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Sample records for chlamydomonas mutants display

  1. Reverse genetics in Chlamydomonas: a platform for isolating insertional mutants

    OpenAIRE

    de Montaigu Amaury; Magneschi Leonardo; Catalanotti Claudia; Yang Wenqiang; Mus Florence; Pootakham Wirulda; Gonzalez-Ballester David; Higuera Jose J; Prior Matthew; Galván Aurora; Fernandez Emilio; Grossman Arthur R

    2011-01-01

    Abstract A method was developed to identify insertional mutants of Chlamydomonas reinhardtii disrupted for selected target genes. The approach relies on the generation of thousands of transformants followed by PCR-based screenings that allow for identification of strains harboring the introduced marker gene within specific genes of interest. Our results highlight the strengths and limitations of two independent screens that differed in the nature of the marker DNA used (PCR-amplified fragment...

  2. Altered Fermentative Metabolism in Chlamydomonas reinhardtii Mutants Lacking Pyruvate Formate Lyase and Both Pyruvate Formate Lyase and Alcohol Dehydrogenase

    Energy Technology Data Exchange (ETDEWEB)

    Catalanotti, C.; Dubini, A.; Subramanian, V.; Yang, W. Q.; Magneschi, L.; Mus, F.; Seibert, M.; Posewitz, M. C.; Grossman, A. R.

    2012-02-01

    Chlamydomonas reinhardtii, a unicellular green alga, often experiences hypoxic/anoxic soil conditions that activate fermentation metabolism. We isolated three Chlamydomonas mutants disrupted for the pyruvate formate lyase (PFL1) gene; the encoded PFL1 protein catalyzes a major fermentative pathway in wild-type Chlamydomonas cells. When the pfl1 mutants were subjected to dark fermentative conditions, they displayed an increased flux of pyruvate to lactate, elevated pyruvate decarboxylation, ethanol accumulation, diminished pyruvate oxidation by pyruvate ferredoxin oxidoreductase, and lowered H2 production. The pfl1-1 mutant also accumulated high intracellular levels of lactate, succinate, alanine, malate, and fumarate. To further probe the system, we generated a double mutant (pfl1-1 adh1) that is unable to synthesize both formate and ethanol. This strain, like the pfl1 mutants, secreted lactate, but it also exhibited a significant increase in the levels of extracellular glycerol, acetate, and intracellular reduced sugars and a decrease in dark, fermentative H2 production. Whereas wild-type Chlamydomonas fermentation primarily produces formate and ethanol, the double mutant reroutes glycolytic carbon to lactate and glycerol. Although the metabolic adjustments observed in the mutants facilitate NADH reoxidation and sustained glycolysis under dark, anoxic conditions, the observed changes could not have been predicted given our current knowledge of the regulation of fermentation metabolism.

  3. Reverse genetics in Chlamydomonas: a platform for isolating insertional mutants

    Directory of Open Access Journals (Sweden)

    de Montaigu Amaury

    2011-07-01

    Full Text Available Abstract A method was developed to identify insertional mutants of Chlamydomonas reinhardtii disrupted for selected target genes. The approach relies on the generation of thousands of transformants followed by PCR-based screenings that allow for identification of strains harboring the introduced marker gene within specific genes of interest. Our results highlight the strengths and limitations of two independent screens that differed in the nature of the marker DNA used (PCR-amplified fragment containing the plasmid-free marker versus entire linearized plasmid with the marker and in the strategies used to maintain and store transformants.

  4. Altered Fermentative Metabolism in Chlamydomonas reinhardtii Mutants Lacking Pyruvate Formate Lyase and Both Pyruvate Formate Lyase and Alcohol Dehydrogenase[W

    Science.gov (United States)

    Catalanotti, Claudia; Dubini, Alexandra; Subramanian, Venkataramanan; Yang, Wenqiang; Magneschi, Leonardo; Mus, Florence; Seibert, Michael; Posewitz, Matthew C.; Grossman, Arthur R.

    2012-01-01

    Chlamydomonas reinhardtii, a unicellular green alga, often experiences hypoxic/anoxic soil conditions that activate fermentation metabolism. We isolated three Chlamydomonas mutants disrupted for the pyruvate formate lyase (PFL1) gene; the encoded PFL1 protein catalyzes a major fermentative pathway in wild-type Chlamydomonas cells. When the pfl1 mutants were subjected to dark fermentative conditions, they displayed an increased flux of pyruvate to lactate, elevated pyruvate decarboxylation, ethanol accumulation, diminished pyruvate oxidation by pyruvate ferredoxin oxidoreductase, and lowered H2 production. The pfl1-1 mutant also accumulated high intracellular levels of lactate, succinate, alanine, malate, and fumarate. To further probe the system, we generated a double mutant (pfl1-1 adh1) that is unable to synthesize both formate and ethanol. This strain, like the pfl1 mutants, secreted lactate, but it also exhibited a significant increase in the levels of extracellular glycerol, acetate, and intracellular reduced sugars and a decrease in dark, fermentative H2 production. Whereas wild-type Chlamydomonas fermentation primarily produces formate and ethanol, the double mutant reroutes glycolytic carbon to lactate and glycerol. Although the metabolic adjustments observed in the mutants facilitate NADH reoxidation and sustained glycolysis under dark, anoxic conditions, the observed changes could not have been predicted given our current knowledge of the regulation of fermentation metabolism. PMID:22353371

  5. An Indexed, Mapped Mutant Library Enables Reverse Genetics Studies of Biological Processes in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Li, Xiaobo; Zhang, Ru; Patena, Weronika; Gang, Spencer S; Blum, Sean R; Ivanova, Nina; Yue, Rebecca; Robertson, Jacob M; Lefebvre, Paul A; Fitz-Gibbon, Sorel T; Grossman, Arthur R; Jonikas, Martin C

    2016-02-01

    The green alga Chlamydomonas reinhardtii is a leading unicellular model for dissecting biological processes in photosynthetic eukaryotes. However, its usefulness has been limited by difficulties in obtaining mutants in specific genes of interest. To allow generation of large numbers of mapped mutants, we developed high-throughput methods that (1) enable easy maintenance of tens of thousands of Chlamydomonas strains by propagation on agar media and by cryogenic storage, (2) identify mutagenic insertion sites and physical coordinates in these collections, and (3) validate the insertion sites in pools of mutants by obtaining >500 bp of flanking genomic sequences. We used these approaches to construct a stably maintained library of 1935 mapped mutants, representing disruptions in 1562 genes. We further characterized randomly selected mutants and found that 33 out of 44 insertion sites (75%) could be confirmed by PCR, and 17 out of 23 mutants (74%) contained a single insertion. To demonstrate the power of this library for elucidating biological processes, we analyzed the lipid content of mutants disrupted in genes encoding proteins of the algal lipid droplet proteome. This study revealed a central role of the long-chain acyl-CoA synthetase LCS2 in the production of triacylglycerol from de novo-synthesized fatty acids.

  6. Chlamydomonas fla mutants reveal a link between deflagellation and intraflagellar transport

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    Quarmby Lynne

    2003-08-01

    Full Text Available Abstract Background Cilia and flagella are often lost in anticipation of mitosis or in response to stress. There are two ways that a cell can lose its flagella: resorption or deflagellation. Deflagellation involves active severing of the axoneme at the base of the flagellum; this process is defective in Chlamydomonas fa mutants. In contrast, resorption has been thought to occur as a consequence of constitutive disassembly at the tip in the absence of continued assembly, which requires intraflagellar transport (IFT. Chlamydomonas fla mutants are unable to build and maintain flagella due to defects in IFT. Results fla10 cells, which are defective in kinesin-II, the anterograde IFT motor, resorb their flagella at the restrictive temperature (33°C, as previously reported. We find that in standard media containing ~300 microM calcium, fla10 cells lose flagella by deflagellation at 33°C. This temperature-induced deflagellation of a fla mutant is not predicted by the IFT-based model for flagellar length control. Other fla mutants behave similarly, losing their flagella by deflagellation instead of resorption, if adequate calcium is available. These data suggest a new model whereby flagellar resorption involves active disassembly at the base of the flagellum via a mechanism with components in common with the severing machinery of deflagellation. As predicted by this model, we discovered that deflagellation stimuli induce resorption if deflagellation is blocked either by mutation in a FA gene or by lack of calcium. Further support for this model comes from our discovery that fla10-fa double mutants resorb their flagella more slowly than fla10 mutants. Conclusions Deflagellation of the fla10 mutant at the restrictive temperature is indicative of an active disassembly signal, which can manifest as either resorption or deflagellation. We propose that when IFT is halted by either an inactivating mutation or a cellular signal, active flagellar disassembly

  7. Phosphoregulation of an Inner Dynein Arm Complex in Chlamydomonas reinhardtii Is Altered in Phototactic Mutant Strains

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    King, Stephen J.; Dutcher, Susan K.

