely targeted to mitochondria and is specifically expressed in rice panicles. Interestingly, we demonstrate that the recessive white-panicle phenotype in the wp3 mutant is inherited in a typical Mendelian manner, while the white-striped leaf phenotype in wp3 is maternally inherited. Our data collectively suggest that the nucleus-encoded mitochondrial protein, WP3, plays an essential role in the regulation of chloroplast development in rice panicles by maintaining functional mitochondria. Therefore, the wp3 mutant is an excellent model in which to explore the communication between the nucleus, mitochondria, and chloroplasts in plant cells.

  1. Expression of Trichoderma reesei β-mannanase in tobacco chloroplasts and its utilization in lignocellulosic woody biomass hydrolysis.

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    Pankaj Agrawal

    Full Text Available Lignocellulosic ethanol offers a promising alternative to conventional fossil fuels. One among the major limitations in the lignocellulosic biomass hydrolysis is unavailability of efficient and environmentally biomass degrading technologies. Plant-based production of these enzymes on large scale offers a cost-effective solution. Cellulases, hemicellulases including mannanases and other accessory enzymes are required for conversion of lignocellulosic biomass into fermentable sugars. β-mannanase catalyzes endo-hydrolysis of the mannan backbone, a major constituent of woody biomass. In this study, the man1 gene encoding β-mannanase was isolated from Trichoderma reesei and expressed via the chloroplast genome. PCR and Southern hybridization analysis confirmed site-specific transgene integration into the tobacco chloroplast genomes and homoplasmy. Transplastomic plants were fertile and set viable seeds. Germination of seeds in the selection medium showed inheritance of transgenes into the progeny without any Mendelian segregation. Expression of endo-β-mannanase for the first time in plants facilitated its characterization for use in enhanced lignocellulosic biomass hydrolysis. Gel diffusion assay for endo-β-mannanase showed the zone of clearance confirming functionality of chloroplast-derived mannanase. Endo-β-mannanase expression levels reached up to 25 units per gram of leaf (fresh weight. Chloroplast-derived mannanase had higher temperature stability (40 °C to 70 °C and wider pH optima (pH 3.0 to 7.0 than E.coli enzyme extracts. Plant crude extracts showed 6-7 fold higher enzyme activity than E.coli extracts due to the formation of disulfide bonds in chloroplasts, thereby facilitating their direct utilization in enzyme cocktails without any purification. Chloroplast-derived mannanase when added to the enzyme cocktail containing a combination of different plant-derived enzymes yielded 20% more glucose equivalents from pinewood than the

  2. Protein profiling of plastoglobules in chloroplasts and chromoplasts. A surprising site for differential accumulation of metabolic enzymes.

    Science.gov (United States)

    Ytterberg, A Jimmy; Peltier, Jean-Benoit; van Wijk, Klaas J

    2006-03-01

    Plastoglobules (PGs) are oval or tubular lipid-rich structures present in all plastid types, but their specific functions are unclear. PGs contain quinones, alpha-tocopherol, and lipids and, in chromoplasts, carotenoids as well. It is not known whether PGs contain any enzymes or regulatory proteins. Here, we determined the proteome of PGs from chloroplasts of stressed and unstressed leaves of Arabidopsis (Arabidopsis thaliana) as well as from pepper (Capsicum annuum) fruit chromoplasts using mass spectrometry. Together, this showed that the proteome of chloroplast PGs consists of seven fibrillins, providing a protein coat and preventing coalescence of the PGs, and an additional 25 proteins likely involved in metabolism of isoprenoid-derived molecules (quinines and tocochromanols), lipids, and carotenoid cleavage. Four unknown ABC1 kinases were identified, possibly involved in regulation of quinone monooxygenases. Most proteins have not been observed earlier but have predicted N-terminal chloroplast transit peptides and lack transmembrane domains, consistent with localization in the PG lipid monolayer particles. Quantitative differences in PG composition in response to high light stress and degreening were determined by differential stable-isotope labeling using formaldehyde. More than 20 proteins were identified in the PG proteome of pepper chromoplasts, including four enzymes of carotenoid biosynthesis and several homologs of proteins observed in the chloroplast PGs. Our data strongly suggest that PGs in chloroplasts form a functional metabolic link between the inner envelope and thylakoid membranes and play a role in breakdown of carotenoids and oxidative stress defense, whereas PGs in chromoplasts are also an active site for carotenoid conversions.

  3. Protein Profiling of Plastoglobules in Chloroplasts and Chromoplasts. A Surprising Site for Differential Accumulation of Metabolic Enzymes1[W

    Science.gov (United States)

    Ytterberg, A. Jimmy; Peltier, Jean-Benoit; van Wijk, Klaas J.

    2006-01-01

    Plastoglobules (PGs) are oval or tubular lipid-rich structures present in all plastid types, but their specific functions are unclear. PGs contain quinones, α-tocopherol, and lipids and, in chromoplasts, carotenoids as well. It is not known whether PGs contain any enzymes or regulatory proteins. Here, we determined the proteome of PGs from chloroplasts of stressed and unstressed leaves of Arabidopsis (Arabidopsis thaliana) as well as from pepper (Capsicum annuum) fruit chromoplasts using mass spectrometry. Together, this showed that the proteome of chloroplast PGs consists of seven fibrillins, providing a protein coat and preventing coalescence of the PGs, and an additional 25 proteins likely involved in metabolism of isoprenoid-derived molecules (quinines and tocochromanols), lipids, and carotenoid cleavage. Four unknown ABC1 kinases were identified, possibly involved in regulation of quinone monooxygenases. Most proteins have not been observed earlier but have predicted N-terminal chloroplast transit peptides and lack transmembrane domains, consistent with localization in the PG lipid monolayer particles. Quantitative differences in PG composition in response to high light stress and degreening were determined by differential stable-isotope labeling using formaldehyde. More than 20 proteins were identified in the PG proteome of pepper chromoplasts, including four enzymes of carotenoid biosynthesis and several homologs of proteins observed in the chloroplast PGs. Our data strongly suggest that PGs in chloroplasts form a functional metabolic link between the inner envelope and thylakoid membranes and play a role in breakdown of carotenoids and oxidative stress defense, whereas PGs in chromoplasts are also an active site for carotenoid conversions. PMID:16461379

  4. Importance of post-translational modifications for functionality of a chloroplast-localized carbonic anhydrase (CAH1 in Arabidopsis thaliana.

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    Stefan Burén

    Full Text Available BACKGROUND: The Arabidopsis CAH1 alpha-type carbonic anhydrase is one of the few plant proteins known to be targeted to the chloroplast through the secretory pathway. CAH1 is post-translationally modified at several residues by the attachment of N-glycans, resulting in a mature protein harbouring complex-type glycans. The reason of why trafficking through this non-canonical pathway is beneficial for certain chloroplast resident proteins is not yet known. Therefore, to elucidate the significance of glycosylation in trafficking and the effect of glycosylation on the stability and function of the protein, epitope-labelled wild type and mutated versions of CAH1 were expressed in plant cells. METHODOLOGY/PRINCIPAL FINDINGS: Transient expression of mutant CAH1 with disrupted glycosylation sites showed that the protein harbours four, or in certain cases five, N-glycans. While the wild type protein trafficked through the secretory pathway to the chloroplast, the non-glycosylated protein formed aggregates and associated with the ER chaperone BiP, indicating that glycosylation of CAH1 facilitates folding and ER-export. Using cysteine mutants we also assessed the role of disulphide bridge formation in the folding and stability of CAH1. We found that a disulphide bridge between cysteines at positions 27 and 191 in the mature protein was required for correct folding of the protein. Using a mass spectrometric approach we were able to measure the enzymatic activity of CAH1 protein. Under circumstances where protein N-glycosylation is blocked in vivo, the activity of CAH1 is completely inhibited. CONCLUSIONS/SIGNIFICANCE: We show for the first time the importance of post-translational modifications such as N-glycosylation and intramolecular disulphide bridge formation in folding and trafficking of a protein from the secretory pathway to the chloroplast in higher plants. Requirements for these post-translational modifications for a fully functional native

  5. The complete chloroplast genome sequence of Ampelopsis: gene organization, comparative analysis and phylogenetic relationships to other angiosperms

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    Gurusamy eRaman

    2016-03-01

    Full Text Available Ampelopsis brevipedunculata is an economically important plant that belongs to the Vitaceae family of angiosperms. The phylogenetic placement of Vitaceae is still unresolved. Recent phylogenetic studies suggested that it should be placed in various alternative families including Caryophyllaceae, asteraceae, Saxifragaceae, Dilleniaceae, or with the rest of the rosid families. However, these analyses provided weak supportive results because they were based on only one of several genes. Accordingly, complete chloroplast genome sequences are required to resolve the phylogenetic relationships among angiosperms. Recent phylogenetic analyses based on the complete chloroplast genome sequence suggested strong support for the position of Vitaceae as the earliest diverging lineage of rosids and placed it as a sister to the remaining rosids. These studies also revealed relationships among several major lineages of angiosperms; however, they highlighted the significance of taxon sampling for obtaining accurate phylogenies. In the present study, we sequenced the complete chloroplast genome of A. brevipedunculata and used these data to assess the relationships among 32 angiosperms, including 18 taxa of rosids. The Ampelopsis chloroplast genome is 161,090 bp in length, and includes a pair of inverted repeats of 26,394 bp that are separated by small and large single copy regions of 19,036 bp and 89,266 bp, respectively. The gene content and order of Ampelopsis is identical to many other unrearranged angiosperm chloroplast genomes, including Vitis and tobacco. A phylogenetic tree constructed based on 70 protein-coding genes of 33 angiosperms showed that both Saxifragales and Vitaceae diverged from the rosid clade and formed two clades with 100% bootstrap value. The position of the Vitaceae is sister to Saxifragales, and both are the basal and earliest diverging lineages. Moreover, Saxifragales forms a sister clade to Vitaceae of rosids. Overall, the results of

  6. Impacts of high ATP supply from chloroplasts and mitochondria on the leaf metabolism of Arabidopsis thaliana

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    Chao eLiang

    2015-10-01

    Full Text Available Chloroplasts and mitochondria are the major ATP producing organelles in plant leaves. Arabidopsis thaliana purple acid phosphatase 2 (AtPAP2 is a phosphatase dually targeted to the outer membranes of both organelles and it plays a role in the import of selected nuclear-encoded proteins into these two organelles. Overexpression (OE of AtPAP2 in Arabidopsis thaliana accelerates plant growth and promotes flowering, seed yield and biomass at maturity. Measurement of ADP/ATP/NADP+/NADPH contents in the leaves of 20-day-old OE and wild-type lines at the end of night and at 1 and 8 h following illumination in a 16/8 h photoperiod revealed that the ATP levels and ATP/NADPH ratios were significantly increased in the OE line at all three time points. The AtPAP2 OE line is therefore a good model to investigate the impact of high energy on the global molecular status of Arabidopsis. In this study, transcriptome, proteome and metabolome profiles of the high ATP transgenic line were examined and compared with those of wild-type plants. A comparison of OE and WT at the end of the night provide valuable information on the impact of higher ATP output from mitochondria on plant physiology, as mitochondrial respiration is the major source of ATP in the dark in leaves. Similarly, comparison of OE and WT following illumination will provide information on the impact of higher energy output from chloroplasts on plant physiology. Overexpression of AtPAP2 was found to significantly affect the transcript and protein abundances of genes encoded by the two organellar genomes. For example, the protein abundances of many ribosomal proteins encoded by the chloroplast genome were higher in the AtPAP2 OE line under both light and dark conditions, while the protein abundances of multiple components of the photosynthetic complexes were lower. RNA-seq data also showed that the transcription of the mitochondrial genome is greatly affected by the availability of energy. These data

  7. Species delimitation and interspecific relationships of the genus Orychophragmus (Brassicaceae inferred from whole chloroplast genomes

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    Huan Hu

    2016-12-01

    Full Text Available IntroductionIt is rather difficult to delimit recently diverged species and construct their interspecific relationships because of insufficient informative variations of sampled DNA fragments (Schluter, 2000; Arnold, 2006. The genome-scale sequence variations were found to increase the phylogenetic resolutions of both high- and low-taxonomic groups (e.g., Yoder et al., 2013; Lamichhaney et al., 2015. It is still expensive to collect nuclear genome variations between species for most none-model genera without the reference genome. However, chloroplast genomes (plastome are relatively easy to be assembled to examine interspecific relationships for phylogenetic analyses, especially in addressing unresolved relationship at low taxonomic levels (Wu et al., 2010; Nock et al., 2011; Yang et al., 2013; Huang et al., 2014; Carbonell-Caballero et al., 2015. Plastomes are haploid with maternal inheritance in most angiosperms (Corriveau and Coleman, 1988; Zhang and Liu, 2003; Hagemann, 2004 and are highly conservative in gene order and genome structure with rare recombinations (Jansen et al., 2007; Moore et al., 2010. In this study, we aimed to examine species delimitation and interspecific relationships in Orychophragmus through assembling chloroplast genomes of multiple individuals of tentatively delimited species (Hu et al., 2015a. Orychophragmus is a small genus in the mustard family (Brassicaceae, Cruciferae distributed in northern, central, and southeastern China (Zhou et al., 2001. Its plants have been widely cultivated as ornamentals, vegetables, or source of seed oil (Sun et al., 2011. Despite controversial species delimitations in the genus (Zhou et al., 1987; Tan et al., 1998; Wu and Zhao, 2003; Al-Shehbaz and Yang, 2000; Zhou et al., 2001; Sun et al., 2012, our recent study based on nuclear (nr ITS sequence variations suggested the recognition of seven species (Hu et al., 2015a. Orychophragmus is sister to Sinalliaria, which is a genus endemic

  8. Chloroplast phylogenomic analyses resolve deep-level relationships of an intractable bamboo tribe Arundinarieae (poaceae).