    1997-01-01

    To gain a further understanding of axonemal dynein regulation, mutant strains of Chlamydomonas reinhardtii that had defects in both phototactic behavior and flagellar motility were identified and characterized. ptm1, ptm2, and ptm3 mutant strains exhibited motility phenotypes that resembled those of known inner dynein arm region mutant strains, but did not have biochemical or genetic phenotypes characteristic of other inner dynein arm mutations. Three other mutant strains had defects in the f class of inner dynein arms. Dynein extracts from the pf9-4 strain were missing the entire f complex. Strains with mutations in pf9/ida1, ida2, or ida3 failed to assemble the f dynein complex and did not exhibit phototactic behavior. Fractionated dynein from mia1-1 and mia2-1 axonemes exhibited a novel f class inner dynein arm biochemical phenotype; the 138-kD f intermediate chain was present in altered phosphorylation forms. In vitro axonemal dynein activity was reduced by the mia1-1 and mia2-1 mutations. The addition of kinase inhibitor restored axonemal dynein activity concomitant with the dephosphorylation of the 138-kD f intermediate chain. Dynein extracts from uni1-1 axonemes, which specifically assemble only one of the two flagella, contained relatively high levels of the altered phosphorylation forms of the 138-kD intermediate chain. We suggest that the f dynein complex may be phosphoregulated asymmetrically between the two flagella to achieve phototactic turning. PMID:9008712

  8. Characterization of a Mutant Deficient for Ammonium and Nitric Oxide Signalling in the Model System Chlamydomonas reinhardtii.

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    Emanuel Sanz-Luque

    Full Text Available The ubiquitous signalling molecule Nitric Oxide (NO is characterized not only by the variety of organisms in which it has been described, but also by the wealth of biological processes that it regulates. In contrast to the expanding repertoire of functions assigned to NO, however, the mechanisms of NO action usually remain unresolved, and genes that work within NO signalling cascades are seldom identified. A recent addition to the list of known NO functions is the regulation of the nitrogen assimilation pathway in the unicellular alga Chlamydomonas reinhardtii, a well-established model organism for genetic and molecular studies that offers new possibilities in the search for mediators of NO signalling. By further exploiting a collection of Chlamydomonas insertional mutant strains originally isolated for their insensitivity to the ammonium (NH4+ nitrogen source, we found a mutant which, in addition to its ammonium insensitive (AI phenotype, was not capable of correctly sensing the NO signal. Similarly to what had previously been described in the AI strain cyg56, the expression of nitrogen assimilation genes in the mutant did not properly respond to treatments with various NO donors. Complementation experiments showed that NON1 (NO Nitrate 1, a gene that encodes a protein containing no known functional domain, was the gene underlying the mutant phenotype. Beyond the identification of NON1, our findings broadly demonstrate the potential for Chlamydomonas reinhardtii to be used as a model system in the search for novel components of gene networks that mediate physiological responses to NO.

  9. Retrograde Signaling and Photoprotection in a gun4 Mutant of Chlamydomonas reinhardtii

    Institute of Scientific and Technical Information of China (English)

    Cinzia Formighieri; Mauro Ceol; Giulia Bonente; Jean-David Rochaix; Roberto Bassi

    2012-01-01

    GUN4 is a regulatory subunit of Mg-chelatase involved in the control of tetrapyrrole synthesis in plants and cyanobacteria.Here,we report the first characterization of a gun4 insertion mutant of the unicellular green alga Chlamydomonas reinhardtii.The mutant contains 50% of chlorophyll as compared to wild-type and accumulates ProtolX.In contrast to the increase in LHCtranscription,the accumulation of most LHC proteins is drastically diminished,implying posttranscriptional down-regulation in the absence of transcriptional coordination.We found that 803 genes change their expression level in gun4 as compared to wild-type,by RNA-Seq,and this wide-ranging effect on transcription is apparent under physiological conditions.Besides LHCs,we identified transcripts encoding enzymes of the tetrapyrrole pathway and factors involved in signal transduction,transcription,and chromatin remodeling.Moreover,we observe perturbations in electron transport with a strongly decreased PSI-to-PSII ratio.This is accompanied by an enhanced activity of the plastid terminal oxidase (PTOX) that could have a physiological role in decreasing photosystem Ⅱ excitation pressure.

  10. A fluorescence-activated cell sorting-based strategy for rapid isolation of high-lipid Chlamydomonas mutants.

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    Terashima, Mia; Freeman, Elizabeth S; Jinkerson, Robert E; Jonikas, Martin C

    2015-01-01

    There is significant interest in farming algae for the direct production of biofuels and valuable lipids. Chlamydomonas reinhardtii is the leading model system for studying lipid metabolism in green algae, but current methods for isolating mutants of this organism with a perturbed lipid content are slow and tedious. Here, we present the Chlamydomonas high-lipid sorting (CHiLiS) strategy, which enables enrichment of high-lipid mutants by fluorescence-activated cell sorting (FACS) of pooled mutants stained with the lipid-sensitive dye Nile Red. This method only takes 5 weeks from mutagenesis to mutant isolation. We developed a staining protocol that allows quantification of lipid content while preserving cell viability. We improved separation of high-lipid mutants from the wild type by using each cell's chlorophyll fluorescence as an internal control. We initially demonstrated 20-fold enrichment of the known high-lipid mutant sta1 from a mixture of sta1 and wild-type cells. We then applied CHiLiS to sort thousands of high-lipid cells from a pool of about 60,000 mutants. Flow cytometry analysis of 24 individual mutants isolated by this approach revealed that about 50% showed a reproducible high-lipid phenotype. We further characterized nine of the mutants with the highest lipid content by flame ionization detection and mass spectrometry lipidomics. All mutants analyzed had a higher triacylglycerol content and perturbed whole-cell fatty acid composition. One arbitrarily chosen mutant was evaluated by microscopy, revealing larger lipid droplets than the wild type. The unprecedented throughput of CHiLiS opens the door to a systems-level understanding of green algal lipid biology by enabling genome-saturating isolation of mutants in key genes.

  11. Carbon dioxide fixation and photoevolution of hydrogen and oxygen in a mutant of Chlamydomonas lacking Photosystem I

    Energy Technology Data Exchange (ETDEWEB)

    Greenbaum, E.; Lee, J.W.; Tevault, C.V. [Oak Ridge National Lab., TN (United States)] [and others

    1995-09-01

    Sustained photoassimilation of atmospheric CO{sub 2} and simultaneous photoevolution of molecular hydrogen and oxygen has been observed in a Photosystem I deficient mutant B4 of Chlamydomonas reinhardtii that contains only Photosystem II. The data indicate that Photosystem II alone is capable of spanning the potential difference between water oxidation/oxygen evolution and ferredoxin reduction. The rates of both CO{sub 2} fixation and hydrogen and oxygen evolution are similar in the mutant to that of the wild-type C. reinhardtii 137c containing both photosystems. The wild-type had stable photosynthetic activity, measured as CO{sub 2} fixation, under both air and anaerobic conditions, while the mutant was stable only under anaerobic conditions. The results are discussed in terms of the fundamental mechanisms and energetics of photosynthesis and possible implications for the evolution of oxygenic photosynthesis.

  12. Interaction of Herbicides and Quinone with the QB-Protein of the Diuron-Resistant Chlamydomonas reinhardtii Mutant Dr2

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    Haworth, Philip; Steinback, Katherine E.

    1987-01-01

    We have used the diuron-resistant Dr2 mutant of Chlamydomonas reinhardtii which is altered in the 32 kilodalton QB-protein at amino acid 219 (valine to isoleucine), to investigate the interactions of herbicides and plastoquinone with the 32 kilodalton QB-protein. The data contained in this report demonstrate that the effects of this mutation are different from those of the more completely characterized mutant which confers extreme resistance to triazines in higher plants. The mutation in C. reinhardtii Dr2 confers only slight resistance to a number of inhibitors of photosynthetic electron transport. Extreme triazine resistance results from an increase in the binding constant of the herbicide with the 32 kilodalton QB-protein, in contrast the diuron binding constant for chloroplasts isolated from wild-type (sensitive) Chlamydomonas and the resistant Dr2 are indistinguishable. We conclude that the altered structure in the 32 kilodalton QB-protein of Dr2 does not directly affect the diuron binding site. This mutation appears to alter the steric properties of the binding protein in such a way that diuron and plastoquinone do not directly compete for binding. This steric perturbation confers mild resistance to other herbicidal inhibitors of photosynthesis and alters the kinetics of QA to QB electron transfer. PMID:16665318

  13. Nickel and low CO2-controlled motility in Chlamydomonas through complementation of a paralyzed flagella mutant with chemically regulated promoters

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    Rosenbaum Joel L

    2011-01-01

    Full Text Available Abstract Background Chlamydomonas reinhardtii is a model system for the biology of unicellular green algae. Chemically regulated promoters, such as the nickel-inducible CYC6 or the low CO2-inducible CAH1 promoter, may prove useful for expressing, at precise times during its cell cycle, proteins with relevant biological functions, or complementing mutants in genes encoding such proteins. To this date, this has not been reported for the above promoters. Results We fused the CYC6 and CAH1 promoters to an HA-tagged RSP3 gene, encoding a protein of the flagellar radial spoke complex. The constructs were used for chemically regulated complementation of the pf14 mutant, carrying an ochre mutation in the RSP3 gene. 7 to 8% of the transformants showed cells with restored motility after induction with nickel or transfer to low CO2 conditions, but not in non-inducing conditions. Maximum complementation (5% motile cells was reached with very different kinetics (5-6 hours for CAH1, 48 hours for CYC6. The two inducible promoters drive much lower levels of RSP3 protein expression than the constitutive PSAD promoter, which shows almost complete rescue of motility. Conclusions To our knowledge, this is the first example of the use of the CYC6 or CAH1 promoters to perform a chemically regulated complementation of a Chlamydomonas mutant. Based on our data, the CYC6 and CAH1 promoters should be capable of fully complementing mutants in genes whose products exert their biological activity at low concentrations.

  14. Partially Functional Outer-Arm Dynein in a Novel Chlamydomonas Mutant Expressing a Truncated γ Heavy Chain▿

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    Liu, Zhongmei; Takazaki, Hiroko; Nakazawa, Yuki; Sakato, Miho; Yagi, Toshiki; Yasunaga, Takuo; King, Stephen M.; Kamiya, Ritsu

    2008-01-01

    The outer dynein arm of Chlamydomonas flagella contains three heavy chains (α, β, and γ), each of which exhibits motor activity. How they assemble and cooperate is of considerable interest. Here we report the isolation of a novel mutant, oda2-t, whose γ heavy chain is truncated at about 30% of the sequence. While the previously isolated γ chain mutant oda2 lacks the entire outer arm, oda2-t retains outer arms that contain α and β heavy chains, suggesting that the N-terminal sequence (corresponding to the tail region) is necessary and sufficient for stable outer-arm assembly. Thin-section electron microscopy and image analysis localize the γ heavy chain to a basal region of the outer-arm image in the axonemal cross section. The motility of oda2-t is lower than that of the wild type and oda11 (lacking the α heavy chain) but higher than that of oda2 and oda4-s7 (lacking the motor domain of the β heavy chain). Thus, the outer-arm dynein lacking the γ heavy-chain motor domain is partially functional. The availability of mutants lacking individual heavy chains should greatly facilitate studies on the structure and function of the outer-arm dynein. PMID:18487347

  15. Rest mutant zebrafish swim erratically and display atypical spatial preferences.

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    Moravec, Cara E; Li, Edward; Maaswinkel, Hans; Kritzer, Mary F; Weng, Wei; Sirotkin, Howard I

    2015-05-01

    The Rest/Nrsf transcriptional repressor modulates expression of a large set of neural specific genes. Many of these target genes have well characterized roles in nervous system processes including development, plasticity and synaptogenesis. However, the impact of Rest-mediated transcriptional regulation on behavior has been understudied due in part to the embryonic lethality of the mouse knockout. To investigate the requirement for Rest in behavior, we employed the zebrafish rest mutant to explore a range of behaviors in adults and larva. Adult rest mutants of both sexes showed abnormal behaviors in a novel environment including increased vertical swimming, erratic swimming patterns and a proclivity for the tank walls. Adult males also had diminished reproductive success. At 6 days post fertilization (dpf), rest mutant larva were hypoactive, but displayed normal evoked responses to light and sound stimuli. Overall, these results provide evidence that rest dysfunction produces atypical swimming patterns and preferences in adults, and reduced locomotor activity in larvae. This study provides the first behavioral analysis of rest mutants and reveals specific behaviors that are modulated by Rest.

  16. Systems-level analysis of nitrogen starvation-induced modifications of carbon metabolism in a Chlamydomonas reinhardtii starchless mutant.

    Science.gov (United States)

    Blaby, Ian K; Glaesener, Anne G; Mettler, Tabea; Fitz-Gibbon, Sorel T; Gallaher, Sean D; Liu, Bensheng; Boyle, Nanette R; Kropat, Janette; Stitt, Mark; Johnson, Shannon; Benning, Christoph; Pellegrini, Matteo; Casero, David; Merchant, Sabeeha S

    2013-11-01

    To understand the molecular basis underlying increased triacylglycerol (TAG) accumulation in starchless (sta) Chlamydomonas reinhardtii mutants, we undertook comparative time-course transcriptomics of strains CC-4348 (sta6 mutant), CC-4349, a cell wall-deficient (cw) strain purported to represent the parental STA6 strain, and three independent STA6 strains generated by complementation of sta6 (CC-4565/STA6-C2, CC-4566/STA6-C4, and CC-4567/STA6-C6) in the context of N deprivation. Despite N starvation-induced dramatic remodeling of the transcriptome, there were relatively few differences (5 × 10(2)) observed between sta6 and STA6, the most dramatic of which were increased abundance of transcripts encoding key regulated or rate-limiting steps in central carbon metabolism, specifically isocitrate lyase, malate synthase, transaldolase, fructose bisphosphatase and phosphoenolpyruvate carboxykinase (encoded by ICL1, MAS1, TAL1, FBP1, and PCK1 respectively), suggestive of increased carbon movement toward hexose-phosphate in sta6 by upregulation of the glyoxylate pathway and gluconeogenesis. Enzyme assays validated the increase in isocitrate lyase and malate synthase activities. Targeted metabolite analysis indicated increased succinate, malate, and Glc-6-P and decreased Fru-1,6-bisphosphate, illustrating the effect of these changes. Comparisons of independent data sets in multiple strains allowed the delineation of a sequence of events in the global N starvation response in C. reinhardtii, starting within minutes with the upregulation of alternative N assimilation routes and carbohydrate synthesis and subsequently a more gradual upregulation of genes encoding enzymes of TAG synthesis. Finally, genome resequencing analysis indicated that (1) the deletion in sta6 extends into the neighboring gene encoding respiratory burst oxidase, and (2) a commonly used STA6 strain (CC-4349) as well as the sequenced reference (CC-503) are not congenic with respect to sta6 (CC-4348

  17. Carbon allocation and element composition in four Chlamydomonas mutants defective in genes related to the CO2 concentrating mechanism.