    Science.gov (United States)

    Ma, Peng-Fei; Zhang, Yu-Xiao; Zeng, Chun-Xia; Guo, Zhen-Hua; Li, De-Zhu

    2014-11-01

    The temperate woody bamboos constitute a distinct tribe Arundinarieae (Poaceae: Bambusoideae) with high species diversity. Estimating phylogenetic relationships among the 11 major lineages of Arundinarieae has been particularly difficult, owing to a possible rapid radiation and the extremely low rate of sequence divergence. Here, we explore the use of chloroplast genome sequencing for phylogenetic inference. We sampled 25 species (22 temperate bamboos and 3 outgroups) for the complete genome representing eight major lineages of Arundinarieae in an attempt to resolve backbone relationships. Phylogenetic analyses of coding versus noncoding sequences, and of different regions of the genome (large single copy and small single copy, and inverted repeat regions) yielded no well-supported contradicting topologies but potential incongruence was found between the coding and noncoding sequences. The use of various data partitioning schemes in analysis of the complete sequences resulted in nearly identical topologies and node support values, although the partitioning schemes were decisively different from each other as to the fit to the data. Our full genomic data set substantially increased resolution along the backbone and provided strong support for most relationships despite the very short internodes and long branches in the tree. The inferred relationships were also robust to potential confounding factors (e.g., long-branch attraction) and received support from independent indels in the genome. We then added taxa from the three Arundinarieae lineages that were not included in the full-genome data set; each of these were sampled for more than 50% genome sequences. The resulting trees not only corroborated the reconstructed deep-level relationships but also largely resolved the phylogenetic placements of these three additional lineages. Furthermore, adding 129 additional taxa sampled for only eight chloroplast loci to the combined data set yielded almost identical

  9. Structural Basis for Redox Regulation of Cytoplasmic and Chloroplastic Triosephosphate Isomerases from Arabidopsis thaliana

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    Laura Margarita López-Castillo

    2016-12-01

    Full Text Available In plants triosephosphate isomerase (TPI interconverts glyceraldehyde 3-phosphate (G3P and dihydroxyacetone phosphate (DHAP during glycolysis, gluconeogenesis, and the Calvin-Benson cycle. The nuclear genome of land plants encodes two tpi genes, one gene product is located in the cytoplasm and the other is imported into the chloroplast. Herein we report the crystal structures of the TPIs from the vascular plant Arabidopsis thaliana (AtTPIs and address their enzymatic modulation by redox agents. Cytoplasmic TPI (cTPI and chloroplast TPI (pdTPI share more than 60% amino acid identity and assemble as (β-α8 dimers with high structural homology. cTPI and pdTPI harbor two and one accessible thiol groups per monomer respectively. cTPI and pdTPI present a cysteine at an equivalent structural position (C13 and C15 respectively and cTPI also contains a specific solvent accessible cysteine at residue 218 (cTPI-C218. Site directed mutagenesis of residues pdTPI-C15, cTPI-C13 and cTPI-C218 to serine substantially decreases enzymatic activity, indicating that the structural integrity of these cysteines is necessary for catalysis. AtTPIs exhibit differential responses to oxidative agents, cTPI is susceptible to oxidative agents such as diamide and H2O2, whereas pdTPI is resistant to inhibition. Incubation of AtTPIs with the sulfhydryl conjugating reagents methylmethane thiosulfonate (MMTS and glutathione inhibits enzymatic activity. However, the concentration necessary to inhibit pdTPI is at least two orders of magnitude higher than the concentration needed to inhibit cTPI. Western-blot analysis indicates that residues cTPI-C13, cTPI-C218, and pdTPI-C15 conjugate with glutathione. In summary, our data indicate that AtTPIs could be redox regulated by the derivatization of specific AtTPI cysteines (cTPI-C13 and pdTPI-C15 and cTPI-C218. Since AtTPIs have evolved by gene duplication, the higher resistance of pdTPI to redox agents may be an adaptive consequence to

  10. FtsZ-less prokaryotic cell division as well as FtsZ- and dynamin-less chloroplast and non-photosynthetic plastid division

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    Shin-Ya eMiyagishima

    2014-09-01

    Full Text Available The chloroplast division machinery is a mixture of a stromal FtsZ-based complex descended from a cyanobacterial ancestor of chloroplasts and a cytosolic dynamin-related protein (DRP 5B-based complex derived from the eukaryotic host. Molecular genetic studies have shown that each component of the division machinery is normally essential for normal chloroplast division. However, several exceptions have been found. In the absence of the FtsZ ring, nonphotosynthetic plastids are able to proliferate, likely by elongation and budding. Depletion of DRP5B impairs, but does not stop chloroplast division. Chloroplasts in glaucophytes, which possesses a peptidoglycan (PG layer, divide without DRP5B. Certain parasitic eukaryotes possess nonphotosynthetic plastids of secondary endosymbiotic origin, but neither FtsZ nor DRP5B is encoded in their genomes. Elucidation of the FtsZ- and/or DRP5B-less chloroplast division mechanism will lead to a better understanding of the function and evolution of the chloroplast division machinery and the finding of the as-yet-unknown mechanism that is likely involved in chloroplast division. Recent studies have shown that FtsZ was lost from a variety of prokaryotes, many of which lost PG by regressive evolution. In addition, even some of the FtsZ-bearing bacteria are able to divide when FtsZ and PG are depleted experimentally. In some cases, alternative mechanisms for cell division, such as budding by an increase of the cell surface-to-volume ratio, are proposed. Although PG is believed to have been lost from chloroplasts other than in glaucophytes, there is some indirect evidence for the existence of PG in chloroplasts. Such information is also useful for understanding how nonphotosynthetic plastids are able to divide in FtsZ-depleted cells and the reason for the retention of FtsZ in chloroplast division. Here we summarize information to facilitate analyses of FtsZ- and/or DRP5B-less chloroplast and nonphotosynthetic plastid

  11. The complete chloroplast genome sequence of Maddenia hypoleuca koehne (Prunoideae, Rosaceae).

    Science.gov (United States)

    Chen, Tao; Zhang, Jing; Liu, Yin; Wang, Hao; Wang, Juan; Chen, Qing; Tang, Hao-Ru; Wang, Xiao-Rong

    2016-11-01

    Maddenia hypoleuca Koehne belonging to family Rosaceae is a native species in China. The complete chloroplast (cp) genome was generated by de novo assembly using low coverage whole genome sequencing data and manual correction. The cp genome was 158 084 bp in length, with GC content of 36.63%. It exhibited a typical quadripartite structure: a pair of large inverted repeat regions (IRs, 26 246 bp each), a large single-copy region (LSC, 86 713 bp), and a small single-copy region (SSC, 18 879 bp). A total of 114 genes were predicted, which included 80 protein-coding genes, 30 tRNA genes, and four rRNA genes. Phylogenetic analysis indicated that M. hypoleuca is most closely related to Prunus padus within the Prunoideae subfamily, which conforms to the traditional classification.

  12. Proliferation of group II introns in the chloroplast genome of the green alga Oedocladium carolinianum (Chlorophyceae

    Directory of Open Access Journals (Sweden)

    Jean-Simon Brouard

    2016-10-01

    Full Text Available Background The chloroplast genome sustained extensive changes in architecture during the evolution of the Chlorophyceae, a morphologically and ecologically diverse class of green algae belonging to the Chlorophyta; however, the forces driving these changes are poorly understood. The five orders recognized in the Chlorophyceae form two major clades: the CS clade consisting of the Chlamydomonadales and Sphaeropleales, and the OCC clade consisting of the Oedogoniales, Chaetophorales, and Chaetopeltidales. In the OCC clade, considerable variations in chloroplast DNA (cpDNA structure, size, gene order, and intron content have been observed. The large inverted repeat (IR, an ancestral feature characteristic of most green plants, is present in Oedogonium cardiacum (Oedogoniales but is lacking in the examined members of the Chaetophorales and Chaetopeltidales. Remarkably, the Oedogonium 35.5-kb IR houses genes that were putatively acquired through horizontal DNA transfer. To better understand the dynamics of chloroplast genome evolution in the Oedogoniales, we analyzed the cpDNA of a second representative of this order, Oedocladium carolinianum. Methods The Oedocladium cpDNA was sequenced and annotated. The evolutionary distances separating Oedocladium and Oedogonium cpDNAs and two other pairs of chlorophycean cpDNAs were estimated using a 61-gene data set. Phylogenetic analysis of an alignment of group IIA introns from members of the OCC clade was performed. Secondary structures and insertion sites of oedogonialean group IIA introns were analyzed. Results The 204,438-bp Oedocladium genome is 7.9 kb larger than the Oedogonium genome, but its repertoire of conserved genes is remarkably similar and gene order differs by only one reversal. Although the 23.7-kb IR is missing the putative foreign genes found in Oedogonium, it contains sequences coding for a putative phage or bacterial DNA primase and a hypothetical protein. Intergenic sequences are 1.5-fold

  13. Membrane protein damage and repair: selective loss of a quinone-protein function in chloroplast membranes

    International Nuclear Information System (INIS)

    Kyle, D.J.; Ohad, I.; Arntzen, C.J.

    1984-01-01

    A loss of electron transport capacity in chloroplast membranes was induced by high-light intensities (photoinhibition). The primary site of inhibition was at the reducing side of photosystem II (PSII) with little damage to the oxidizing side or to the reaction center core of PSII. Addition of herbicides (atrazine or diuron) partially protected the membrane from photoinhibition; these compounds displace the bound plastoquinone (designated as Q/sub B/), which functions as the secondary electron acceptor on the reducing side of PSII. Loss of function of the 32-kilodalton Q/sub B/ apoprotein was demonstrated by a loss of binding sites for [ 14 C]atraazine. We suggest that quinone anions, which may interact with molecular oxygen to produce an oxygen radical, selectively damage the apoprotein of the secondary acceptor of PSII, thus rendering it inactive and thereby blocking photosynthetic electron flow under conditions of high photon flux densities. 21 references, 4 figures, 2 tables

  14. Enhanced hydrogen production by coupled system of Halobacterium halobium and chloroplast after entrapment within reverse micelles

    Energy Technology Data Exchange (ETDEWEB)

    Singh, A.; Dubey, R.S. [Banaras Hindhu University, Varanasi (India). Dept. of Biochemistry; Pandey, K.D. [Banaras Hindhu University, Varanasi (India). Dept. of Botany

    1999-08-01

    Reverse micelles were used for the enhanced rate of photoproduction of hydrogen using the coupled system of Halobacterium halobium and chloroplasts organelles. Different combinations of organic solvents and surfactants were used for generating reverse micelles. A several fold enhancement in the rate of H{sub 2} production was observed when the coupled system was entrapped within reverse micelles as compared to the aqueous suspension where no detectable H{sub 2} was produced. The coupled system immobilized in reverse micelles formed by sodium lauryl sulfate and carbon tetrachloride yielded maximum rate of H{sub 2} evolution. The optimum temperature for such hydrogen production was 40{sup o}C using light of 520-570 nm wavelength and 100 lux intensity. (author)

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

    Science.gov (United States)

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

    2014-01-01

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

  16. Identification and Analysis of the Chloroplast rpoC1 Gene Differentially Expressed in Wild Ginseng

    Directory of Open Access Journals (Sweden)

    Lee Kwang-Ho

    2012-06-01

    Full Text Available Panax ginseng is a well-known herbal medicine in traditional Asian medicine, and wild ginseng is widely accepted to be more active than cultivated ginseng in chemoprevention. However, little has actually been reported on the difference between wild ginseng and cultivated ginseng. Thus, to identify and analyze those differences, we used suppressive subtraction hybridization (SSH sequences with microarrays, realtime polymerase chain reaction (PCR, and reverse transcription PCRs (RT-PCRs. One of the clones isolated in this research was the chloroplast rpoC1 gene, a β subunit of RNA polymerase. Real-time RT-PCR results showed that the expression of the rpoC1 gene was significantly upregulated in wild ginseng as compared to cultivated ginseng, so, we conclude that the rpoC1 gene may be one of the important markers of wild ginseng.

  17. Modifications at the A-domain of the chloroplast import receptor Toc159.

    Science.gov (United States)

    Agne, Birgit; Kessler, Felix

    2010-11-01

    Two families of GTPases, the Toc34 and Toc159 GTPase families, take on the task of preprotein recognition at the translocon at the outer membrane of chloroplasts (TOC translocon). The major Toc159 family members have highly acidic N-terminal domains (A-domains) that are non-essential and so far have escaped functional characterization. But recently, interest in the role of the A-domain has strongly increased. The new data of three independent studies provide evidence that the Toc159 A-domain I) participates in preprotein selectivity, II) has typical features of intrinsically unfolded proteins and III) is highly phosphorylated and possibly released from the rest of the protein by a proteolytic event. This hints to a complex regulation of A-domain function that is important for the maintenance of the preprotein selectivity at the TOC translocons.

  18. Light Quality Affects Chloroplast Electron Transport Rates Estimated from Chl Fluorescence Measurements.

    Science.gov (United States)

    Evans, John R; Morgan, Patrick B; von Caemmerer, Susanne

    2017-10-01

    Chl fluorescence has been used widely to calculate photosynthetic electron transport rates. Portable photosynthesis instruments allow for combined measurements of gas exchange and Chl fluorescence. We analyzed the influence of spectral quality of actinic light on Chl fluorescence and the calculated electron transport rate, and compared this with photosynthetic rates measured by gas exchange in the absence of photorespiration. In blue actinic light, the electron transport rate calculated from Chl fluorescence overestimated the true rate by nearly a factor of two, whereas there was closer agreement under red light. This was consistent with the prediction made with a multilayer leaf model using profiles of light absorption and photosynthetic capacity. Caution is needed when interpreting combined measurements of Chl fluorescence and gas exchange, such as the calculation of CO2 partial pressure in leaf chloroplasts. © Crown copyright 2017.

  19. Photosynthesis and chloroplast genes are involved in water-use efficiency in common bean.

    Science.gov (United States)

    Ruiz-Nieto, Jorge E; Aguirre-Mancilla, César L; Acosta-Gallegos, Jorge A; Raya-Pérez, Juan C; Piedra-Ibarra, Elías; Vázquez-Medrano, Josefina; Montero-Tavera, Victor

    2015-01-01

    A recent proposal to mitigate the effects of climatic change and reduce water consumption in agriculture is to develop cultivars with high water-use efficiency. The aims of this study were to characterize this trait as a differential response mechanism to water-limitation in two bean cultivars contrasting in their water stress tolerance, to isolate and identify gene fragments related to this response in a model cultivar, as well as to evaluate transcription levels of genes previously identified. Keeping CO2 assimilation through a high photosynthesis rate under limited conditions was the physiological response which allowed the cultivar model to maintain its growth and seed production with less water. Chloroplast genes stood out among identified genetic elements, which confirmed the importance of photosynthesis in such response. ndhK, rpoC2, rps19, rrn16, ycf1 and ycf2 genes were expressed only in response to limited water availability. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  20. In silico analysis of Simple Sequence Repeats from chloroplast genomes of Solanaceae species

    Directory of Open Access Journals (Sweden)

    Evandro Vagner Tambarussi

    2009-01-01

    Full Text Available The availability of chloroplast genome (cpDNA sequences of Atropa belladonna, Nicotiana sylvestris, N.tabacum, N. tomentosiformis, Solanum bulbocastanum, S. lycopersicum and S. tuberosum, which are Solanaceae species,allowed us to analyze the organization of cpSSRs in their genic and intergenic regions. In general, the number of cpSSRs incpDNA ranged from 161 in S. tuberosum to 226 in N. tabacum, and the number of intergenic cpSSRs was higher than geniccpSSRs. The mononucleotide repeats were the most frequent in studied species, but we also identified di-, tri-, tetra-, pentaandhexanucleotide repeats. Multiple alignments of all cpSSRs sequences from Solanaceae species made the identification ofnucleotide variability possible and the phylogeny was estimated by maximum parsimony. Our study showed that the plastomedatabase can be exploited for phylogenetic analysis and biotechnological approaches.

  1. Functional proteomics of barley and barley chloroplasts – strategies, methods and perspectives

    DEFF Research Database (Denmark)

    Petersen, Jørgen; Rogowska-Wrzesinska, Adelina; Jensen, Ole Nørregaard

    2013-01-01

    Barley (Hordeum vulgare) is an important cereal grain that is used in a range of products for animal and human consumption. Crop yield and seed quality has been optimized during decades by plant breeding programs supported by biotechnology and molecular biology techniques. The recently completed...... whole-genome sequencing of barley revealed approximately 26,100 open reading frames, which provides a foundation for detailed molecular studies of barley by functional genomics and proteomics approaches. Such studies will provide further insights into the mechanisms of, for example, drought and stress...... tolerance, micronutrient utilization, and photosynthesis in barley. In the present review we present the current state of proteomics research for investigations of barley chloroplasts, i.e., the organelle that contain the photosynthetic apparatus in the plant. We describe several different proteomics...

  2. Chloroplastic and stomatal aspects of ozone-induced reduction of net photosynthesis in plants

    Energy Technology Data Exchange (ETDEWEB)

    Torsethaugen, Gro

    1998-09-01

    The present thesis relates to ozone-induced reduction of photosynthesis in plants. As a photochemical oxidant O{sub 3} is formed by the interaction of hydrocarbons, nitrogen oxides and oxygen in sunlight. Ozone (O{sub 3}) is the most phytotoxic of all the air pollutants and is known to reduce plant growth and net photosynthesis, cause stomatal closure, induce visible injury, accelerate senescence and induce or inhibit transcription of a variety of genes with a corresponding increase/decrease in protein products. The underlying cellular mechanisms for many of these changes are unknown. Following fields are investigated: Ozone-induced reduction of net photosynthesis; ozone and the photosynthetic apparatus in the chloroplasts; ozone and stomata; ozone effects on plant membranes; protection against ozone injury in plants. 249 refs., 22 figs., 4 tabs.