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    Memmola, Francesco; Mukherjee, Bratati; Moroney, James V; Giordano, Mario

    2014-09-01

    Four mutants of Chlamydomonas reinhardtii with defects in different components of the CO2 concentrating mechanism (CCM) or in Rubisco activase were grown autotrophically at high pCO2 and then transferred to low pCO2, in order to study the role of different components of the CCM on carbon allocation and elemental composition. To study carbon allocation, we measured the relative size of the main organic pools by Fourier Transform Infrared spectroscopy. Total reflection X-ray fluorescence was used to analyze the elemental composition of algal cells. Our data show that although the organic pools increased their size at high CO2 in all strains, their stoichiometry was highly homeostatic, i.e., the ratios between carbohydrates and proteins, lipid and proteins, and carbohydrates and lipids, did not change significantly. The only exception was the wild-type 137c, in which proteins decreased relative to carbohydrates and lipids, when the cells were transferred to low CO2. It is noticeable that the two wild types used in this study responded differently to the transition from high to low CO2. Malfunctions of the CCM influenced the concentration of several elements, somewhat altering cell elemental stoichiometry: especially the C/P and N/P ratios changed appreciably in almost all strains as a function of the growth CO2 concentration, except in 137c and the Rubisco activase mutant rca1. In strain cia3, defective in the lumenal carbonic anhydrase (CA), the cell quotas of P, S, Ca, Mn, Fe, and Zn were about 5-fold higher at low CO2 than at high CO2. A Principle Components Analysis showed that, mostly because of its elemental composition, cia3 behaved in a substantially different way from all other strains, at low CO2. The lumenal CA thus plays a crucial role, not only for the correct functioning of the CCM, but also for element utilization. Not surprisingly, growth at high CO2 attenuated differences among strains.

  18. Colony mutants of compatible nocardiae displaying variations in recombining capacity.

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    Brownell, G H; Walsh, R S

    1972-03-01

    Colonial morphology mutants of Nocardia erythropolis were isolated following ultraviolet (UV) irradiation. The alleles rou-1/smo-1 were located by recombinant analysis and found to be linked to previously mapped characters. On the basis of recombinant class type patterns obtained from various selective characters it was postulated that the rou-1 allele may span a region of unique nucleotides in the Mat-Ce genome. Recombination frequencies of rou-1 and smo-2 bearing mutants of the Mat-Ce mating type were found to differ by over 1000 fold. Attempts to demonstrate that low recombination frequencies produced by the Smo mutants were due to Rec(-) genes were unsuccessful. No increased sensitivity to either UV or X irradiation was observed by the Smo mutants. Acriflavine treatment of either Rou or Smo colony mutants failed to accelerate reversion or to alter the recombining potentials of the mutants.

  19. Relationships between PSII-independent hydrogen bioproduction and starch metabolism as evidenced from isolation of starch catabolism mutants in the green alga Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Chochois, Vincent; Constans, Laure; Beyly, Audrey; Soliveres, Melanie; Peltier, Gilles; Cournac, Laurent [CEA, DSV, IBEB, Laboratoire de Bioenergetique et Biotechnologie des Bacteries and Microalgues, Saint Paul Lez Durance, F-13108 (France); CNRS, UMR Biologie Vegetale and Microbiologie Environnementales, Saint Paul lez Durance, F-13108 (France); Aix-Marseille Universite, Saint Paul lez Durance, F-13108 (France); Dauvillee, David; Ball, Steven [Univ Lille Nord de France, F-59000 Lille (France); USTL, UGSF, F-59650 Villeneuve d' Ascq (France); CNRS, UMR 8576, F-59650 Villeneuve d' Ascq (France)

    2010-10-15

    Sulfur deprivation, which is considered as an efficient way to trigger long-term hydrogen photoproduction in unicellular green algae has two major effects: a decrease in PSII which allows anaerobiosis to be reached and carbohydrate (starch) storage. Starch metabolism has been proposed as one of the major factors of hydrogen production, particularly during the PSII-independent (or indirect) pathway. While starch biosynthesis has been characterized in the green alga Chlamydomonas reinhardtii, little remains known concerning starch degradation. In order to gain a better understanding of starch catabolism pathways and identify those steps likely to limit the starch-dependent hydrogen production, we have designed a genetic screening procedure aimed at isolating mutants of the green alga C. reinhardtii affected in starch mobilization. Using two different screening protocols, the first one based on aerobic starch degradation in the dark and the second one on anaerobic starch degradation in the light, eighteen mutants were isolated among a library of 15,000 insertion mutants, eight (std1-8) with the first screen and ten (sda1-10) with the second. Most of the mutant strains isolated in this study showed a reduction or a delay in the PSII-independent hydrogen production. Further characterization of these mutants should allow the identification of molecular determinants of starch-dependent hydrogen production and supply targets for future biotechnological improvements. (author)

  20. Identification and molecular characterization of the second Chlamydomonas gun4 mutant, gun4-II [v2; ref status: indexed, http://f1000r.es/1id

    Directory of Open Access Journals (Sweden)

    Phillip B Grovenstein

    2013-07-01

    Full Text Available The green micro-alga Chlamydomonas reinhardtii is an elegant model organism to study oxygenic photosynthesis. Chlorophyll (Chl and heme are major tetrapyrroles that play an essential role in photosynthesis and respiration. These tetrapyrroles are synthesized via a common branched pathway that involves mainly enzymes, encoded by nuclear genes. One of the enzymes in the pathway is Mg chelatase (MgChel. MgChel catalyzes insertion of Mg2+ into protoporphyrin IX (PPIX, proto to form Magnesium-protoporphyrin IX (MgPPIX, Mgproto, the first biosynthetic intermediate in the Chl branch. The GUN4 (genomes uncoupled 4 protein is not essential for the MgChel activity but has been shown to significantly stimulate its activity. We have isolated a light sensitive mutant, 6F14, by random DNA insertional mutagenesis. 6F14 cannot tolerate light intensities higher than 90-100 μmol photons m-2 s-1. It shows a light intensity dependent progressive photo-bleaching. 6F14 is incapable of photo-autotrophic growth under light intensity higher than 100 μmol photons m-2 s-1. PCR based analyses show that in 6F14 the insertion of the plasmid outside the GUN4 locus has resulted in a genetic rearrangement of the GUN4 gene and possible deletions in the genomic region flanking the GUN4 gene. Our gun4 mutant has a Chl content very similar to that in the wild type in the dark and is very sensitive to fluctuations in the light intensity in the environment unlike the earlier identified Chlamydomonas gun4 mutant. Complementation with a functional copy of the GUN4 gene restored light tolerance, Chl biosynthesis and photo-autotrophic growth under high light intensities in 6F14. 6F14 is the second gun4 mutant to be identified in C. reinhardtii. Additionally, we show that our two gun4 complements over-express the GUN4 protein and show a higher Chl content per cell compared to that in the wild type strain.

  1. D1-arginine257 mutants (R257E, K, and Q) of Chlamydomonas reinhardtii have a lowered QB redox potential: analysis of thermoluminescence and fluorescence measurements

    Science.gov (United States)

    Rose, Stuart; Minagawa, Jun; Seufferheld, Manfredo; Padden, Sean; Svensson, Bengt; Kolling, Derrick R. J.; Crofts, Antony R.; Govindjee

    2009-01-01

    Arginine257 (R257), in the de-helix that caps the QB site of the D1 protein, has been shown by mutational studies to play a key role in the sensitivity of Photosystem II (PS II) to bicarbonate-reversible binding of the formate anion. In this article, the role of this residue has been further investigated through D1 mutations (R257E, R257Q, and R257K) in Chlamydomonas reinhardtii. We have investigated the activity of the QB site by studying differences from wild type on the steady-state turnover of PS II, as assayed through chlorophyll (Chl) a fluorescence yield decay after flash excitation. The effects of p-benzoquinone (BQ, which oxidizes reduced QB, QB−) and 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU, which blocks electron flow from QA− to QB) were measured. The equilibrium constants of the two-electron gate were obtained through thermoluminescence measurements. The thermoluminescence properties were changed in the mutants, especially when observed after pretreatment with 100 μM BQ. A theoretical analysis of the thermoluminescence data, based mainly on the recombination pathways model of Rappaport et al. (2005), led to the conclusion that the free-energy difference for the recombination of QB− with S2 was reduced by 20–40 mV in the three mutants (D1-R257K, D1-R257Q, and D1-R257E); this was interpreted to be due to a lowering of the redox potential of QB/QB−. Further, since the recombination of QA− with S2 was unaffected, we suggest that no significant change in redox potential of QA/QA− occurred in these three mutants. The maximum variable Chl a fluorescence yield is lowered in the mutants, in the order R257K > R257Q > R257E, compared to wild type. Our analysis of the binary oscillations in Chl a fluorescence following pretreatment of cells with BQ showed that turnover of the QB site was relatively unaffected in the three mutants. The mutant D1-R257E had the lowest growth rate and steady-state activity and showed the weakest binary oscillations

  2. Light inhibition of mitochondrial respiration in a mutant of Chlamydomonas reinhardtii devoid of ribulose-1,5-bisphosphate carboxylase/oxygenase activity.