  3. Complete chloroplast genome sequence of a major economic species, Ziziphus jujuba (Rhamnaceae).

    Science.gov (United States)

    Ma, Qiuyue; Li, Shuxian; Bi, Changwei; Hao, Zhaodong; Sun, Congrui; Ye, Ning

    2017-02-01

    Ziziphus jujuba is an important woody plant with high economic and medicinal value. Here, we analyzed and characterized the complete chloroplast (cp) genome of Z. jujuba, the first member of the Rhamnaceae family for which the chloroplast genome sequence has been reported. We also built a web browser for navigating the cp genome of Z. jujuba ( http://bio.njfu.edu.cn/gb2/gbrowse/Ziziphus_jujuba_cp/ ). Sequence analysis showed that this cp genome is 161,466 bp long and has a typical quadripartite structure of large (LSC, 89,120 bp) and small (SSC, 19,348 bp) single-copy regions separated by a pair of inverted repeats (IRs, 26,499 bp). The sequence contained 112 unique genes, including 78 protein-coding genes, 30 transfer RNAs, and four ribosomal RNAs. The genome structure, gene order, GC content, and codon usage are similar to other typical angiosperm cp genomes. A total of 38 tandem repeats, two forward repeats, and three palindromic repeats were detected in the Z. jujuba cp genome. Simple sequence repeat (SSR) analysis revealed that most SSRs were AT-rich. The homopolymer regions in the cp genome of Z. jujuba were verified and manually corrected by Sanger sequencing. One-third of mononucleotide repeats were found to be erroneously sequenced by the 454 pyrosequencing, which resulted in sequences of 1-4 bases shorter than that by the Sanger sequencing. Analyzing the cp genome of Z. jujuba revealed that the IR contraction and expansion events resulted in ycf1 and rps19 pseudogenes. A phylogenetic analysis based on 64 protein-coding genes showed that Z. jujuba was closely related to members of the Elaeagnaceae family, which will be helpful for phylogenetic studies of other Rosales species. The complete cp genome sequence of Z. jujuba will facilitate population, phylogenetic, and cp genetic engineering studies of this economic plant.

  4. Complete chloroplast genome of the multifunctional crop globe artichoke and comparison with other Asteraceae.

    Science.gov (United States)

    Curci, Pasquale L; De Paola, Domenico; Danzi, Donatella; Vendramin, Giovanni G; Sonnante, Gabriella

    2015-01-01

    With over 20,000 species, Asteraceae is the second largest plant family. High-throughput sequencing of nuclear and chloroplast genomes has allowed for a better understanding of the evolutionary relationships within large plant families. Here, the globe artichoke chloroplast (cp) genome was obtained by a combination of whole-genome and BAC clone high-throughput sequencing. The artichoke cp genome is 152,529 bp in length, consisting of two single-copy regions separated by a pair of inverted repeats (IRs) of 25,155 bp, representing the longest IRs found in the Asteraceae family so far. The large (LSC) and the small (SSC) single-copy regions span 83,578 bp and 18,641 bp, respectively. The artichoke cp sequence was compared to the other eight Asteraceae complete cp genomes available, revealing an IR expansion at the SSC/IR boundary. This expansion consists of 17 bp of the ndhF gene generating an overlap between the ndhF and ycf1 genes. A total of 127 cp simple sequence repeats (cpSSRs) were identified in the artichoke cp genome, potentially suitable for future population studies in the Cynara genus. Parsimony-informative regions were evaluated and allowed to place a Cynara species within the Asteraceae family tree. The eight most informative coding regions were also considered and tested for "specific barcode" purpose in the Asteraceae family. Our results highlight the usefulness of cp genome sequencing in exploring plant genome diversity and retrieving reliable molecular resources for phylogenetic and evolutionary studies, as well as for specific barcodes in plants.

  5. The hydrogen peroxide-sensitive proteome of the chloroplast in vitro and in vivo

    Directory of Open Access Journals (Sweden)

    Meenakumari eMuthuramalingam

    2013-03-01

    Full Text Available Hydrogen peroxide (H2O2 evolves during cellular metabolism and accumulates under various stresses causing serious redox imbalances. Many proteomics studies aiming to identify proteins sensitive to H2O2 used concentrations that were above the physiological range. Here the chloroplast proteins were subjected to partial oxidation by exogenous addition of H2O2 equivalent to 10% of available protein thiols which allowed for the identification of the primary targets of oxidation. The chosen redox proteomic approach employed differential labeling of non-oxidized and oxidized thiols using sequential alkylation with NEM and biotin maleimide. The in vitro identified proteins are involved in carbohydrate metabolism, photosynthesis, redox homeostasis and nitrogen assimilation. By using methyl viologen that induces oxidative stress in vivo, mostly the same primary targets of oxidation were identified and several oxidation sites were annotated. RubisCO was a primary oxidation target. Due to its high abundance, RubisCO is suggested to act as a chloroplast redox buffer to maintain a suitable redox state, even in the presence of increased ROS release. 2-Cys Prxs undergo redox-dependent modifications and play important roles in antioxidant defense and signaling. The identification of 2-Cys Prx was expected based on its high affinity to H2O2 and is considered as a proof of concept for the approach. Targets of Trx, such as phosphoribulokinase, glyceraldehyde-3-phosphate dehydrogenase (GAPDH, transketolase and sedoheptulose-1,7-bisphosphatase have at least one regulatory disulfide bridge which supports the conclusion that the identified proteins undergo reversible thiol oxidation. In conclusion, the presented approach enabled the identification of early targets of H2O2 oxidation within the cellular proteome under physiological experimental conditions.

  6. Effective plague vaccination via oral delivery of plant cells expressing F1-V antigens in chloroplasts.

    Science.gov (United States)

    Arlen, Philip A; Singleton, Michael; Adamovicz, Jeffrey J; Ding, Yi; Davoodi-Semiromi, Abdolreza; Daniell, Henry

    2008-08-01

    The chloroplast bioreactor is an alternative to fermentation-based systems for production of vaccine antigens and biopharmaceuticals. We report here expression of the plague F1-V fusion antigen in chloroplasts. Site-specific transgene integration and homoplasmy were confirmed by PCR and Southern blotting. Mature leaves showed the highest level of transgene expression on the third day of continuous illumination, with a maximum level of 14.8% of the total soluble protein. Swiss Webster mice were primed with adjuvant-containing subcutaneous (s.c.) doses of F1-V and then boosted with either adjuvanted s.c. doses (s.c. F1-V mice) or unadjuvanted oral doses (oral F1-V mice). Oral F1-V mice had higher prechallenge serum immunoglobulin G1 (IgG1) titers than s.c. F1-V mice. The corresponding serum levels of antigen-specific IgG2a and IgA were 2 and 3 orders of magnitude lower, respectively. After vaccination, mice were exposed to an inhaled dose of 1.02 x 10(6) CFU of aerosolized Yersinia pestis CO92 (50% lethal dose, 6.8 x 10(4) CFU). All control animals died within 3 days. F1-V given s.c. (with adjuvant) protected 33% of the immunized mice, while 88% of the oral F1-V mice survived aerosolized Y. pestis challenge. A comparison of splenic Y. pestis CFU counts showed that there was a 7- to 10-log reduction in the mean bacterial burden in survivors. Taken together, these data indicate that oral booster doses effectively elicit protective immune responses in vivo. In addition, this is the first report of a plant-derived oral vaccine that protected animals from live Y. pestis challenge, bringing the likelihood of lower-cost vaccines closer to reality.

  7. Inclusion of chloroplast genes that have undergone expansion misleads phylogenetic reconstruction in the Chlorophyta.

    Science.gov (United States)

    Novis, Phil M; Smissen, Rob; Buckley, Thomas R; Gopalakrishnan, Kishore; Visnovsky, Gabriel

    2013-11-01

    Chlorophytes comprise a substantial proportion of green plant diversity. However, sister-group relationships and circumscription of the classes Chlorophyceae, Trebouxiophyceae, and Ulvophyceae have been problematic to resolve. Some analyses support a sister relationship between the trebouxiophycean Leptosira and chlorophyceans, potentially altering the circumscription of two classes, also supported by a shared fragmentation in the chloroplast gene rpoB. We sought to determine whether the latter is a synapomorphy or whether the supporting analyses are vulnerable to systematic bias. We sequenced a portion of rpoB spanning the fragmented region in strains for which it had not previously been sampled: four Chlorophyceae, six counterclockwise (CCW) group (ulvophyceans and trebouxiophyceans) and one streptophyte. We then explored the effect of subsampling proteins and taxa on phylogenetic reconstruction from a data set of 41 chloroplast proteins. None of the CCW or streptophyte strains possessed the split in rpoB, including inferred near relatives of Leptosira, but it was found in all chlorophycean strains. We reconstructed alternative phylogenies (Leptosira + Chlorophyceae and Leptosira + Chlorellales) using two different protein groups (Rpo and Rps), both subject to coding-region expansion. A conserved region of RpoB remained suitable for analysis of more recent divergences. The Rps sequences can explain earlier findings linking Leptosira with the Chlorophyceae and should be excluded from phylogenetic analyses attempting to resolve deep nodes because their expansion violates the assumptions of substitution models. We reaffirm that Leptosira is a trebouxiophycean and that fragmentation of rpoB has occurred at least twice in chlorophyte evolution.

  8. Genetic population structure of the desert shrub species lycium ruthenicum inferred from chloroplast dna

    International Nuclear Information System (INIS)

    Chen, H.; Yonezawa, T.

    2014-01-01

    Lycium ruthenicum (Solananeae), a spiny shrub mostly distributed in the desert regions of north and northwest China, has been shown to exhibit high tolerance to the extreme environment. In this study, the phylogeography and evolutionary history of L. ruthenicum were examined, on the basis of 80 individuals from eight populations. Using the sequence variations of two spacer regions of chloroplast DNA (trnH-psbA and rps16-trnK) , the absence of a geographic component in the chloroplast DNA genetic structure was identified (GST = 0.351, NST = 0.304, NST< GST), which was consisted with the result of SAMOVA, suggesting weak phylogeographic structure of this species. Phylogenetic and network analyses showed that a total of 10 haplotypes identified in the present study clustered into two clades, in which clade I harbored the ancestral haplotypes that inferred two independent glacial refugia in the middle of Qaidam Basin and the western Inner Mongolia. The existence of regional evolutionary differences was supported by GENETREE, which revealed that one of the population in Qaidam Basin and the two populations in Tarim Basin had experienced rapid expansion, and the other populations retained relatively stable population size during the Pleistocene . Given the results of long-term gene flow and pairwise differences, strong gene flow was insufficient to reduce the genetic differentiation among populations or within populations, probably due to the genetic composition containing a common haplotype and the high number of private haplotypes fixed for most of the population. The divergence times of different lineages were consistent with the rapid uplift phases of the Qinghai-Tibetan Plateau and the initiation and expansion of deserts in northern China, suggesting that the origin and evolution of L. ruthenicum were strongly influenced by Quaternary environment changes. (author)

  9. A comparison of chloroplast genome sequences in Aconitum (Ranunculaceae: a traditional herbal medicinal genus

    Directory of Open Access Journals (Sweden)

    Hanghui Kong

    2017-11-01

    Full Text Available The herbal medicinal genus Aconitum L., belonging to the Ranunculaceae family, represents the earliest diverging lineage within the eudicots. It currently comprises of two subgenera, A. subgenus Lycoctonum and A. subg. Aconitum. The complete chloroplast (cp genome sequences were characterized in three species: A. angustius, A. finetianum, and A. sinomontanum in subg. Lycoctonum and compared to other Aconitum species to clarify their phylogenetic relationship and provide molecular information for utilization of Aconitum species particularly in Eastern Asia. The length of the chloroplast genome sequences were 156,109 bp in A. angustius, 155,625 bp in A. finetianum and 157,215 bp in A. sinomontanum, with each species possessing 126 genes with 84 protein coding genes (PCGs. While genomic rearrangements were absent, structural variation was detected in the LSC/IR/SSC boundaries. Five pseudogenes were identified, among which Ψrps19 and Ψycf1 were in the LSC/IR/SSC boundaries, Ψrps16 and ΨinfA in the LSC region, and Ψycf15 in the IRb region. The nucleotide variability (Pi of Aconitum was estimated to be 0.00549, with comparably higher variations in the LSC and SSC than the IR regions. Eight intergenic regions were revealed to be highly variable and a total of 58–62 simple sequence repeats (SSRs were detected in all three species. More than 80% of SSRs were present in the LSC region. Altogether, 64.41% and 46.81% of SSRs are mononucleotides in subg. Lycoctonum and subg. Aconitum, respectively, while a higher percentage of di-, tri-, tetra-, and penta- SSRs were present in subg. Aconitum. Most species of subg. Aconitum in Eastern Asia were first used for phylogenetic analyses. The availability of the complete cp genome sequences of these species in subg. Lycoctonum will benefit future phylogenetic analyses and aid in germplasm utilization in Aconitum species.

  10. A comparison of chloroplast genome sequences in Aconitum (Ranunculaceae): a traditional herbal medicinal genus.

    Science.gov (United States)

    Kong, Hanghui; Liu, Wanzhen; Yao, Gang; Gong, Wei

    2017-01-01

    The herbal medicinal genus Aconitum L., belonging to the Ranunculaceae family, represents the earliest diverging lineage within the eudicots. It currently comprises of two subgenera, A . subgenus Lycoctonum and A . subg. Aconitum . The complete chloroplast (cp) genome sequences were characterized in three species: A. angustius , A. finetianum , and A. sinomontanum in subg. Lycoctonum and compared to other Aconitum species to clarify their phylogenetic relationship and provide molecular information for utilization of Aconitum species particularly in Eastern Asia. The length of the chloroplast genome sequences were 156,109 bp in A. angustius , 155,625 bp in A. finetianum and 157,215 bp in A. sinomontanum , with each species possessing 126 genes with 84 protein coding genes (PCGs). While genomic rearrangements were absent, structural variation was detected in the LSC/IR/SSC boundaries. Five pseudogenes were identified, among which Ψ rps 19 and Ψ ycf 1 were in the LSC/IR/SSC boundaries, Ψ rps 16 and Ψ inf A in the LSC region, and Ψ ycf 15 in the IRb region. The nucleotide variability ( Pi ) of Aconitum was estimated to be 0.00549, with comparably higher variations in the LSC and SSC than the IR regions. Eight intergenic regions were revealed to be highly variable and a total of 58-62 simple sequence repeats (SSRs) were detected in all three species. More than 80% of SSRs were present in the LSC region. Altogether, 64.41% and 46.81% of SSRs are mononucleotides in subg. Lycoctonum and subg. Aconitum , respectively, while a higher percentage of di-, tri-, tetra-, and penta- SSRs were present in subg. Aconitum . Most species of subg. Aconitum in Eastern Asia were first used for phylogenetic analyses. The availability of the complete cp genome sequences of these species in subg. Lycoctonum will benefit future phylogenetic analyses and aid in germplasm utilization in Aconitum species.

  11. Testing the Role of the N-Terminal Tail of D1 in the Maintenance of Photosystem II in Tobacco Chloroplasts

    Czech Academy of Sciences Publication Activity Database

    Michoux, F.; Ahmad, N.; Wei, Z-Y.; Belgio, Erica; Ruban, A.V.; Nixon, P.

    2016-01-01

    Roč. 7, 21 June (2016), s. 844 ISSN 1664-462X Institutional support: RVO:61388971 Keywords : D1 degradation * PSII repair * chloroplast transformation Subject RIV: EF - Botanics Impact factor: 4.298, year: 2016

  12. Chloroplast genes as genetic markers for inferring patterns of change, maternal ancestry and phylogenetic relationships among Eleusine species.

    Science.gov (United States)

    Agrawal, Renuka; Agrawal, Nitin; Tandon, Rajesh; Raina, Soom Nath

    2014-01-01

    Assessment of phylogenetic relationships is an important component of any successful crop improvement programme, as wild relatives of the crop species often carry agronomically beneficial traits. Since its domestication in East Africa, Eleusine coracana (2n = 4x = 36), a species belonging to the genus Eleusine (x = 8, 9, 10), has held a prominent place in the semi-arid regions of India, Nepal and Africa. The patterns of variation between the cultivated and wild species reported so far and the interpretations based upon them have been considered primarily in terms of nuclear events. We analysed, for the first time, the phylogenetic relationship between finger millet (E. coracana) and its wild relatives by species-specific chloroplast deoxyribonucleic acid (cpDNA) polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and chloroplast simple sequence repeat (cpSSR) markers/sequences. Restriction fragment length polymorphism of the seven amplified chloroplast genes/intergenic spacers (trnK, psbD, psaA, trnH-trnK, trnL-trnF, 16S and trnS-psbC), nucleotide sequencing of the chloroplast trnK gene and chloroplast microsatellite polymorphism were analysed in all nine known species of Eleusine. The RFLP of all seven amplified chloroplast genes/intergenic spacers and trnK gene sequences in the diploid (2n = 16, 18, 20) and allotetraploid (2n = 36, 38) species resulted in well-resolved phylogenetic trees with high bootstrap values. Eleusine coracana, E. africana, E. tristachya, E. indica and E. kigeziensis did not show even a single change in restriction site. Eleusine intermedia and E. floccifolia were also shown to have identical cpDNA fragment patterns. The cpDNA diversity in Eleusine multiflora was found to be more extensive than that of the other eight species. The trnK gene sequence data complemented the results obtained by PCR-RFLP. The maternal lineage of all three allotetraploid species (AABB, AADD) was the same, with E. indica being the

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

    Science.gov (United States)

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

    2016-08-01

    Calcium is important for chloroplast, not only in its photosynthetic but also nonphotosynthetic functions. Multiple Ca(2+)/H(+) transporters and channels have been described and studied in the plasma membrane and organelle membranes of plant cells; however, the molecular identity and physiological roles of chloroplast Ca(2+)/H(+) antiporters have remained unknown. Here we report the identification and characterization of a member of the UPF0016 family, CCHA1 (a chloroplast-localized potential Ca(2+)/H(+) antiporter), in Arabidopsis thaliana. We observed that the ccha1 mutant plants developed pale green leaves and showed severely stunted growth along with impaired photosystem II (PSII) function. CCHA1 localizes to the chloroplasts, and the levels of the PSII core subunits and the oxygen-evolving complex were significantly decreased in the ccha1 mutants compared with the wild type. In high Ca(2+) concentrations, Arabidopsis CCHA1 partially rescued the growth defect of yeast gdt1Δ null mutant, which is defective in a Ca(2+)/H(+) antiporter. The ccha1 mutant plants also showed significant sensitivity to high concentrations of CaCl2 and MnCl2, as well as variation in pH. Taken these results together, we propose that CCHA1 might encode a putative chloroplast-localized Ca(2+)/H(+) antiporter with critical functions in the regulation of PSII and in chloroplast Ca(2+) and pH homeostasis in Arabidopsis. Copyright © 2016 The Author. Published by Elsevier Inc. All rights reserved.

  14. The promoter of the Arabidopsis thaliana plastocyanin gene contains a far upstream enhancer-like element involved in chloroplast-dependent expression.

    Science.gov (United States)

    Vorst, O; Kock, P; Lever, A; Weterings, B; Weisbeek, P; Smeekens, S

    1993-12-01

    Plastocyanin is part of the photosynthetic electron transport chain in the chloroplast and is encoded in the nucleus. Expression of the Arabidopsis thaliana plastocyanin gene is organ specific: high mRNA levels are observed in young green parts of the plant. Furthermore, expression is dependent on the presence of light and functional chloroplasts. When grown in the presence of norflurazon under white light conditions, resulting in the photo-oxidative destruction of the chloroplast, plastocyanin mRNA levels are strongly reduced. A -1579 to -9 promoter fragment confers light-regulated and chloroplast-dependent expression to the beta-glucuronidase reporter gene in transgenic tobacco plants. This suggests that regulation takes place at the level of transcription. A plastocyanin promoter deletion series ranging from -1579 to -121 which was also tested in tobacco, revealed the presence of a strong positive regulating element (PRE) in the -1579 to -705 region. Deletion of this part of the promoter resulted in a approximately 100-fold reduction of GUS expression as measured in mature leaves. Surprisingly, this enhancer-like element was capable of stimulating transcription from a position downstream of its reporter. Moreover, it could also activate a truncated CaMV 35S promoter. Deletion of this element coincides with the loss of chloroplast-dependency of reporter gene expression, as judged by norflurazon treatment of transgenic seedlings. So, the activity of the PRE itself might depend on the presence of functional chloroplasts.

  15. Complete chloroplast genome sequence of green foxtail (Setaria viridis), a promising model system for C4 photosynthesis.

    Science.gov (United States)

    Wang, Shuo; Gao, Li-Zhi

    2016-09-01

    The complete chloroplast genome of green foxtail (Setaria viridis), a promising model system for C4 photosynthesis, is first reported in this study. The genome harbors a large single copy (LSC) region of 81 016 bp and a small single copy (SSC) region of 12 456  bp separated by a pair of inverted repeat (IRa and IRb) regions of 22 315 bp. GC content is 38.92%. The proportion of coding sequence is 57.97%, comprising of 111 (19 duplicated in IR regions) unique genes, 71 of which are protein-coding genes, four are rRNA genes, and 36 are tRNA genes. Phylogenetic analysis indicated that S. viridis was clustered with its cultivated species S. italica in the tribe Paniceae of the family Poaceae. This newly determined chloroplast genome will provide valuable genetic resources to assist future studies on C4 photosynthesis in grasses.

  16. Abscisic acid refines the synthesis of chloroplast proteins in maize (Zea mays in response to drought and light.

    Directory of Open Access Journals (Sweden)

    Xiuli Hu

    Full Text Available To better understand abscisic acid (ABA regulation of the synthesis of chloroplast proteins in maize (Zea mays L. in response to drought and light, we compared leaf proteome differences between maize ABA-deficient mutant vp5 and corresponding wild-type Vp5 green and etiolated seedlings exposed to drought stress. Proteins extracted from the leaves of Vp5 and vp5 seedlings were used for two-dimensional electrophoresis (2-DE and subsequent matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF mass spectrometry (MS. After Coomassie brilliant blue staining, approximately 450 protein spots were reproducibly detected on 2-DE gels. A total of 36 differentially expressed protein spots in response to drought and light were identified using MALDI-TOF MS and their subcellular localization was determined based on the annotation of reviewed accession in UniProt Knowledgebase and the software prediction. As a result, corresponding 13 proteins of the 24 differentially expressed protein spots were definitely localized in chloroplasts and their expression was in an ABA-dependent way, including 6 up-regulated by both drought and light, 5 up-regulated by drought but down-regulated by light, 5 up-regulated by light but down-regulated by drought; 5 proteins down-regulated by drought were mainly those involved in photosynthesis and ATP synthesis. Thus, the results in the present study supported the vital role of ABA in regulating the synthesis of drought- and/or light-induced proteins in maize chloroplasts and would facilitate the functional characterization of ABA-induced chloroplast proteins in C(4 plants.

  17. Distinctive Architecture of the Chloroplast Genome in the Chlorodendrophycean Green Algae Scherffelia dubia and Tetraselmis sp. CCMP 881.

    Science.gov (United States)

    Turmel, Monique; de Cambiaire, Jean-Charles; Otis, Christian; Lemieux, Claude

    2016-01-01

    The Chlorodendrophyceae is a small class of green algae belonging to the core Chlorophyta, an assemblage that also comprises the Pedinophyceae, Trebouxiophyceae, Ulvophyceae and Chlorophyceae. Here we describe for the first time the chloroplast genomes of chlorodendrophycean algae (Scherffelia dubia, 137,161 bp; Tetraselmis sp. CCMP 881, 100,264 bp). Characterized by a very small single-copy (SSC) region devoid of any gene and an unusually large inverted repeat (IR), the quadripartite structures of the Scherffelia and Tetraselmis genomes are unique among all core chlorophytes examined thus far. The lack of genes in the SSC region is offset by the rich and atypical gene complement of the IR, which includes genes from the SSC and large single-copy regions of prasinophyte and streptophyte chloroplast genomes having retained an ancestral quadripartite structure. Remarkably, seven of the atypical IR-encoded genes have also been observed in the IRs of pedinophycean and trebouxiophycean chloroplast genomes, suggesting that they were already present in the IR of the common ancestor of all core chlorophytes. Considering that the relationships among the main lineages of the core Chlorophyta are still unresolved, we evaluated the impact of including the Chlorodendrophyceae in chloroplast phylogenomic analyses. The trees we inferred using data sets of 79 and 108 genes from 71 chlorophytes indicate that the Chlorodendrophyceae is a deep-diverging lineage of the core Chlorophyta, although the placement of this class relative to the Pedinophyceae remains ambiguous. Interestingly, some of our phylogenomic trees together with our comparative analysis of gene order data support the monophyly of the Trebouxiophyceae, thus offering further evidence that the previously observed affiliation between the Chlorellales and Pedinophyceae is the result of systematic errors in phylogenetic reconstruction.

  18. The complete structure of the chloroplast 70S ribosome in complex with translation factor pY.

    Science.gov (United States)

    Bieri, Philipp; Leibundgut, Marc; Saurer, Martin; Boehringer, Daniel; Ban, Nenad

    2017-02-15

    Chloroplasts are cellular organelles of plants and algae that are responsible for energy conversion and carbon fixation by the photosynthetic reaction. As a consequence of their endosymbiotic origin, they still contain their own genome and the machinery for protein biosynthesis. Here, we present the atomic structure of the chloroplast 70S ribosome prepared from spinach leaves and resolved by cryo-EM at 3.4 Å resolution. The complete structure reveals the features of the 4.5S rRNA, which probably evolved by the fragmentation of the 23S rRNA, and all five plastid-specific ribosomal proteins. These proteins, required for proper assembly and function of the chloroplast translation machinery, bind and stabilize rRNA including regions that only exist in the chloroplast ribosome. Furthermore, the structure reveals plastid-specific extensions of ribosomal proteins that extensively remodel the mRNA entry and exit site on the small subunit as well as the polypeptide tunnel exit and the putative binding site of the signal recognition particle on the large subunit. The translation factor pY, involved in light- and temperature-dependent control of protein synthesis, is bound to the mRNA channel of the small subunit and interacts with 16S rRNA nucleotides at the A-site and P-site, where it protects the decoding centre and inhibits translation by preventing tRNA binding. The small subunit is locked by pY in a non-rotated state, in which the intersubunit bridges to the large subunit are stabilized. © 2016 The Authors. Published under the terms of the CC BY NC ND 4.0 license.

  19. Inhibition of light modulation of chloroplast enzyme activity by sulfite. One of the lethal effects of SO/sub 2/

    Energy Technology Data Exchange (ETDEWEB)

    Anderson, L E; Duggan, J X

    1977-01-01

    The capacity of a particulate pea (Pisum sativum L.) leaf chloroplast system for light-modulation of enzyme activity is diminished by brief exposure to sodium sulfite and, when intact seedlings are exposed to atmospheric SO/sub 2/, the same system is inactivated. The destructive effect of this pollutant on green plants may therefore be due to disruption of the mechanism for control of carbon dioxide fixation.

  20. Hypergravity of 10 g Changes Plant Growth, Anatomy, Chloroplast Size, and Photosynthesis in the Moss Physcomitrella patens

    Science.gov (United States)

    Takemura, Kaori; Watanabe, Rina; Kameishi, Ryuji; Sakaguchi, Naoya; Kamachi, Hiroyuki; Kume, Atsushi; Karahara, Ichirou; Hanba, Yuko T.; Fujita, Tomomichi

    2017-12-01

    The photosynthetic and anatomical responses of bryophytes to changes in gravity will provide crucial information for estimating how these plant traits evolved to adapt to changes in gravity in land plant history. We performed long-term hypergravity experiments at 10 g for 4 and 8 weeks using the moss Physcomitrella patens with two centrifuges equipped with lighting systems that enable long-term plant growth under hypergravity with irradiance. The aims of this study are (1) to quantify changes in the anatomy and morphology of P. patens, and (2) to analyze the post-effects of hypergravity on photosynthesis by P. patens in relation to these changes. We measured photosynthesis by P. patens for a population of gametophores (e.g., canopy) in Petri dishes and plant culture boxes. Gametophore numbers increased by 9% for a canopy of P. patens, with 24-27% increases in chloroplast sizes (diameter and thickness) in leaf cells. In a canopy of P. patens, the area-based photosynthesis rate ( A canopy) was increased by 57% at 10 g. The increase observed in A canopy was associated with greater plant numbers and chloroplast sizes, both of which involved enhanced CO2 diffusion from the atmosphere to chloroplasts in the canopies of P. patens. These results suggest that changes in gravity are important environmental stimuli to induce changes in plant growth and photosynthesis by P. patens, in which an alteration in chloroplast size is one of the key traits. We are now planning an ISS experiment to investigate the responses of P. patens to microgravity.

  1. The Chloroplast Division Protein ARC6 Acts to Inhibit Disassembly of GDP-bound FtsZ2.

    Science.gov (United States)

    Sung, Min Woo; Shaik, Rahamthulla; TerBush, Allan D; Osteryoung, Katherine W; Vitha, Stanislav; Holzenburg, Andreas

    2018-05-16

    Chloroplasts host photosynthesis and fulfill other metabolic functions that are essential to plant life. They have to divide by binary fission to maintain their numbers throughout cycles of cell division. Chloroplast division is achieved by a complex ring-shaped division machinery located on both the inner (stromal) and the outer (cytosolic) side of the chloroplast envelope. The inner division ring (termed the Z ring) is formed by the assembly of tubulin-like FtsZ1 and FtsZ2 proteins. ARC6 is a key chloroplast division protein that interacts with the Z ring. ARC6 spans the inner envelope membrane, is known to stabilize or maintain the Z ring, and anchors the Z ring to the inner membrane through interaction with FtsZ2. The underlying mechanism of Z-ring stabilization is not well understood. Here, biochemical and structural characterization of ARC6 was conducted using light scattering, sedimentation, and light and transmission electron microscopy. The recombinant protein was purified as a dimer. The results indicated that a truncated form of ARC6 (tARC6), representing the stromal portion of ARC6, affects FtsZ2 assembly without forming higher-order structures, and exerts its effect via FtsZ2 dynamics. tARC6 prevented GDP-induced FtsZ2 disassembly and caused a significant net increase in FtsZ2 assembly when GDP was present. Single particle analysis and 3D reconstruction were performed to elucidate the structural basis of ARC6 activity. Together, the data reveal that a dimeric form of tARC6 binds to FtsZ2 filaments and does not increase FtsZ polymerization rates but rather inhibits GDP-associated FtsZ2 disassembly. Published under license by The American Society for Biochemistry and Molecular Biology, Inc.

  2. Oral delivery of Acid Alpha Glucosidase epitopes expressed in plant chloroplasts suppresses antibody formation in treatment of Pompe mice.