    Science.gov (United States)

    Gans, P; Rebeille, F

    1988-01-01

    The effect of light on mitochondrial respiration has been investigated in Chlamydomonas reinhardtii rcl-u-1-10-6C, a mutant devoid of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) activity. No CO2 uptake was observed in the light, confirming that there was no Rubisco activity, but the CO2 evolution rate was diminished by 65 to 80%. This inhibition was ascribable to a decrease in the tricarboxylic acid cycle (Krebs cycle) activity. At the same time, O2 evolution associated with stimulation of the O2 uptake appears. Darkness or addition of DCMU fully reversed the effect of light, indicating that the inhibitory process is linked to photosystem activities. Levels of pyridine nucleotides (NAD(H) and NADP(H)) and adenine nucleotides (ATP and ADP), the most probable mediators of the interaction between photosynthesis and respiration, were measured in dark and in light. During a dark to light transition the level of NADPH increased significantly whereas the NAD(H) pool remained almost fully oxidized. The level of ADP was always extremely low. These results suggest that the inhibition of Krebs cycle activity is due to a competition for cytosolic ADP between chloroplastic photophosphorylations and oxidative phosphorylations.

  3. A nuclear mutant of Chlamydomonas that exhibits increased sensitivity to UV irradiation, reduced recombination of nuclear genes, and altered transmission of chloroplast genes.

    Science.gov (United States)

    Rosen, H; Newman, S M; Boynton, J E; Gillham, N W

    1991-01-01

    Meiotic progeny of Chlamydomonas reinhardtii normally receive chloroplast genomes only from the mt+ parent. However, exceptional zygotes, which transmit the chloroplast genomes of both parents or, more rarely, only those of the mt- parent, arise at a low frequency. Mutations at the mt(+)-linked mat-3 locus were found previously to elevate the transmission of chloroplast genomes from the mt- parent, resulting in a much higher than normal frequency of exceptional zygotes. In this paper we demonstrate that an ultraviolet-sensitive nuclear mutation mapping at the uvsE1 locus, which is unlinked to mating type, also promotes chloroplast genome transmission from the mt- parent. This mutant, which was previously shown to reduce recombination of nuclear genes in meiosis, acts synergistically with the mat-3-3 mutation to produce an extremely high frequency of exceptional zygotes. Through the use of restriction fragment length polymorphisms existing in the chloroplast genomes of C. reinhardtii and the interfertile strain C. smithii, we show that chloroplast DNA fragments from the mt- parent normally begin to disappear shortly after zygote formation. However, this process appears to be blocked totally in the absence of wild-type uvsE1 and mat-3 gene products. Our findings are consistent with the hypothesis that both gene products contribute to the mechanism responsible for uniparental inheritance of the chloroplast genome from the mt+ parent.

  4. Systems-Level Analysis of Nitrogen Starvation–Induced Modifications of Carbon Metabolism in a Chlamydomonas reinhardtii Starchless Mutant[W

    Science.gov (United States)

    Blaby, Ian K.; Glaesener, Anne G.; Mettler, Tabea; Fitz-Gibbon, Sorel T.; Gallaher, Sean D.; Liu, Bensheng; Boyle, Nanette R.; Kropat, Janette; Stitt, Mark; Johnson, Shannon; Benning, Christoph; Pellegrini, Matteo; Casero, David; Merchant, Sabeeha S.

    2013-01-01

    To understand the molecular basis underlying increased triacylglycerol (TAG) accumulation in starchless (sta) Chlamydomonas reinhardtii mutants, we undertook comparative time-course transcriptomics of strains CC-4348 (sta6 mutant), CC-4349, a cell wall–deficient (cw) strain purported to represent the parental STA6 strain, and three independent STA6 strains generated by complementation of sta6 (CC-4565/STA6-C2, CC-4566/STA6-C4, and CC-4567/STA6-C6) in the context of N deprivation. Despite N starvation–induced dramatic remodeling of the transcriptome, there were relatively few differences (5 × 102) observed between sta6 and STA6, the most dramatic of which were increased abundance of transcripts encoding key regulated or rate-limiting steps in central carbon metabolism, specifically isocitrate lyase, malate synthase, transaldolase, fructose bisphosphatase and phosphoenolpyruvate carboxykinase (encoded by ICL1, MAS1, TAL1, FBP1, and PCK1 respectively), suggestive of increased carbon movement toward hexose-phosphate in sta6 by upregulation of the glyoxylate pathway and gluconeogenesis. Enzyme assays validated the increase in isocitrate lyase and malate synthase activities. Targeted metabolite analysis indicated increased succinate, malate, and Glc-6-P and decreased Fru-1,6-bisphosphate, illustrating the effect of these changes. Comparisons of independent data sets in multiple strains allowed the delineation of a sequence of events in the global N starvation response in C. reinhardtii, starting within minutes with the upregulation of alternative N assimilation routes and carbohydrate synthesis and subsequently a more gradual upregulation of genes encoding enzymes of TAG synthesis. Finally, genome resequencing analysis indicated that (1) the deletion in sta6 extends into the neighboring gene encoding respiratory burst oxidase, and (2) a commonly used STA6 strain (CC-4349) as well as the sequenced reference (CC-503) are not congenic with respect to sta6 (CC-4348

  5. Transport and arrangement of the outer-dynein-arm docking complex in the flagella of Chlamydomonas mutants that lack outer dynein arms.

    Science.gov (United States)

    Wakabayashi, K; Takada, S; Witman, G B; Kamiya, R

    2001-04-01

    The outer dynein arms of Chlamydomonas flagella are attached to a precise site on the outer doublet microtubules and repeat at a regular interval of 24 nm. This binding is mediated by the outer dynein arm docking complex (ODA-DC), which is composed of three protein subunits. In this study, antibodies against the 83- and 62-kD subunits (DC83 and DC62) of the ODA-DC were used to analyze its state of association with outer arm components within the cytoplasm, and its localization in the axonemes of oda mutants. Immunoprecipitation indicates that DC83 and DC62 are preassembled within the cytoplasm, but that they are not associated with outer arm dynein. Both proteins are lost or greatly diminished in oda1 and oda3, mutants in the structural genes of DC62 and DC83, respectively, demonstrating that their association is necessary for their stable presence in the cytoplasm. Immunoelectron microscopy indicates that DC83 repeats at 24-nm intervals along the length of the doublet microtubules of oda6, which lacks outer arms; thus, outer arm periodicity may be determined by the ODA-DC. Flagellar regeneration and temporary dikaryon experiments indicate that the ODA-DC can be rapidly transported into the flagellum and assembled on the doublet microtubules independently of the outer arms and independently of flagellar growth. Unexpectedly, the intensity of ODA-DC labeling decreased toward the distal ends of axonemes of oda6 but not wild-type cells, suggesting that the outer arms reciprocally contribute to the assembly/stability of the ODA-DC.