    Science.gov (United States)

    Su, Jin; Sherman, Alexandra; Doerfler, Phillip A; Byrne, Barry J; Herzog, Roland W; Daniell, Henry

    2015-10-01

    Deficiency of acid alpha glucosidase (GAA) causes Pompe disease in which the patients systemically accumulate lysosomal glycogen in muscles and nervous systems, often resulting in infant mortality. Although enzyme replacement therapy (ERT) is effective in treating patients with Pompe disease, formation of antibodies against rhGAA complicates treatment. In this report, we investigated induction of tolerance by oral administration of GAA expressed in chloroplasts. Because full-length GAA could not be expressed, N-terminal 410-amino acids of GAA (as determined by T-cell epitope mapping) were fused with the transmucosal carrier CTB. Tobacco transplastomic lines expressing CTB-GAA were generated through site-specific integration of transgenes into the chloroplast genome. Homoplasmic lines were confirmed by Southern blot analysis. Despite low-level expression of CTB-GAA in chloroplasts, yellow or albino phenotype of transplastomic lines was observed due to binding of GAA to a chloroplast protein that has homology to mannose-6 phosphate receptor. Oral administration of the plant-made CTB-GAA fusion protein even at 330-fold lower dose (1.5 μg) significantly suppressed immunoglobulin formation against GAA in Pompe mice injected with 500 μg rhGAA per dose, with several-fold lower titre of GAA-specific IgG1 and IgG2a. Lyophilization increased CTB-GAA concentration by 30-fold (up to 190 μg per g of freeze-dried leaf material), facilitating long-term storage at room temperature and higher dosage in future investigations. This study provides the first evidence that oral delivery of plant cells is effective in reducing antibody responses in ERT for lysosomal storage disorders facilitating further advances in clinical investigations using plant cell culture system or in vitro propagation. © 2015 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

  3. Ruminal protozoal contribution to the duodenal flow of fatty acids following feeding of steers on forages differing in chloroplast content.

    Science.gov (United States)

    Huws, S A; Lee, M R F; Kingston-Smith, A H; Kim, E J; Scott, M B; Tweed, J K S; Scollan, N D

    2012-12-28

    Ruminant products are criticised for their SFA content relative to PUFA, although n-6:n-3 PUFA is desirable for human health ( content of rumen protozoa offers a potentially novel approach to enhance PUFA flow to the duodenum and subsequent incorporation into meat and milk. We evaluated protozoal contribution to duodenal n-3 PUFA flow due to intracellular chloroplast content. A total of six Holstein × Friesian steers were fed, in a two-period changeover design, either straw:concentrate (S:C, 60:40; DM basis; S:C, low chloroplast) or fresh perennial ryegrass (PRG; high chloroplast). Following 12 d adaptation to diet, ruminal protozoal and whole duodenal samples were obtained. N and fatty acid content of whole duodenum and rumen protozoal samples were assessed and protozoal 18S rDNA quantitative PCR performed, enabling calculation of protozoal N flow. The ratio of individual fatty acids:N in rumen protozoal samples was calculated to obtain protozoal fatty acid flows. Based on total fatty acid flow, contribution (%) of protozoa to individual fatty acid flows was calculated. Protozoal fatty acid data and microscopical observations revealed that protozoa were enriched with 18 : 3n-3 following PRG feeding, compared with the S:C diet, due to increased intracellular chloroplast content. However, duodenal protozoal 18S rDNA concentration post PRG feeding was low, indicating rumen retention of the protozoa. Nutrition influences the 18 : 3n-3 content of protozoa; the challenge is to increase protozoal flow to the small intestine, while maintaining sustainable rumen densities.

  4. Complete chloroplast genome sequence of common bermudagrass (Cynodon dactylon (L.) Pers.) and comparative analysis within the family Poaceae.

    Science.gov (United States)

    Huang, Ya-Yi; Cho, Shu-Ting; Haryono, Mindia; Kuo, Chih-Horng

    2017-01-01

    Common bermudagrass (Cynodon dactylon (L.) Pers.) belongs to the subfamily Chloridoideae of the Poaceae family, one of the most important plant families ecologically and economically. This grass has a long connection with human culture but its systematics is relatively understudied. In this study, we sequenced and investigated the chloroplast genome of common bermudagrass, which is 134,297 bp in length with two single copy regions (LSC: 79,732 bp; SSC: 12,521 bp) and a pair of inverted repeat (IR) regions (21,022 bp). The annotation contains a total of 128 predicted genes, including 82 protein-coding, 38 tRNA, and 8 rRNA genes. Additionally, our in silico analyses identified 10 sets of repeats longer than 20 bp and predicted the presence of 36 RNA editing sites. Overall, the chloroplast genome of common bermudagrass resembles those from other Poaceae lineages. Compared to most angiosperms, the accD gene and the introns of both clpP and rpoC1 genes are missing. Additionally, the ycf1, ycf2, ycf15, and ycf68 genes are pseudogenized and two genome rearrangements exist. Our phylogenetic analysis based on 47 chloroplast protein-coding genes supported the placement of common bermudagrass within Chloridoideae. Our phylogenetic character mapping based on the parsimony principle further indicated that the loss of the accD gene and clpP introns, the pseudogenization of four ycf genes, and the two rearrangements occurred only once after the most recent common ancestor of the Poaceae diverged from other monocots, which could explain the unusual long branch leading to the Poaceae when phylogeny is inferred based on chloroplast sequences.

  5. New tools for chloroplast genetic engineering allow the synthesis of human growth hormone in the green alga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Wannathong, Thanyanan; Waterhouse, Janet C; Young, Rosanna E B; Economou, Chloe K; Purton, Saul

    2016-06-01

    In recent years, there has been an increasing interest in the exploitation of microalgae in industrial biotechnology. Potentially, these phototrophic eukaryotes could be used for the low-cost synthesis of valuable recombinant products such as bioactive metabolites and therapeutic proteins. The algal chloroplast in particular represents an attractive target for such genetic engineering, both because it houses major metabolic pathways and because foreign genes can be targeted to specific loci within the chloroplast genome, resulting in high-level, stable expression. However, routine methods for chloroplast genetic engineering are currently available only for one species-Chlamydomonas reinhardtii-and even here, there are limitations to the existing technology, including the need for an expensive biolistic device for DNA delivery, the lack of robust expression vectors, and the undesirable use of antibiotic resistance markers. Here, we describe a new strain and vectors for targeted insertion of transgenes into a neutral chloroplast locus that (i) allow scar-less fusion of a transgenic coding sequence to the promoter/5'UTR element of the highly expressed endogenous genes psaA or atpA, (ii) employ the endogenous gene psbH as an effective but benign selectable marker, and (iii) ensure the successful integration of the transgene construct in all transformant lines. Transformation is achieved by a simple and cheap method of agitation of a DNA/cell suspension with glass beads, with selection based on the phototrophic rescue of a cell wall-deficient ΔpsbH strain. We demonstrate the utility of these tools in the creation of a transgenic line that produces high levels of functional human growth hormone.

  6. Development of Orally Bioavailable Therapeutics by the Chloroplast Expression System to Counter Muscle Degeneration, Weakness, and Fibrosis in DMD

    Science.gov (United States)

    2016-11-01

    strength in the diaphragm muscles . However, by 2 months of treatment the benefits were reduced back to untreated controls. We are continuing to analyze...Enzyme 2 Angiotensin-(1-7) Fibrosis Duchenne Muscular Dystrophy Mdx Skeletal muscle Chloroplast expression Oral bioavailability 3... treatment regimen to determine if any acute functional benefit was achieved. While no significant improvement was observed in EDL muscles , the diaphragms

  7. Photo-protection in the centric diatom Coscinodiscus granii is not controlled by chloroplast high-light avoidance movement.

    Directory of Open Access Journals (Sweden)

    Johannes Wilhelm Goessling

    2016-01-01

    Full Text Available Diatoms are important phototrophs in the worlds’ oceans contributing approximately 40% of the global primary photosynthetic production. This is partially explained by their capacity to exploit environments with variable light conditions, but there is limited knowledge on how diatoms cope with changes in the spectral composition and intensity of light. In this study, the influence of light quality and high irradiance on photosynthesis in the centric diatom Coscinodiscus granii was investigated with microscopic imaging and variable chlorophyll fluorescence techniques. Determination of the wavelength-dependent functional absorption cross-section of photosystem (PS II revealed that absorption of blue light (BL and red light (RL was 2.3-fold and 0.8-fold that of white light (WL, respectively. Hence, BL was more efficiently converted into photo-chemical energy. Excessive energy from BL was dissipated via non-photochemical quenching (NPQ mechanisms, while RL apparently induced only negligible NPQ even at high irradiance. A dose dependent increase of cells exhibiting an altered chloroplast distribution was observed after exposure to high levels of BL and WL, but not RL. However, no effective quantum yield of PSII was measured in the majority of cells with an altered chloroplast distribution, and positive Sytox green® death staining confirmed that most of these cells were dead. We conclude that although Coscinodiscus granii can sustain high irradiance it does not perform chloroplast high-light avoidance movements for photo-protection.

  8. Crystallization of the c[subscript 14]-rotor of the chloroplast ATP synthase reveals that it contains pigments

    Energy Technology Data Exchange (ETDEWEB)

    Varco-Merth, Benjamin; Fromme, Raimund; Wang, Meitian; Fromme, Petra (AZU)

    2008-08-27

    The ATP synthase is one of the most important enzymes on earth as it couples the transmembrane electrochemical potential of protons to the synthesis of ATP from ADP and inorganic phosphage, providing the main ATP source of almost all higher life on earth. During ATP synthesis, stepwise protonation of a conserved carboxylate on each protein subunit of an oligomeric ring of 10--15 c-subunits is commonly thought to drive rotation of the rotor moiety (c{sub 10-14}{gamma}{sup {epsilon}}) relative to stator moiety ({alpha}{sub 3}{beta}{sub 3}{delta}ab{sub 2}). Here we report the isolation and crystallization of the c{sub 14}-ring of subunit c from the spinach chloroplast enzyme diffracting as far as 2.8 {angstrom}. Though ATP synthase was not previously know to contain any pigments, the crystals of the c-subunit possessed a strong yellow color. The pigment analysis revaled that they contain 1 chlorophyll and 2 carotenoids, thereby showing for the first time that the chloroplast ATP synthase contains cofactors, leading to the question of the possible roles of the functions of the pigments in the chloroplast ATP synthase.

  9. The Complete Chloroplast Genome Sequence of Tree of Heaven (Ailanthus altissima (Mill. (Sapindales: Simaroubaceae, an Important Pantropical Tree

    Directory of Open Access Journals (Sweden)

    Josphat K. Saina

    2018-03-01

    Full Text Available Ailanthus altissima (Mill. Swingle (Simaroubaceae is a deciduous tree widely distributed throughout temperate regions in China, hence suitable for genetic diversity and evolutionary studies. Previous studies in A. altissima have mainly focused on its biological activities, genetic diversity and genetic structure. However, until now there is no published report regarding genome of this plant species or Simaroubaceae family. Therefore, in this paper, we first characterized A. altissima complete chloroplast genome sequence. The tree of heaven chloroplast genome was found to be a circular molecule 160,815 base pairs (bp in size and possess a quadripartite structure. The A. altissima chloroplast genome contains 113 unique genes of which 79 and 30 are protein coding and transfer RNA (tRNA genes respectively and also 4 ribosomal RNA genes (rRNA with overall GC content of 37.6%. Microsatellite marker detection identified A/T mononucleotides as majority SSRs in all the seven analyzed genomes. Repeat analyses of seven Sapindales revealed a total of 49 repeats in A. altissima, Rhus chinensis, Dodonaea viscosa, Leitneria floridana, while Azadirachta indica, Boswellia sacra, and Citrus aurantiifolia had a total of 48 repeats. The phylogenetic analysis using protein coding genes revealed that A. altissima is a sister to Leitneria floridana and also suggested that Simaroubaceae is a sister to Rutaceae family. The genome information reported here could be further applied for evolution and invasion, population genetics, and molecular studies in this plant species and family.

  10. Complete chloroplast genome and 45S nrDNA sequences of the medicinal plant species Glycyrrhiza glabra and Glycyrrhiza uralensis.

    Science.gov (United States)

    Kang, Sang-Ho; Lee, Jeong-Hoon; Lee, Hyun Oh; Ahn, Byoung Ohg; Won, So Youn; Sohn, Seong-Han; Kim, Jung Sun

    2017-10-06

    Glycyrrhiza uralensis and G. glabra, members of the Fabaceae, are medicinally important species that are native to Asia and Europe. Extracts from these plants are widely used as natural sweeteners because of their much greater sweetness than sucrose. In this study, the three complete chloroplast genomes and five 45S nuclear ribosomal (nr)DNA sequences of these two licorice species and an interspecific hybrid are presented. The chloroplast genomes of G. glabra, G. uralensis and G. glabra × G. uralensis were 127,895 bp, 127,716 bp and 127,939 bp, respectively. The three chloroplast genomes harbored 110 annotated genes, including 76 protein-coding genes, 30 tRNA genes and 4 rRNA genes. The 45S nrDNA sequences were either 5,947 or 5,948 bp in length. Glycyrrhiza glabra and G. glabra × G. uralensis showed two types of nrDNA, while G. uralensis contained a single type. The complete 45S nrDNA sequence unit contains 18S rRNA, ITS1, 5.8S rRNA, ITS2 and 26S rRNA. We identified simple sequence repeat and tandem repeat sequences. We also developed four reliable markers for analysis of Glycyrrhiza diversity authentication.

  11. Effects of some inhibitors of protein synthesis on the chloroplast fine structure, CO2 fixation and the Hill reaction activity

    Directory of Open Access Journals (Sweden)

    S. Więckowski

    2015-01-01

    Full Text Available A comparative study concerning the effects of chloramphenicol (100 μg ml-1, actidione (10 μg ml-1, 5-bromouracil (190 μg ml-1, actinomycin D (30 μg ml-1 and DL-ethionine (800 μg ml-1 on the chloroplast fine structure, 14CO2 incorporation and the Hill reaction activity was the subject of the experiments presented in this paper. The experiments were conducted on bean seedlings under the conditions when chlorophyll accumulation was inhibited only partially. The results obtained indicate that chloromphenicol is responsible for the reduction of the number of grana per section of plastid and for the formation of numerous vesicles in the stroma. In the presence of actidione, actinomycin D or DL-ethionine the lamellae are poorly differentiated into .stroma and granum regions and there occur disturbances in the typical orientation of lamellae within chloroplasts. Only in the presence of 5-bromouracil the development of chloroplast structure resemble that in control plants. A comparison of the results obtained with those published earlier (Więckowski et al., 1974; Ficek and Więckowski, 1974 shows that such processes as assimilatory pigment accumulation, the rate of CO2 fixation, the Hill reaction activity, and the development of lamellar system are suppressed in a different extent by the inhibitors used.

  12. Regulation of Ribulose-1,5-Bisphosphate Carboxylase Activity by the Activase System in Lysed Spinach Chloroplasts

    Science.gov (United States)

    Parry, Martin A. J.; Keys, Alfred J.; Foyer, Christine H.; Furbank, Robert T.; Walker, David A.

    1988-01-01

    Ribulose-1,5-bisphosphate (RuBP) carboxylase in lysed spinach (Spinacia oleracea L. cv virtuosa) chloroplasts that had been partly inactivated at low CO2 and Mg2+ by incubating in darkness with 4 millimolar partially purified RuBP was reactivated by light. If purified RuBP was used to inhibit dark activation of the enzyme, reactivation by light was not observed unless fructose-1,6-bisphosphate, ATP, or ADP plus inorganic phosphate were also added. Presumably, ADP plus inorganic phosphate acted as an ATP-generating system with a requirement for the generation of ΔpH across the thylakoid membrane. When the RuBP obtained from Sigma Chemical Co. was used, light did not reactivate the enzyme. There was no direct correlation between ΔpH and activation. Therefore, thylakoids are required in the ribulose-1,5-bisphosphate carboxylase activase system largely to synthesize ATP. Inactivation of RuBP carboxylase in isolated chloroplasts or in the lysed chloroplast system was not promoted simply by a transition from light to dark conditions but was caused by low CO2 and Mg2+. PMID:16666184

  13. Preprotein import into chloroplasts via the Toc and Tic complexes is regulated by redox signals in Pisum sativum.

    Science.gov (United States)

    Stengel, Anna; Benz, J Philipp; Buchanan, Bob B; Soll, Jürgen; Bölter, Bettina

    2009-11-01

    The import of nuclear-encoded preproteins is necessary to maintain chloroplast function. The recognition and transfer of most precursor proteins across the chloroplast envelopes are facilitated by two membrane-inserted protein complexes, the translocons of the chloroplast outer and inner envelope (Toc and Tic complexes, respectively). Several signals have been invoked to regulate the import of preproteins. In our study, we were interested in redox-based import regulation mediated by two signals: regulation based on thiols and on the metabolic NADP+/NADPH ratio. We sought to identify the proteins participating in the regulation of these transport pathways and to characterize the preprotein subgroups whose import is redox-dependent. Our results provide evidence that the formation and reduction of disulfide bridges in the Toc receptors and Toc translocation channel have a strong influence on import yield of all tested preproteins that depend on the Toc complex for translocation. Furthermore, the metabolic NADP+/NADPH ratio influences not only the composition of the Tic complex, but also the import efficiency of most, but not all, preproteins tested. Thus, several Tic subcomplexes appear to participate in the translocation of different preprotein subgroups, and the redox-active components of these complexes likely play a role in regulating transport.

  14. Multi-functional roles for the polypeptide transport associated domains of Toc75 in chloroplast protein import

    Science.gov (United States)

    Paila, Yamuna D; Richardson, Lynn GL; Inoue, Hitoshi; Parks, Elizabeth S; McMahon, James; Inoue, Kentaro; Schnell, Danny J

    2016-01-01

    Toc75 plays a central role in chloroplast biogenesis in plants as the membrane channel of the protein import translocon at the outer envelope of chloroplasts (TOC). Toc75 is a member of the Omp85 family of bacterial and organellar membrane insertases, characterized by N-terminal POTRA (polypeptide-transport associated) domains and C-terminal membrane-integrated β-barrels. We demonstrate that the Toc75 POTRA domains are essential for protein import and contribute to interactions with TOC receptors, thereby coupling preprotein recognition at the chloroplast surface with membrane translocation. The POTRA domains also interact with preproteins and mediate the recruitment of molecular chaperones in the intermembrane space to facilitate membrane transport. Our studies are consistent with the multi-functional roles of POTRA domains observed in other Omp85 family members and demonstrate that the domains of Toc75 have evolved unique properties specific to the acquisition of protein import during endosymbiotic evolution of the TOC system in plastids. DOI: http://dx.doi.org/10.7554/eLife.12631.001 PMID:26999824

  15. Membrane Composition and Physiological Activity of Plastids from an Oenothera Plastome Mutator-Induced Chloroplast Mutant 1

    Science.gov (United States)

    Johnson, Ellen M.; Sears, Barbara B.

    1990-01-01

    Plastids were isolated from a plastome mutator-induced mutant (pm7) of Oenothera hookeri and were analyzed for various physiological and biochemical attributes. No photosynthetic electron transport activity was detected in the mutant plastids. This is consistent with previous ultrastructural analysis showing the absence of thylakoid membranes in the pm7 plastids and with the observation of aberrant processing and accumulation of chloroplast proteins in the mutant. In comparison to wild type, the mutant tissue lacks chlorophyll, and has significant differences in levels of four fatty acids. The analyses did not reveal any differences in carotenoid levels nor in the synthesis of several chloroplast lipids. The consequences of the altered composition of the chloroplast membrane are discussed in terms of their relation to the aberrant protein processing of the pm7 plastids. The pigment, fatty acid, and lipid measurements were also performed on two distinct nuclear genotypes (A/A and A/C) which differ in their compatibility with the plastid genome (type I) contained in these lines. In these cases, only chlorophyll concentrations differed significantly. PMID:16667256

  16. Constitutive Transcription and Stable RNA Accumulation in Plastids during the Conversion of Chloroplasts to Chromoplasts in Ripening Tomato Fruits 1

    Science.gov (United States)

    Marano, María Rosa; Carrillo, Néstor

    1992-01-01

    The size distribution of plastid transcripts during chromoplast differentiation in ripening tomato (Lycopersicon esculentum L.) fruits was determined using northern blot analysis. Hybridization of total cellular RNA from leaves and fruits with several tobacco chloroplast DNA probes showed distinct transcript patterns in chloroplasts and chromoplasts. We also compared transcriptional rates by probing immobilized DNA fragments of small size (representing about 85% of the plastid genome) with run-on transcripts from tomato plastids. The relative rates of transcription of the various DNA regions were very similar in chloro- and chromoplasts. Parallel determination of the steady-state levels of plastid RNA showed no strict correlation between synthesis rate and RNA accumulation. Differences in the relative abundance of transcripts between chloro- and chromoplasts were not very pronounced and were limited to a small number of genes. The results indicate that the conversion of chloroplasts to chromoplasts at the onset of tomato fruit ripening proceeds with no important variations in the relative transcription rates and with only moderate changes in the relative stability of plastid-encoded transcripts. Images Figure 1 Figure 4 PMID:16653091

  17. Re-exploration of U's Triangle Brassica Species Based on Chloroplast Genomes and 45S nrDNA Sequences.

    Science.gov (United States)

    Kim, Chang-Kug; Seol, Young-Joo; Perumal, Sampath; Lee, Jonghoon; Waminal, Nomar Espinosa; Jayakodi, Murukarthick; Lee, Sang-Choon; Jin, Seungwoo; Choi, Beom-Soon; Yu, Yeisoo; Ko, Ho-Cheol; Choi, Ji-Weon; Ryu, Kyoung-Yul; Sohn, Seong-Han; Parkin, Isobel; Yang, Tae-Jin

    2018-05-09

    The concept of U's triangle, which revealed the importance of polyploidization in plant genome evolution, described natural allopolyploidization events in Brassica using three diploids [B. rapa (A genome), B. nigra (B), and B. oleracea (C)] and derived allotetraploids [B. juncea (AB genome), B. napus (AC), and B. carinata (BC)]. However, comprehensive understanding of Brassica genome evolution has not been fully achieved. Here, we performed low-coverage (2-6×) whole-genome sequencing of 28 accessions of Brassica as well as of Raphanus sativus [R genome] to explore the evolution of six Brassica species based on chloroplast genome and ribosomal DNA variations. Our phylogenomic analyses led to two main conclusions. (1) Intra-species-level chloroplast genome variations are low in the three allotetraploids (2~7 SNPs), but rich and variable in each diploid species (7~193 SNPs). (2) Three allotetraploids maintain two 45SnrDNA types derived from both ancestral species with maternal dominance. Furthermore, this study sheds light on the maternal origin of the AC chloroplast genome. Overall, this study clarifies the genetic relationships of U's triangle species based on a comprehensive genomics approach and provides important genomic resources for correlative and evolutionary studies.

  18. Transcriptomic and Functional Analyses Reveal That PpGLK1 Regulates Chloroplast Development in Peach (Prunus persica

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    Min Chen

    2018-01-01

    Full Text Available Peach is an ideal species for fruit tree research because of its small, fully sequenced genome. Chloroplast development is dependent on the tight cooperation between the nuclear and plastid genomes, and is regulated by GLK transcription factors. In this work, the pigment content was monitored and the chloroplast-to-chromoplast conversion during the fruit ripening was visualized by transmission electron microscopy. Localization and expression analyses showed that PpGLK1 was located in the nucleus and expressed mainly in the leaves and fruit skin. A transcriptome analysis showed that PpGLK1 and its target genes were significantly differentially expressed in ripening peach fruit skin. PpGLK1 silencing affected chlorophyll accumulation in peach leaves and fruits. Overexpression of PpGLK1 rescued the phenotypes of the Arabidopsis Atglk1Atglk2 double mutant and the tomato uniform ripening mutant. The results of a yeast two-hybrid analysis showed that PpGLK1 is autoactivated and that PpGLK1 (301-542 a.a. interacted with PpARF5. Together, our results indicate that PpGLK1 regulates chloroplast development in green tissues in peach. Therefore, it may be a promising target gene for improving the production and quality of peach by genetic engineering and breeding approaches.

  19. Optimization of codon composition and regulatory elements for expression of human insulin like growth factor-1 in transgenic chloroplasts and evaluation of structural identity and function.

    Science.gov (United States)

    Daniell, Henry; Ruiz, Gricel; Denes, Bela; Sandberg, Laurence; Langridge, William

    2009-04-03

    Transgenic chloroplasts are potential bioreactors for recombinant protein production, especially for achievement of high levels of protein expression and proper folding. Production of therapeutic proteins in leaves provides transgene containment by elimination of reproductive structures. Therefore, in this study, human Insulin like Growth Factor-1 is expressed in transgenic chloroplasts for evaluation of structural identity and function. Expression of the synthetic Insulin like Growth Factor 1 gene (IGF-1s, 60% AT) was observed in transformed E. coli. However, no native IGF-1 gene (IGF-1n, 41% AT) product was detected in the western blots in E. coli. Site-specific integration of the transgenes into the tobacco chloroplast genome was confirmed after transformation using PCR. Southern blot analysis confirmed that the transgenic lines were homoplasmic. The transgenic plant lines had IGF-1s expression levels of 11.3% of total soluble protein (TSP). The IGF-1n plants contained 9.5% TSP as IGF-1n, suggesting that the chloroplast translation machinery is more flexible than E. coli in codon preference and usage. The expression of IGF-1 was increased up to 32% TSP under continuous illumination by the chloroplast light regulatory elements. IgG-Sepharose affinity column chromatographic separation of Z domain containing chloroplast derived IGF-1 protein, single and two dimensional electrophoresis methods and mass spectrometer analysis confirmed the identity of human IGF-1 in transgenic chloroplasts. Two spots analyzed from 2-D focusing/phoresis acrylamide gel showed the correct amino acid sequence of human IGF-1 and the S. aureus Z-tag. Cell proliferation assays in human HU-3 cells demonstrated the biological activity of chloroplast derived IGF-1 even in the presence of the S. aureus Z tag. This study demonstrates that the human Insulin like Growth Factor-1 expressed in transgenic chloroplasts is identical to the native protein and is fully functional. The ability to use plant

  20. Optimization of codon composition and regulatory elements for expression of human insulin like growth factor-1 in transgenic chloroplasts and evaluation of structural identity and function

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    Sandberg Laurence

    2009-04-01

    Full Text Available Abstract Background Transgenic chloroplasts are potential bioreactors for recombinant protein production, especially for achievement of high levels of protein expression and proper folding. Production of therapeutic proteins in leaves provides transgene containment by elimination of reproductive structures. Therefore, in this study, human Insulin like Growth Factor-1 is expressed in transgenic chloroplasts for evaluation of structural identity and function. Results Expression of the synthetic Insulin like Growth Factor 1 gene (IGF-1s, 60% AT was observed in transformed E. coli. However, no native IGF-1 gene (IGF-1n, 41% AT product was detected in the western blots in E. coli. Site-specific integration of the transgenes into the tobacco chloroplast genome was confirmed after transformation using PCR. Southern blot analysis confirmed that the transgenic lines were homoplasmic. The transgenic plant lines had IGF-1s expression levels of 11.3% of total soluble protein (TSP. The IGF-1n plants contained 9.5% TSP as IGF-1n, suggesting that the chloroplast translation machinery is more flexible than E. coli in codon preference and usage. The expression of IGF-1 was increased up to 32% TSP under continuous illumination by the chloroplast light regulatory elements. IgG-Sepharose affinity column chromatographic separation of Z domain containing chloroplast derived IGF-1 protein, single and two dimensional electrophoresis methods and mass spectrometer analysis confirmed the identity of human IGF-1 in transgenic chloroplasts. Two spots analyzed from 2-D focusing/phoresis acrylamide gel showed the correct amino acid sequence of human IGF-1 and the S. aureus Z-tag. Cell proliferation assays in human HU-3 cells demonstrated the biological activity of chloroplast derived IGF-1 even in the presence of the S. aureus Z tag. Conclusion This study demonstrates that the human Insulin like Growth Factor-1 expressed in transgenic chloroplasts is identical to the native

  1. Relationship of tightly bound ADP and ATP to control and catalysis by chloroplast ATP synthase

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, J.; Xue, Z.; Du, Z.; Melese, T.; Boyer, P.D.

    1988-07-12

    Whether the tightly bound ADP that can cause a pronounced inhibition of ATP hydrolysis by the chloroplast ATP synthase and F/sub 1/ ATPase (CF/sub 1/) is bound at catalytic sites or at noncatalytic regulatory sites or both has been uncertain. The authors have used photolabeling by 2-azido-ATP and 2-azido-ADP to ascertain the location, with Mg/sup 2 +/ activation, of tightly bound ADP (a) that inhibits the hydrolysis of ATP by chloroplast ATP synthase, (b) that can result in an inhibited form of CF/sub 1/ that slowly regains activity during ATP hydrolysis, and (c) that arises when low concentrations of ADP markedly inhibit the hydrolysis of GTP by CF/sub 1/. The data show that in all instances the inhibition is associated with ADP binding without inorganic phosphate (P/sub i/) at catalytic sites. After photophosphorylation of ADP or 2-azido-ADP with (/sup 32/P)P/sub i/, similar amounts of the corresponding triphosphates are present on washed thylakoid membranes. Trials with appropriately labeled substrates show that a small portion of the tightly bound 2-azido-ATP gives rise to covalent labeling with an ATP moiety at noncatalytic sites but that most of the bound 2-azido-ATP gives rise to covalent labeling with an ATP moiety at noncatalytic sites but that most of the bound 2-azido-ATP gives rise to covalent labeling by an ADP moiety at a catalytic site. They also report the occurrence of a 1-2-min delay in the onset of the Mg/sup 2 +/-induced inhibition after addition of CF/sub 1/ to solutions containing Mg/sup 2 +/ and ATP, and that this delay is not associated with the filling of noncatalytic sites. A rapid burst of P/sub i/ formation is followed by a much lower, constant steady-state rate. The burst is not observed with GTP as a substrate or with Ca/sup 2 +/ as the activating cation.

  2. Complete chloroplast genome sequence of Elodea canadensis and comparative analyses with other monocot plastid genomes.