  6. Isolation of anti-T cell receptor scFv mutants by yeast surface display.

    Science.gov (United States)

    Kieke, M C; Cho, B K; Boder, E T; Kranz, D M; Wittrup, K D

    1997-11-01

    Yeast surface display and sorting by flow cytometry have been used to isolate mutants of an scFv that is specific for the Vbeta8 region of the T cell receptor. Selection was based on equilibrium binding by two fluorescently labeled probes, a soluble Vbeta8 domain and an antibody to the c-myc epitope tag present at the carboxy-terminus of the scFv. The mutants that were selected in this screen included a scFv with threefold increased affinity for the Vbeta8 and scFv clones that were bound with reduced affinities by the anti-c-myc antibody. The latter finding indicates that the yeast display system may be used to map conformational epitopes, which cannot be revealed by standard peptide screens. Equilibrium antigen binding constants were estimated within the surface display format, allowing screening of isolated mutants without necessitating subcloning and soluble expression. Only a relatively small library of yeast cells (3 x 10[5]) displaying randomly mutagenized scFv was screened to identify these mutants, indicating that this system will provide a powerful tool for engineering the binding properties of eucaryotic secreted and cell surface proteins.

  7. A rice mutant displaying a heterochronically elongated internode carries a 100 kb deletion

    Institute of Scientific and Technical Information of China (English)

    Mika Hayashi-Tsugane; Masahiko Maekawa; Qian Qian; Hirokazu Kobayashi; Shigeru Iida; Kazuo Tsugane

    2011-01-01

    We have isolated a recessive rice mutant,designated as indeterminate growth(ing),which displays creeping and apparent heterochronic phenotypes in the vegetative period with lanky and winding culms.Rough mapping and subsequent molecular characterization revealed that the ing mutant carries a large deletion,which corresponds to a 103 kb region in the Nipponbare genome,containing nine annotated genes on chromosome 3.Of these annotated genes,the SLRI gene encoding a DELLA protein is the only one that is well characterized in its function,and its null mutation,which is caused by a single base deletion in the middle of the intronless SLR1 gene,confers a slender phenotype that bears close resemblance to the ing mutant phenotype.The primary cause of the ing mutant phenotype is the deletion of the SLR1 gene,and the ing mutant appears to be the first characterized mutant having the entire SLRI sequence deleted.Our results also suggest that the deleted region of 103 kb does not contain an indispensable gene,whose dysfunction must result in a lethal phenotype.

  8. D1-arginine257 mutants (R257E, K, and Q) of Chlamydomonas reinhardtii have a lowered QB redox potential: analysis of thermoluminescence and fluorescence measurements.

    Science.gov (United States)

    Rose, Stuart; Minagawa, Jun; Seufferheld, Manfredo; Padden, Sean; Svensson, Bengt; Kolling, Derrick R J; Crofts, Antony R; Govindjee

    2008-01-01

    Arginine257 (R257), in the de-helix that caps the Q(B) site of the D1 protein, has been shown by mutational studies to play a key role in the sensitivity of Photosystem II (PS II) to bicarbonate-reversible binding of the formate anion. In this article, the role of this residue has been further investigated through D1 mutations (R257E, R257Q, and R257K) in Chlamydomonas reinhardtii. We have investigated the activity of the Q(B) site by studying differences from wild type on the steady-state turnover of PS II, as assayed through chlorophyll (Chl) a fluorescence yield decay after flash excitation. The effects of p-benzoquinone (BQ, which oxidizes reduced Q(B), Q(B)(-) ) and 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU, which blocks electron flow from Q(A)(-) to Q(B)) were measured. The equilibrium constants of the two-electron gate were obtained through thermoluminescence measurements. The thermoluminescence properties were changed in the mutants, especially when observed after pretreatment with 100 microM BQ. A theoretical analysis of the thermoluminescence data, based mainly on the recombination pathways model of Rappaport et al. (2005), led to the conclusion that the free-energy difference for the recombination of Q(B)(-) with S(2) was reduced by 20-40 mV in the three mutants (D1-R257K, D1-R257Q, and D1-R257E); this was interpreted to be due to a lowering of the redox potential of Q(B)/Q(B)(-). Further, since the recombination of Q(A)(-) with S(2) was unaffected, we suggest that no significant change in redox potential of Q(A)/Q(A)(-) occurred in these three mutants. The maximum variable Chl a fluorescence yield is lowered in the mutants, in the order R257K > R257Q > R257E, compared to wild type. Our analysis of the binary oscillations in Chl a fluorescence following pretreatment of cells with BQ showed that turnover of the Q(B) site was relatively unaffected in the three mutants. The mutant D1-R257E had the lowest growth rate and steady-state activity and showed

  9. Streptococcus pyogenes polymyxin B-resistant mutants display enhanced ExPortal integrity.

    Science.gov (United States)

    Port, Gary C; Vega, Luis A; Nylander, Andrew B; Caparon, Michael G

    2014-07-01

    The ExPortal protein secretion organelle in Streptococcus pyogenes is an anionic phospholipid-containing membrane microdomain enriched in Sec translocons and postsecretion protein biogenesis factors. Polymyxin B binds to and disrupts ExPortal integrity, resulting in defective secretion of several toxins. To gain insight into factors that influence ExPortal organization, a genetic screen was conducted to select for spontaneous polymyxin B-resistant mutants displaying enhanced ExPortal integrity. Whole-genome resequencing of 25 resistant mutants revealed from one to four mutations per mutant genome clustered primarily within a core set of 10 gene groups. Construction of mutants with individual deletions or insertions demonstrated that 7 core genes confer resistance and enhanced ExPortal integrity through loss of function, while 3 were likely due to gain of function and/or combinatorial effects. Core resistance genes include a transcriptional regulator of lipid biosynthesis, several genes involved in nutrient acquisition, and a variety of genes involved in stress responses. Two members of the latter class also function as novel regulators of the secreted SpeB cysteine protease. Analysis of the most frequently isolated mutation, a single nucleotide deletion in a track of 9 consecutive adenine residues in pstS, encoding a component of a high-affinity Pi transporter, suggests that this sequence functions as a molecular switch to facilitate stress adaptation. Together, these data suggest the existence of a membrane stress response that promotes enhanced ExPortal integrity and resistance to cationic antimicrobial peptides.

  10. Selection of functional T cell receptor mutants from a yeast surface-display library.

    Science.gov (United States)

    Kieke, M C; Shusta, E V; Boder, E T; Teyton, L; Wittrup, K D; Kranz, D M

    1999-05-11

    The heterodimeric alphabeta T cell receptor (TCR) for antigen is the key determinant of T cell specificity. The structure of the TCR is very similar to that of antibodies, but the engineering of TCRs by directed evolution with combinatorial display libraries has not been accomplished to date. Here, we report that yeast surface display of a TCR was achieved only after the mutation of specific variable region residues. These residues are located in two regions of the TCR, at the interface of the alpha- and beta-chains and in the beta-chain framework region that is thought to be in proximity to the CD3 signal-transduction complex. The mutations are encoded naturally in many antibody variable regions, indicating specific functional differences that have not been appreciated between TCRs and antibodies. The identification of these residues provides an explanation for the inherent difficulties in the display of wild-type TCRs compared with antibodies. Yeast-displayed mutant TCRs bind specifically to the peptide/MHC antigen, enabling engineering of soluble T cell receptors as specific T cell antagonists. This strategy of random mutagenesis followed by selection for surface expression may be of general use in the directed evolution of other eukaryotic proteins that are refractory to display.

  11. Enhanced Adsorption and Recovery of Uranyl Ions by NikR Mutant-Displaying Yeast

    Directory of Open Access Journals (Sweden)

    Kouichi Kuroda

    2014-04-01

    Full Text Available Uranium is one of the most important metal resources, and the technology for the recovery of uranyl ions (UO22+ from aqueous solutions is required to ensure a semi-permanent supply of uranium. The NikR protein is a Ni2+-dependent transcriptional repressor of the nickel-ion uptake system in Escherichia coli, but its mutant protein (NikRm is able to selectively bind uranyl ions in the interface of the two monomers. In this study, NikRm protein with ability to adsorb uranyl ions was displayed on the cell surface of Saccharomyces cerevisiae. To perform the binding of metal ions in the interface of the two monomers, two metal-binding domains (MBDs of NikRm were tandemly fused via linker peptides and displayed on the yeast cell surface by fusion with the cell wall-anchoring domain of yeast α-agglutinin. The NikRm-MBD-displaying yeast cells with particular linker lengths showed the enhanced adsorption of uranyl ions in comparison to the control strain. By treating cells with citrate buffer (pH 4.3, the uranyl ions adsorbed on the cell surface were recovered. Our results indicate that the adsorption system by yeast cells displaying tandemly fused MBDs of NikRm is effective for simple and concentrated recovery of uranyl ions, as well as adsorption of uranyl ions.