    Science.gov (United States)

    Huotari, Tea; Korpelainen, Helena

    2012-10-15

    Elodea canadensis is an aquatic angiosperm native to North America. It has attracted great attention due to its invasive nature when transported to new areas in its non-native range. We have determined the complete nucleotide sequence of the chloroplast (cp) genome of Elodea. Taxonomically Elodea is a basal monocot, and only few monocot cp genomes representing early lineages of monocots have been sequenced so far. The genome is a circular double-stranded DNA molecule 156,700 bp in length, and has a typical structure with large (LSC 86,194 bp) and small (SSC 17,810 bp) single-copy regions separated by a pair of inverted repeats (IRs 26,348 bp each). The Elodea cp genome contains 113 unique genes and 16 duplicated genes in the IR regions. A comparative analysis showed that the gene order and organization of the Elodea cp genome is almost identical to that of Amborella trichopoda, a basal angiosperm. The structure of IRs in Elodea is unique among monocot species with the whole cp genome sequenced. In Elodea and another monocot Lemna minor the borders between IRs and LSC are located upstream of rps 19 gene and downstream of trnH-GUG gene, while in most monocots, IR has extended to include both trnH and rps 19 genes. A phylogenetic analysis conducted using Bayesian method, based on the DNA sequences of 81 chloroplast genes from 17 monocot taxa provided support for the placement of Elodea together with Lemna as a basal monocot and the next diverging lineage of monocots after Acorales. In comparison with other monocots, the Elodea cp genome has gone through only few rearrangements or gene losses. IR of Elodea has a unique structure among the monocot species studied so far as its structure is similar to that of a basal angiosperm Amborella. This result together with phylogenetic analyses supports the placement of Elodea as a basal monocot to the next diverging lineage of monocots after Acorales. So far, only few cp genomes representing early lineages of monocots have been

  3. Complete chloroplast genomes from apomictic Taraxacum (Asteraceae): Identity and variation between three microspecies

    Science.gov (United States)

    Majeský, Ľuboš; Schwarzacher, Trude; Gornall, Richard; Heslop-Harrison, Pat

    2017-01-01

    Chloroplast DNA sequences show substantial variation between higher plant species, and less variation within species, so are typically excellent markers to investigate evolutionary, population and genetic relationships and phylogenies. We sequenced the plastomes of Taraxacum obtusifrons Markl. (O978); T. stridulum Trávniček ined. (S3); and T. amplum Markl. (A978), three apomictic triploid (2n = 3x = 24) dandelions from the T. officinale agg. We aimed to characterize the variation in plastomes, define relationships and correlations with the apomictic microspecies status, and refine placement of the microspecies in the evolutionary or phylogenetic context of the Asteraceae. The chloroplast genomes of accessions O978 and S3 were identical and 151,322 bp long (where the nuclear genes are known to show variation), while A978 was 151,349 bp long. All three genomes contained 135 unique genes, with an additional copy of the trnF-GGA gene in the LSC region and 20 duplicated genes in the IR region, along with short repeats, the typical major Inverted Repeats (IR1 and IR2, 24,431bp long), and Large and Small Single Copy regions (LSC 83,889bp and SSC 18,571bp in O978). Between the two Taraxacum plastomes types, we identified 28 SNPs. The distribution of polymorphisms suggests some parts of the Taraxacum plastome are evolving at a slower rate. There was a hemi-nested inversion in the LSC region that is common to Asteraceae, and an SSC inversion from ndhF to rps15 found only in some Asteraceae lineages. A comparative repeat analysis showed variation between Taraxacum and the phylogenetically close genus Lactuca, with many more direct repeats of 40bp or more in Lactuca (1% larger plastome than Taraxacum). When individual genes and non-coding regions were for Asteraceae phylogeny reconstruction, not all showed the same evolutionary scenario suggesting care is needed for interpretation of relationships if a limited number of markers are used. Studying genotypic diversity in

  4. Functional chloroplasts in metazoan cells - a unique evolutionary strategy in animal life

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    Krug Patrick J

    2009-12-01

    Full Text Available Abstract Background Among metazoans, retention of functional diet-derived chloroplasts (kleptoplasty is known only from the sea slug taxon Sacoglossa (Gastropoda: Opisthobranchia. Intracellular maintenance of plastids in the slug's digestive epithelium has long attracted interest given its implications for understanding the evolution of endosymbiosis. However, photosynthetic ability varies widely among sacoglossans; some species have no plastid retention while others survive for months solely on photosynthesis. We present a molecular phylogenetic hypothesis for the Sacoglossa and a survey of kleptoplasty from representatives of all major clades. We sought to quantify variation in photosynthetic ability among lineages, identify phylogenetic origins of plastid retention, and assess whether kleptoplasty was a key character in the radiation of the Sacoglossa. Results Three levels of photosynthetic activity were detected: (1 no functional retention; (2 short-term retention lasting about one week; and (3 long-term retention for over a month. Phylogenetic analysis of one nuclear and two mitochondrial loci revealed reciprocal monophyly of the shelled Oxynoacea and shell-less Plakobranchacea, the latter comprising a monophyletic Plakobranchoidea and paraphyletic Limapontioidea. Only species in the Plakobranchoidea expressed short- or long-term kleptoplasty, most belonging to a speciose clade of slugs bearing parapodia (lateral flaps covering the dorsum. Bayesian ancestral character state reconstructions indicated that functional short-term retention arose once in the last common ancestor of Plakobranchoidea, and independently evolved into long-term retention in four derived species. Conclusion We propose a sequential progression from short- to long-term kleptoplasty, with different adaptations involved in each step. Short-term kleptoplasty likely arose as a deficiency in plastid digestion, yielding additional energy via the release of fixed carbon

  5. Exogenous spermidine is enhancing tomato tolerance to salinity-alkalinity stress by regulating chloroplast antioxidant system and chlorophyll metabolism.

    Science.gov (United States)

    Li, Jianming; Hu, Lipan; Zhang, Li; Pan, Xiongbo; Hu, Xiaohui

    2015-12-29

    Salinity-alkalinity stress is known to adversely affect a variety of processes in plants, thus inhibiting growth and decreasing crop yield. Polyamines protect plants against a variety of environmental stresses. However, whether exogenous spermidine increases the tolerance of tomato seedlings via effects on chloroplast antioxidant enzymes and chlorophyll metabolism is unknown. In this study, we examined the effect of exogenous spermidine on chlorophyll synthesis and degradation pathway intermediates and related enzyme activities, as well as chloroplast ultrastructure, gene expression, and antioxidants in salinity-alkalinity-stressed tomato seedlings. Salinity-alkalinity stress disrupted chlorophyll metabolism and hindered uroorphyrinogen III conversion to protoporphyrin IX. These effects were more pronounced in seedlings of cultivar Zhongza No. 9 than cultivar Jinpengchaoguan. Under salinity-alkalinity stress, exogenous spermidine alleviated decreases in the contents of total chlorophyll and chlorophyll a and b in seedlings of both cultivars following 4 days of stress. With extended stress, exogenous spermidine reduced the accumulation of δ-aminolevulinic acid, porphobilinogen, and uroorphyrinogen III and increased the levels of protoporphyrin IX, Mg-protoporphyrin IX, and protochlorophyllide, suggesting that spermidine promotes the conversion of uroorphyrinogen III to protoporphyrin IX. The effect occurred earlier in cultivar Jinpengchaoguan than in cultivar Zhongza No. 9. Exogenous spermidine also alleviated the stress-induced increases in malondialdehyde content, superoxide radical generation rate, chlorophyllase activity, and expression of the chlorophyllase gene and the stress-induced decreases in the activities of antioxidant enzymes, antioxidants, and expression of the porphobilinogen deaminase gene. In addition, exogenous spermidine stabilized the chloroplast ultrastructure in stressed tomato seedlings. The tomato cultivars examined exhibited different

  6. Overexpression of chloroplast NADPH-dependent thioredoxin reductase in Arabidopsis enhances leaf growth and elucidates in-vivo function of reductase and thioredoxin domains

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    Jouni eToivola

    2013-10-01

    Full Text Available Plant chloroplasts have versatile thioredoxin systems including two thioredoxin reductases and multiple types of thioredoxins. Plastid-localized NADPH-dependent thioredoxin reductase (NTRC contains both reductase (NTRd and thioredoxin (TRXd domains in a single polypeptide and forms homodimers. To study the action of NTRC and NTRC domains in vivo, we have complemented the ntrc knockout line of Arabidopsis with the wild type and full-length NTRC genes, in which 2-Cys motifs either in NTRd, or in TRXd were inactivated. The ntrc line was also transformed either with the truncated NTRd or TRXd alone. Overexpression of wild-type NTRC promoted plant growth by increasing leaf size and biomass yield of the rosettes. Complementation of the ntrc line with the full-length NTRC gene containing an active reductase but an inactive thioredoxin domain, or vice versa, recovered wild-type chloroplast phenotype and, partly, rosette biomass production, indicating that the NTRC domains are capable of interacting with other chloroplast thioredoxin systems. Overexpression of truncated NTRd or TRXd in ntrc background did not restore wild-type phenotype. Modelling of the 3-dimensional structure of the NTRC dimer indicates extensive interactions between the NTR domains and the TRX domains further stabilize the dimeric structure. The long linker region between the NTRd and TRXd, however, allows flexibility for the position of the TRXd in the dimer. Supplementation of the TRXd in the NTRC homodimer model by free chloroplast thioredoxins indicated that TRXf is the most likely partner to interact with NTRC. We propose that overexpression of NTRC promotes plant biomass yield both directly by stimulation of chloroplast biosynthetic and protected pathways controlled by NTRC and indirectly via free chloroplast thioredoxins. Our data indicate that overexpression of chloroplast thiol redox-regulator has a potential to increase biofuel yield in plant and algal species suitable for

  7. The role of autophagy in chloroplast degradation and chlorophagy in immune defenses during Pst DC3000 (AvrRps4 infection.

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    Junjian Dong

    Full Text Available BACKGROUND: Chlorosis of leaf tissue normally observed during pathogen infection may result from the degradation of chloroplasts. There is a growing evidence to suggest that the chloroplast plays a significant role during pathogen infection. Although most degradation of the organelles and cellular structures in plants is mediated by autophagy, its role in chloroplast catabolism during pathogen infection is largely unknown. RESULTS: In this study, we investigated the function of autophagy in chloroplast degradation during avirulent Pst DC3000 (AvrRps4 infection. We examined the expression of defensive marker genes and suppression of bacterial growth using the electrolyte leakage assay in normal light (N and low light (L growing environments of wild-type and atg5-1 plants during pathogen treatment. Stroma-targeted GFP proteins (CT-GFP were observed with LysoTracker Red (LTR staining of autophagosome-like structures in the vacuole. The results showed that Arabidopsis expressed a significant number of small GFP-labeled bodies when infected with avirulent Pst DC3000 (AvrRps4. While barely detectable, there were small GFP-labeled bodies in plants with the CT-GFP expressing atg5-1 mutation. The results showed that chloroplast degradation depends on autophagy and this may play an important role in inhibiting pathogen growth. CONCLUSION: Autophagy plays a role in chloroplast degradation in Arabidopsis during avirulent Pst DC3000 (AvrRps4 infection. Autophagy dependent chloroplast degradation may be the primary source of reactive oxygen species (ROS as well as the pathogen-response signaling molecules that induce the defense response.

  8. Transcriptional Slippage and RNA Editing Increase the Diversity of Transcripts in Chloroplasts: Insight from Deep Sequencing of Vigna radiata Genome and Transcriptome.

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    Ching-Ping Lin

    Full Text Available We performed deep sequencing of the nuclear and organellar genomes of three mungbean genotypes: Vigna radiata ssp. sublobata TC1966, V. radiata var. radiata NM92 and the recombinant inbred line RIL59 derived from a cross between TC1966 and NM92. Moreover, we performed deep sequencing of the RIL59 transcriptome to investigate transcript variability. The mungbean chloroplast genome has a quadripartite structure including a pair of inverted repeats separated by two single copy regions. A total of 213 simple sequence repeats were identified in the chloroplast genomes of NM92 and RIL59; 78 single nucleotide variants and nine indels were discovered in comparing the chloroplast genomes of TC1966 and NM92. Analysis of the mungbean chloroplast transcriptome revealed mRNAs that were affected by transcriptional slippage and RNA editing. Transcriptional slippage frequency was positively correlated with the length of simple sequence repeats of the mungbean chloroplast genome (R2=0.9911. In total, 41 C-to-U editing sites were found in 23 chloroplast genes and in one intergenic spacer. No editing site that swapped U to C was found. A combination of bioinformatics and experimental methods revealed that the plastid-encoded RNA polymerase-transcribed genes psbF and ndhA are affected by transcriptional slippage in mungbean and in main lineages of land plants, including three dicots (Glycine max, Brassica rapa, and Nicotiana tabacum, two monocots (Oryza sativa and Zea mays, two gymnosperms (Pinus taeda and Ginkgo biloba and one moss (Physcomitrella patens. Transcript analysis of the rps2 gene showed that transcriptional slippage could affect transcripts at single sequence repeat regions with poly-A runs. It showed that transcriptional slippage together with incomplete RNA editing may cause sequence diversity of transcripts in chloroplasts of land plants.

  9. Two new fern chloroplasts and decelerated evolution linked to the long generation time in tree ferns.

    Science.gov (United States)

    Zhong, Bojian; Fong, Richard; Collins, Lesley J; McLenachan, Patricia A; Penny, David

    2014-04-30

    We report the chloroplast genomes of a tree fern (Dicksonia squarrosa) and a "fern ally" (Tmesipteris elongata), and show that the phylogeny of early land plants is basically as expected, and the estimates of divergence time are largely unaffected after removing the fastest evolving sites. The tree fern shows the major reduction in the rate of evolution, and there has been a major slowdown in the rate of mutation in both families of tree ferns. We suggest that this is related to a generation time effect; if there is a long time period between generations, then this is probably incompatible with a high mutation rate because otherwise nearly every propagule would probably have several lethal mutations. This effect will be especially strong in organisms that have large numbers of cell divisions between generations. This shows the necessity of going beyond phylogeny and integrating its study with other properties of organisms. © The Author(s) 2014. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  10. Reversible pH-dependent activation/inactivation of CF(1-ATPase of spinach chloroplasts

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    A. P. Khomochkin

    2017-08-01

    Full Text Available The aim of the work was to study the reverse pH-dependent regulation of the enzymatic activity of the catalytic part of ATP synthase (EC 3.6.3.14 of chloroplast – coupling factor CF1. It was shown that the short-term incubation of isolated CF1 in the media with pH 4.5 or 3.5 leads to inactivation of Ca2+-ATPase, which is rapidly (t1/2 ~ 1 min restored in the medium containing 0.5-10 mM bicarbonate at pH 7.8. After acid treatment, the rate of Mg2+-ATPase reaction was also stimulated in the presence of 1 mM bicarbonate (рН 7.8; 37 °С. The increase in Ca2+– and Mg2+-АТР activity of CF1 associated with the addition of NaHCO3 solution was completely eliminated after the introduction of 50 mM acetazolamide – a specific inhibitor of carbonic anhydrase. The obtained results suggest the existence of the bound bicarbonate in the CF1 structure, which apparently participates in proton transfer.

  11. Carbonic anhydrase activity of integral-functional complexes of thylakoid membranes of spinach chloroplasts

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    A. V. Semenihin

    2015-06-01

    Full Text Available Isolated thylakoid membranes were disrupted by treatment with nonionic detergents digitonin or dodecyl maltoside. Solubilized polypeptide complexes were separated by native gel charge shift electrophoresis. The position of ATP-synthase complex and its isolated catalytic part (CF1 within gel was determined using the color reaction for ATPase activity. Due to the presence of cytochromes, the red band in unstained gels corresponded to the cytochrome b6f complex. Localization of the cytochrome b6f complex, ATP synthase and coupling CF1 in the native gel was confirmed by their subunit composition determined after SDS-electrophoretic analysis. Carbonic anhydrase (CA activity in polypeptide zones of PS II, cytochrome b6f complex, and ATP-synthase CF1 was identified in native gels using indicator bromothymol blue. CA activity of isolated CF1 in solution was determined by infrared gas analysis as the rate of bicarbonate dehydration. The water-soluble acetazolamide, an inhibitor of CA, unlike lipophilic ethoxyzolamide inhibited CA activity of CF1. Thus, it was shown for the first time that ATP-synthase has a component which is capable of catalyzing the interconversion of forms of carbonic acid associated with proton exchange. The data obtained suggest the presence of multiple forms of carbonic anhydrase in the thylakoid membranes of spinach chloroplasts and confirm their involvement in the proton transfer to the ATP synthase.

  12. Rotation of nucleotide sites is not required for the enzymatic activity of chloroplast coupling factor

    Energy Technology Data Exchange (ETDEWEB)

    Musier, K.M.; Hammes, G.G.

    1987-09-22

    New heterobifunctional photoaffinity cross-linking reagents, 6-maleimido-N-(4-benzoylphenyl)hexanamide, 12-maleimido-N-(4-benzoylphenyl)dodecanamide, and 12-(/sup 14/C)maleimido-N-(4-benzoylphenyo)dodecanamide, were synthesized to investigate the mechanism of ATP hydrolysis by chloroplast coupling factor 1. These reagents react with sulfhydryl groups on the ..gamma..-polypeptide. Subsequent photolysis cross-links the ..gamma..-polypeptide covalently to ..cap alpha..- and ..beta..-polypeptides. The cross-linkers prevent major movements of the ..gamma..-polypeptide with respect to the ..cap alpha..- and ..beta..-polypeptides but are sufficiently long to permit some flexibility in the enzyme structure. When approx. 50% of the ..gamma..-polypeptide was cross-linked to a ..cap alpha..- and ..beta..-polypeptides, a 7% loss in ATPase activity was observed for the longer cross-linker and a 12% loss for the shorter. These results indicate that large movements of ..cap alpha..- and ..beta..-polypeptides with respect to the ..gamma..-polypeptide are not essential for catalysis. In particular, rotation of the polypeptide chains to crease structurally equivalent sites during catalysis is not a required feature of the enzyme mechanism.

  13. Comparative Analysis of the Complete Chloroplast Genomes of Four Aconitum Medicinal Species

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    Jing Meng

    2018-04-01

    Full Text Available Aconitum (Ranunculaceae consists of approximately 400 species distributed in the temperate regions of the northern hemisphere. Many species are well-known herbs, mainly used for analgesia and anti-inflammatory purposes. This genus is well represented in China and has gained widespread attention for its toxicity and detoxification properties. In southwestern China, several Aconitum species, called ‘Dula’ in the Yi Nationality, were often used to control the poisonous effects of other Aconitum plants. In this study, the complete chloroplast (cp genomes of these species were determined for the first time through Illumina paired-end sequencing. Our results indicate that their cp genomes ranged from 151,214 bp (A. episcopale to 155,769 bp (A. delavayi in length. A total of 111–112 unique genes were identified, including 85 protein-coding genes, 36–37 tRNA genes and eight ribosomal RNA genes (rRNA. We also analyzed codon usage, IR expansion or contraction and simple sequence repeats in the cp genomes. Eight variable regions were identified and these may potentially be useful as specific DNA barcodes for species identification of Aconitum. Phylogenetic analysis revealed that all five studied species formed a new clade and were resolved with 100% bootstrap support. This study will provide genomic resources and potential plastid markers for DNA barcoding, further taxonomy and germplasm exploration of Aconitum.

  14. Unique haplotypes of cacao trees as revealed by trnH-psbA chloroplast DNA

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    Nidia Gutiérrez-López

    2016-04-01

    Full Text Available Cacao trees have been cultivated in Mesoamerica for at least 4,000 years. In this study, we analyzed sequence variation in the chloroplast DNA trnH-psbA intergenic spacer from 28 cacao trees from different farms in the Soconusco region in southern Mexico. Genetic relationships were established by two analysis approaches based on geographic origin (five populations and genetic origin (based on a previous study. We identified six polymorphic sites, including five insertion/deletion (indels types and one transversion. The overall nucleotide diversity was low for both approaches (geographic = 0.0032 and genetic = 0.0038. Conversely, we obtained moderate to high haplotype diversity (0.66 and 0.80 with 10 and 12 haplotypes, respectively. The common haplotype (H1 for both networks included cacao trees from all geographic locations (geographic approach and four genetic groups (genetic approach. This common haplotype (ancient derived a set of intermediate haplotypes and singletons interconnected by one or two mutational steps, which suggested directional selection and event purification from the expansion of narrow populations. Cacao trees from Soconusco region were grouped into one cluster without any evidence of subclustering based on AMOVA (FST = 0 and SAMOVA (FST = 0.04393 results. One population (Mazatán showed a high haplotype frequency; thus, this population could be considered an important reservoir of genetic material. The indels located in the trnH-psbA intergenic spacer of cacao trees could be useful as markers for the development of DNA barcoding.

  15. Complete sequence and comparative analysis of the chloroplast genome of Plinia trunciflora

    Directory of Open Access Journals (Sweden)

    Maria Eguiluz

    2017-11-01

    Full Text Available Abstract Plinia trunciflora is a Brazilian native fruit tree from the Myrtaceae family, also known as jaboticaba. This species has great potential by its fruit production. Due to the high content of essential oils in their leaves and of anthocyanins in the fruits, there is also an increasing interest by the pharmaceutical industry. Nevertheless, there are few studies focusing on its molecular biology and genetic characterization. We herein report the complete chloroplast (cp genome of P. trunciflora using high-throughput sequencing and compare it to other previously sequenced Myrtaceae genomes. The cp genome of P. trunciflora is 159,512 bp in size, comprising inverted repeats of 26,414 bp and single-copy regions of 88,097 bp (LSC and 18,587 bp (SSC. The genome contains 111 single-copy genes (77 protein-coding, 30 tRNA and four rRNA genes. Phylogenetic analysis using 57 cp protein-coding genes demonstrated that P. trunciflora, Eugenia uniflora and Acca sellowiana form a cluster with closer relationship to Syzygium cumini than with Eucalyptus. The complete cp sequence reported here can be used in evolutionary and population genetics studies, contributing to resolve the complex taxonomy of this species and fill the gap in genetic characterization.

  16. Complete sequence and comparative analysis of the chloroplast genome of Plinia trunciflora

    Science.gov (United States)

    Eguiluz, Maria; Yuyama, Priscila Mary; Guzman, Frank; Rodrigues, Nureyev Ferreira; Margis, Rogerio

    2017-01-01

    Abstract Plinia trunciflora is a Brazilian native fruit tree from the Myrtaceae family, also known as jaboticaba. This species has great potential by its fruit production. Due to the high content of essential oils in their leaves and of anthocyanins in the fruits, there is also an increasing interest by the pharmaceutical industry. Nevertheless, there are few studies focusing on its molecular biology and genetic characterization. We herein report the complete chloroplast (cp) genome of P. trunciflora using high-throughput sequencing and compare it to other previously sequenced Myrtaceae genomes. The cp genome of P. trunciflora is 159,512 bp in size, comprising inverted repeats of 26,414 bp and single-copy regions of 88,097 bp (LSC) and 18,587 bp (SSC). The genome contains 111 single-copy genes (77 protein-coding, 30 tRNA and four rRNA genes). Phylogenetic analysis using 57 cp protein-coding genes demonstrated that P. trunciflora, Eugenia uniflora and Acca sellowiana form a cluster with closer relationship to Syzygium cumini than with Eucalyptus. The complete cp sequence reported here can be used in evolutionary and population genetics studies, contributing to resolve the complex taxonomy of this species and fill the gap in genetic characterization. PMID:29111566

  17. The Complete Chloroplast Genome of Wild Rice (Oryza minuta) and Its Comparison to Related Species.

    Science.gov (United States)

    Asaf, Sajjad; Waqas, Muhammad; Khan, Abdul L; Khan, Muhammad A; Kang, Sang-Mo; Imran, Qari M; Shahzad, Raheem; Bilal, Saqib; Yun, Byung-Wook; Lee, In-Jung

    2017-01-01

    Oryza minuta , a tetraploid wild relative of cultivated rice (family Poaceae), possesses a BBCC genome and contains genes that confer resistance to bacterial blight (BB) and white-backed (WBPH) and brown (BPH) plant hoppers. Based on the importance of this wild species, this study aimed to understand the phylogenetic relationships of O. minuta with other Oryza species through an in-depth analysis of the composition and diversity of the chloroplast (cp) genome. The analysis revealed a cp genome size of 135,094 bp with a typical quadripartite structure and consisting of a pair of inverted repeats separated by small and large single copies, 139 representative genes, and 419 randomly distributed microsatellites. The genomic organization, gene order, GC content and codon usage are similar to those of typical angiosperm cp genomes. Approximately 30 forward, 28 tandem and 20 palindromic repeats were detected in the O . minuta cp genome. Comparison of the complete O. minuta cp genome with another eleven Oryza species showed a high degree of sequence similarity and relatively high divergence of intergenic spacers. Phylogenetic analyses were conducted based on the complete genome sequence, 65 shared genes and matK gene showed same topologies and O. minuta forms a single clade with parental O. punctata . Thus, the complete O . minuta cp genome provides interesting insights and valuable information that can be used to identify related species and reconstruct its phylogeny.

  18. Synthesis and transfer of galactolipids in the chloroplast envelope membranes of Arabidopsis thaliana.

    Science.gov (United States)

    Kelly, Amélie A; Kalisch, Barbara; Hölzl, Georg; Schulze, Sandra; Thiele, Juliane; Melzer, Michael; Roston, Rebecca L; Benning, Christoph; Dörmann, Peter

    2016-09-20

    Galactolipids [monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG)] are the hallmark lipids of photosynthetic membranes. The galactolipid synthases MGD1 and DGD1 catalyze consecutive galactosyltransfer reactions but localize to the inner and outer chloroplast envelopes, respectively, necessitating intermembrane lipid transfer. Here we show that the N-terminal sequence of DGD1 (NDGD1) is required for galactolipid transfer between the envelopes. Different diglycosyllipid synthases (DGD1, DGD2, and Chloroflexus glucosyltransferase) were introduced into the dgd1-1 mutant of Arabidopsis in fusion with N-terminal extensions (NDGD1 and NDGD2) targeting to the outer envelope. Reconstruction of DGDG synthesis in the outer envelope membrane was observed only with diglycosyllipid synthase fusion proteins carrying NDGD1, indicating that NDGD1 enables galactolipid translocation between envelopes. NDGD1 binds to phosphatidic acid (PA) in membranes and mediates PA-dependent membrane fusion in vitro. These findings provide a mechanism for the sorting and selective channeling of lipid precursors between the galactolipid pools of the two envelope membranes.

  19. Challenge towards plant recombinant protein expression: instability in nuclear and chloroplast transformation

    International Nuclear Information System (INIS)

    Amiri, M.; Jalali-Javaran, M.; Ehsani, P.; Haddad, R.

    2016-01-01

    It is crucial to maintain the stability of transgene and its expression level. It seems the transformation method and the target organ can influence this instability. To this aim, two transformation systems, Agrobacterium-mediated and particle bombardment systems which have been applied to introduce tissue plasminogen activator (tPA) into nuclear and chloroplast respectively, have been compared to determine transformation efficiency and tPA expression and stability. The presence of tPA gene in transformants has been confirmed by PCR analysis. The gene expression in nuclear transformants and homoplasmy in transplastomic plants have been assayed by ELISA and southern blot, respectively. Some of the Agrobacterium-derived transformants have shown the heritability and stability of the integrated T-DNA harboring the transgene which encodes the tissue plasminogen activator and instability of its expression in T1 generation. Using Southern blot analysis of bombardment-mediated transformants has surprisingly led to detecting the inheritability of tPA. There are several factors lead to silencing of transgene in transgenic plants which should be considered. Possible reasons for these silencing are like vector designing, methylation, copy number, and genome rearrangement.

  20. Phylogenetic relationships of Vepris (Rutaceae inferred from chloroplast, nuclear, and morphological data.

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    Cynthia M Morton

    Full Text Available The tribe Toddalieae Hook. F. (Rutaceae has been controversial since its inception by Bentham and Hooker. The nine taxa examined, Acronychia J.R. & G.Foster, Diphasia Pierre, Diphasiopsis Mendonca, Fagaropsis Mildbr.ex. Siebenl., Oricia Pierre, Teclea Delile, Toddaliopsis Engl., Toddalia Juss. and Vepris Comm. ex. A. Juss, have been recognized under the tribe Toddalieae or Tribes Acronychia, Phellodendron and Toddalia. More recently Araliopsis Engl., Diphasia, Diphasiopsis, Oricia, Teclea, and Toddaliopsis have been incorporated into the genus Vepris, while Toddalia and Fagaropsis have continued to be recognized as closely related. For this study, sequence data of one non-coding chloroplast region (trnL-F and one nuclear region (ITS and various morphological characters, based on Mziray's taxonomic studies were examined to try to elucidate these relationships. This study found that the taxa Diphasia, Diphasiopsis, Oricia, Teclea, Toddaliopsis, Vepris, Toddalia eugeniifolia Engl. and Toddalia glomerata F. Hoffm. form a monophyletic group. Due to the amount of intrageneric and intraspecific variation, species delimitations were difficult to determine; however, these genera should be united into Vepris. The analyses also confirmed that Toddalia asiatica (L. Lam., Zanthoxylon sp. and Fagaropsis angolensis (Engl. H.M. Gardner are the closest relatives to this group.

  1. Chloroplast Redox Imbalance Governs Phenotypic Plasticity: the Grand Design of Photosynthesis Revisited

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    Norman eHuner

    2012-11-01

    Full Text Available 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 PSII 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 Arabidopsis thaliana are discussed in terms of retrograde regulation and the grand design of photosynthesis originally proposed by Daniel Arnon in 1982.

  2. Temperature dependence of energy-transducing functions and inhibitor sensitivity in chloroplasts

    Energy Technology Data Exchange (ETDEWEB)

    Schuurmans, J.J.; Veerman, E.C.I.; Francke, J.A.; Torres-Pereira, J.M.G.; Kraayenhof, R.

    1984-01-01

    A comparative analysis of the temperature dependence of energy-transducing reactions in spinach (Spinacia oleracea) chloroplasts and their sensitivity for uncouplers and energy-transfer inhibitors at different temperatures is presented. Arrhenius plots reveal two groups of transitions, around 19/sup 0/C and around 12/sup 0/C. Activities that show transitions around 19/sup 0/C include linear electron flow from water to ferricyanide, its coupled photophosphorylation, the dark-release of the fluorescent probe atebrin, and the slow component of the 515 nm (carotenoid) absorbance decay after a flash. The transitions around 12/sup 0/C are observed with pyocyanine-mediated cyclic photophosphorylation, light- and dithioerythritol-activated ATP hydrolysis, the dark-release of protons, and the fast 515 nm decay component. It is suggested that both groups of temperature transitions are determined by proton displacements in different domains of the exposed thylakoid membranes. The effects of various uncouplers and an energy-transfer inhibitor are temperature dependent. Some uncouplers also show a different relative inhibition of proton uptake and ATP synthesis at lower temperatures. The efficiency of energy transduction (ATP/e/sub 3/) varied with temperature and was optimal around 10/sup 0/C.

  3. Challenge towards plant recombinant protein expression: instability in nuclear and chloroplast transformation

    Energy Technology Data Exchange (ETDEWEB)

    Amiri, M.; Jalali-Javaran, M.; Ehsani, P.; Haddad, R.

    2016-07-01

    It is crucial to maintain the stability of transgene and its expression level. It seems the transformation method and the target organ can influence this instability. To this aim, two transformation systems, Agrobacterium-mediated and particle bombardment systems which have been applied to introduce tissue plasminogen activator (tPA) into nuclear and chloroplast respectively, have been compared to determine transformation efficiency and tPA expression and stability. The presence of tPA gene in transformants has been confirmed by PCR analysis. The gene expression in nuclear transformants and homoplasmy in transplastomic plants have been assayed by ELISA and southern blot, respectively. Some of the Agrobacterium-derived transformants have shown the heritability and stability of the integrated T-DNA harboring the transgene which encodes the tissue plasminogen activator and instability of its expression in T1 generation. Using Southern blot analysis of bombardment-mediated transformants has surprisingly led to detecting the inheritability of tPA. There are several factors lead to silencing of transgene in transgenic plants which should be considered. Possible reasons for these silencing are like vector designing, methylation, copy number, and genome rearrangement.

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

    Science.gov (United States)

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

    2015-04-27

    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.

  5. Complete sequence and comparative analysis of the chloroplast genome of Plinia trunciflora.

    Science.gov (United States)

    Eguiluz, Maria; Yuyama, Priscila Mary; Guzman, Frank; Rodrigues, Nureyev Ferreira; Margis, Rogerio

    2017-01-01

    Plinia trunciflora is a Brazilian native fruit tree from the Myrtaceae family, also known as jaboticaba. This species has great potential by its fruit production. Due to the high content of essential oils in their leaves and of anthocyanins in the fruits, there is also an increasing interest by the pharmaceutical industry. Nevertheless, there are few studies focusing on its molecular biology and genetic characterization. We herein report the complete chloroplast (cp) genome of P. trunciflora using high-throughput sequencing and compare it to other previously sequenced Myrtaceae genomes. The cp genome of P. trunciflora is 159,512 bp in size, comprising inverted repeats of 26,414 bp and single-copy regions of 88,097 bp (LSC) and 18,587 bp (SSC). The genome contains 111 single-copy genes (77 protein-coding, 30 tRNA and four rRNA genes). Phylogenetic analysis using 57 cp protein-coding genes demonstrated that P. trunciflora, Eugenia uniflora and Acca sellowiana form a cluster with closer relationship to Syzygium cumini than with Eucalyptus. The complete cp sequence reported here can be used in evolutionary and population genetics studies, contributing to resolve the complex taxonomy of this species and fill the gap in genetic characterization.