  12. Enhanced adsorption and recovery of uranyl ions by NikR mutant-displaying yeast.

    Science.gov (United States)

    Kuroda, Kouichi; Ebisutani, Kazuki; Iida, Katsuya; Nishitani, Takashi; Ueda, Mitsuyoshi

    2014-04-11

    Uranium is one of the most important metal resources, and the technology for the recovery of uranyl ions (UO22+) from aqueous solutions is required to ensure a semi-permanent supply of uranium. The NikR protein is a Ni2+-dependent transcriptional repressor of the nickel-ion uptake system in Escherichia coli, but its mutant protein (NikRm) is able to selectively bind uranyl ions in the interface of the two monomers. In this study, NikRm protein with ability to adsorb uranyl ions was displayed on the cell surface of Saccharomyces cerevisiae. To perform the binding of metal ions in the interface of the two monomers, two metal-binding domains (MBDs) of NikRm were tandemly fused via linker peptides and displayed on the yeast cell surface by fusion with the cell wall-anchoring domain of yeast α-agglutinin. The NikRm-MBD-displaying yeast cells with particular linker lengths showed the enhanced adsorption of uranyl ions in comparison to the control strain. By treating cells with citrate buffer (pH 4.3), the uranyl ions adsorbed on the cell surface were recovered. Our results indicate that the adsorption system by yeast cells displaying tandemly fused MBDs of NikRm is effective for simple and concentrated recovery of uranyl ions, as well as adsorption of uranyl ions.

  13. Efficient Heterologous Transformation of Chlamydomonas reinhardtii npq2 Mutant with the Zeaxanthin Epoxidase Gene Isolated and Characterized from Chlorella zofingiensis

    Directory of Open Access Journals (Sweden)

    Herminia Rodríguez

    2012-09-01

    Full Text Available In the violaxanthin cycle, the violaxanthin de-epoxidase and zeaxanthin epoxidase catalyze the inter-conversion between violaxanthin and zeaxanthin in both plants and green algae. The zeaxanthin epoxidase gene from the green microalga Chlorella zofingiensis (Czzep has been isolated. This gene encodes a polypeptide of 596 amino acids. A single copy of Czzep has been found in the C. zofingiensis genome by Southern blot analysis. qPCR analysis has shown that transcript levels of Czzep were increased after zeaxanthin formation under high light conditions. The functionality of Czzep gene by heterologous genetic complementation in the Chlamydomonas mutant npq2, which lacks zeaxanthin epoxidase (ZEP activity and accumulates zeaxanthin in all conditions, was analyzed. The Czzep gene was adequately inserted in the pSI105 vector and expressed in npq2. The positive transformants were able to efficiently convert zeaxanthin into violaxanthin, as well as to restore their maximum quantum efficiency of the PSII (Fv/Fm. These results show that Chlamydomonas can be an efficient tool for heterologous expression and metabolic engineering for biotechnological applications.

  14. Escherichia coli mar and acrAB mutants display no tolerance to simple alcohols.

    Science.gov (United States)

    Ankarloo, Jonas; Wikman, Susanne; Nicholls, Ian A

    2010-03-31

    The inducible Mar phenotype of Escherichia coli is associated with increased tolerance to multiple hydrophobic antibiotics as well as some highly hydrophobic organic solvents such as cyclohexane, mediated mainly through the AcrAB/TolC efflux system. The influence of water miscible alcohols ethanol and 1-propanol on a Mar constitutive mutant and a mar deletion mutant of E. coli K-12, as well as the corresponding strains carrying the additional acrAB deletion, was investigated. In contrast to hydrophobic solvents, all strains were killed in exponential phase by 1-propanol and ethanol at rates comparable to the parent strain. Thus, the Mar phenotype does not protect E. coli from killing by these more polar solvents. Surprisingly, AcrAB does not contribute to an increased alcohol tolerance. In addition, sodium salicylate, at concentrations known to induce the mar operon, was unable to increase 1-propanol or ethanol tolerance. Rather, the toxicity of both solvents was increased in the presence of sodium salicylate. Collectively, the results imply that the resilience of E. coli to water miscible alcohols, in contrast to more hydrophobic solvents, does not depend upon the AcrAB/TolC efflux system, and suggests a lower limit for substrate molecular size and functionality. Implications for the application of microbiological systems in environments containing high contents of water miscible organic solvents, e.g., phage display screening, are discussed.

  15. Identification and molecular characterization of a Chlamydomonas reinhardtii mutant that shows a light intensity dependent progressive chlorophyll deficiency [v1; ref status: indexed, http://f1000r.es/1b6

    Directory of Open Access Journals (Sweden)

    Phillip B Grovenstein

    2013-06-01

    Full Text Available The green micro-alga Chlamydomonas reinhardtii is an elegant model organism to study all aspects of oxygenic photosynthesis. Chlorophyll (Chl and heme are major tetrapyrroles that play an essential role in energy metabolism in photosynthetic organisms. These tetrapyrroles are synthesized via a common branched pathway that involves mainly nuclear encoded enzymes. One of the enzymes in the pathway is Mg chelatase (MgChel which inserts Mg2+ into protoporphyrin IX (PPIX, proto to form Magnesium-protoporphyrin IX (MgPPIX, Mgproto, the first biosynthetic intermediate in the Chl branch. The GUN4 (genomes uncoupled 4 protein is not essential for the MgChel activity but has been shown to significantly stimulate its activity. We have isolated a light sensitive mutant, 6F14, by random DNA insertional mutagenesis. 6F14 cannot tolerate light intensities higher than 90-100 μmol photons m-2 s-1. It shows a light intensity dependent progressive photo-bleaching. 6F14 is incapable of photo-autotrophic growth under light intensity higher than 100 μmol photons m-2 s-1. PCR based analyses show that in 6F14 the insertion of the plasmid outside the GUN4 locus has resulted in a genetic rearrangement of the GUN4 gene and possible deletions in the genomic region flanking the GUN4 gene. Our gun4 mutant has a Chl content very similar to that in the wild type in the dark and is very sensitive to fluctuations in the light intensity in the environment unlike the earlier identified Chlamydomonas gun4 mutant. Complementation with a functional copy of the GUN4 gene restored light tolerance, Chl biosynthesis and photo-autotrophic growth under high light intensities in 6F14. 6F14 is the second gun4 mutant to be identified in C. reinhardtii. Additionally, we show that our two gun4 complements over-express the GUN4 protein and show a higher Chl content per cell compared to that in the wild type strain.

  16. Autosomal mutants of proton-exposed kidney cells display frequent loss of heterozygosity on nonselected chromosomes.

    Science.gov (United States)

    Grygoryev, Dmytro; Dan, Cristian; Gauny, Stacey; Eckelmann, Bradley; Ohlrich, Anna P; Connolly, Marissa; Lasarev, Michael; Grossi, Gianfranco; Kronenberg, Amy; Turker, Mitchell S

    2014-05-01

    High-energy protons found in the space environment can induce mutations and cancer, which are inextricably linked. We hypothesized that some mutants isolated from proton-exposed kidneys arose through a genome-wide incident that causes loss of heterozygosity (LOH)-generating mutations on multiple chromosomes (termed here genomic LOH). To test this hypothesis, we examined 11 pairs of nonselected chromosomes for LOH events in mutant cells isolated from the kidneys of mice exposed to 4 or 5 Gy of 1 GeV protons. The mutant kidney cells were selected for loss of expression of the chromosome 8-encoded Aprt gene. Genomic LOH events were also assessed in Aprt mutants isolated from isogenic cultured kidney epithelial cells exposed to 5 Gy of protons in vitro. Control groups were spontaneous Aprt mutants and clones isolated without selection from the proton-exposed kidneys or cultures. The in vivo results showed significant increases in genomic LOH events in the Aprt mutants from proton-exposed kidneys when compared with spontaneous Aprt mutants and when compared with nonmutant (i.e., nonselected) clones from the proton-exposed kidneys. A bias for LOH events affecting chromosome 14 was observed in the proton-induced Aprt mutants, though LOH for this chromosome did not confer increased radiation resistance. Genomic LOH events were observed in Aprt mutants isolated from proton-exposed cultured kidney cells; however the incidence was fivefold lower than in Aprt mutants isolated from exposed intact kidneys, suggesting a more permissive environment in the intact organ and/or the evolution of kidney clones prior to their isolation from the tissue. We conclude that proton exposure creates a subset of viable cells with LOH events on multiple chromosomes, that these cells form and persist in vivo, and that they can be isolated from an intact tissue by selection for a mutation on a single chromosome.

  17. Mouse mutants for the nicotinic acetylcholine receptor ß2 subunit display changes in cell adhesion and neurodegeneration response genes.

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    Carol M Rubin

    Full Text Available Mice lacking expression of the ß2 subunit of the neuronal nicotinic acetylcholine receptor (CHRNB2 display abnormal retinal waves and a dispersed projection of retinal ganglion cell (RGC axons to their dorsal lateral geniculate nuclei (dLGNs. Transcriptomes of LGN tissue from two independently generated Chrnb2-/- mutants and from wildtype mice were obtained at postnatal day 4 (P4, during the normal period of segregation of eye-specific afferents to the LGN. Microarray analysis reveals reduced expression of genes located on the cell membrane or in extracellular space, and of genes active in cell adhesion and calcium signaling. In particular, mRNA for cadherin 1 (Cdh1, a known axon growth regulator, is reduced to nearly undetectable levels in the LGN of P4 mutant mice and Lypd2 mRNA is similarly suppressed. Similar analysis of retinal tissue shows increased expression of crumbs 1 (Crb1 and chemokine (C-C motif ligand 21 (Ccl21 mRNAs in Chrnb2-/- mutant animals. Mutations in these genes are associated with retinal neuronal degeneration. The retinas of Chrnb2-/- mutants are normal in appearance, but the increased expression of these genes may also be involved in the abnormal projection patterns of RGC to the LGN. These data may provide the tools to distinguish the interplay between neural activity and molecular expression. Finally, comparison of the transcriptomes of the two different Chrnb2-/- mutant strains reveals the effects of genetic background upon gene expression.

  18. A Novel ABA Insensitive Mutant of Lotus japonicus with a Wilty Phenotype Displays Unaltered Nodulation Regulation

    Institute of Scientific and Technical Information of China (English)

    Bandana Biswas; Pick Kuen Chan; Peter M Gresshoff

    2009-01-01

    An ABA insensitive mutant,Beyma,was isolated in Lotus japonicus MG-20 from an EMS mutagenesis pop-ulation using root growth inhibition to applied ABA as the screening criterion.(The name 'Beyma" was taken from the Australian Aboriginal language,Wagiman,beyma,meaning 'drying up'.) The stable mutant that segregates as a dominant Mendelian mutation is insensitive to ABA induced inhibition of germination,vegetative growth,stomatal opening,as well as nodulation.Tissue ABA levels were normal,suggesting a sensitivity rather than biosynthesis mutation.It is slow-growing (50-70% of wild-type MG-20) and has a near-constitutive wilty phenotype associated with its inability to regulate stomatal opening.Whilst showing a wide range of ABA insensitive phenotypes,Beyma did not show alteration of nodule number control,as,in the absence of added ABA,the number and patterning (but not size) of nodules formed in the mutant were similar to that of MG-20.Split root experiments on MG-20 showed that application of ABA on one side of the root inhibited nodulation locally but not systemically.We propose that ABA is not involved directly in systemic autoregulation of nodulation (AON).

  19. Generation of a phage-display library of single-domain camelid VH H antibodies directed against Chlamydomonas reinhardtii antigens, and characterization of VH Hs binding cell-surface antigens.

    Science.gov (United States)

    Jiang, Wenzhi; Rosenberg, Julian N; Wauchope, Akelia D; Tremblay, Jacqueline M; Shoemaker, Charles B; Weeks, Donald P; Oyler, George A

    2013-11-01

    Single-domain antibodies (sdAbs) are powerful tools for the detection, quantification, purification and subcellular localization of proteins of interest in biological research. We have generated camelid (Lama pacos) heavy chain-only variable VH domain (VH H) libraries against antigens in total cell lysates from Chlamydomonas reinhardtii. The sdAbs in the sera from immunized animals and VH H antibody domains isolated from the library show specificity to C. reinhardtii and lack of reactivity to antigens from four other algae: Chlorella variabilis, Coccomyxa subellipsoidea, Nannochloropsis oceanica and Thalassiosira pseudonana. Antibodies were produced against a diverse representation of antigens as evidenced by sera ELISA and protein-blot analyses. A phage-display library consisting of the VH H region contained at least 10(6) individual transformants, and thus should represent a wide range of C. reinhardtii antigens. The utility of the phage library was demonstrated by using live C. reinhardtii cells to pan for VH H clones with specific recognition of cell-surface epitopes. The lead candidate VH H clones (designated B11 and H10) bound to C. reinhardtii with EC50 values ≤ 0.5 nm. Treatment of cells with VH H B11 fused to the mCherry or green fluorescent proteins allowed brilliant and specific staining of the C. reinhardtii cell wall and analysis of cell-wall genesis during cell division. Such high-complexity VH H antibody libraries for algae will be valuable tools for algal researchers and biotechnologists.

  20. Mutants of circadian-associated PRR genes display a novel and visible phenotype as to light responses during de-etiolation of Arabidopsis thaliana seedlings.

    Science.gov (United States)

    Kato, Takahiko; Murakami, Masaya; Nakamura, Yuko; Ito, Shogo; Nakamichi, Norihito; Yamashino, Takafumi; Mizuno, Takeshi

    2007-03-01

    In Arabidopsis thaliana, it is currently accepted that certain mutants with lesions in clock-associated genes commonly display hallmarked phenotypes with regard to three characteristic biological events: (i) altered rhythmic expression of circadian-controlled genes, (ii) changes in flowering time, and (iii) altered sensitivity to red light in elongation of hypocotyls. During the course of examination of the clock-associated mutants of PSEUDO-RESPONSE REGULATORS, PRRs, including TOC1 (PRR1), we found that they commonly show another visible phenotype of anomalous greening responses upon the onset to light exposure of etiolated seedlings. These findings are indicative of a novel link between circadian rhythms and chloroplast development.

  1. Synthesizing and salvaging NAD: lessons learned from Chlamydomonas reinhardtii.

    Directory of Open Access Journals (Sweden)

    Huawen Lin

    2010-09-01

    Full Text Available The essential coenzyme nicotinamide adenine dinucleotide (NAD+ plays important roles in metabolic reactions and cell regulation in all organisms. Bacteria, fungi, plants, and animals use different pathways to synthesize NAD+. Our molecular and genetic data demonstrate that in the unicellular green alga Chlamydomonas NAD+ is synthesized from aspartate (de novo synthesis, as in plants, or nicotinamide, as in mammals (salvage synthesis. The de novo pathway requires five different enzymes: L-aspartate oxidase (ASO, quinolinate synthetase (QS, quinolate phosphoribosyltransferase (QPT, nicotinate/nicotinamide mononucleotide adenylyltransferase (NMNAT, and NAD+ synthetase (NS. Sequence similarity searches, gene isolation and sequencing of mutant loci indicate that mutations in each enzyme result in a nicotinamide-requiring mutant phenotype in the previously isolated nic mutants. We rescued the mutant phenotype by the introduction of BAC DNA (nic2-1 and nic13-1 or plasmids with cloned genes (nic1-1 and nic15-1 into the mutants. NMNAT, which is also in the de novo pathway, and nicotinamide phosphoribosyltransferase (NAMPT constitute the nicotinamide-dependent salvage pathway. A mutation in NAMPT (npt1-1 has no obvious growth defect and is not nicotinamide-dependent. However, double mutant strains with the npt1-1 mutation and any of the nic mutations are inviable. When the de novo pathway is inactive, the salvage pathway is essential to Chlamydomonas for the synthesis of NAD+. A homolog of the human SIRT6-like gene, SRT2, is upregulated in the NS mutant, which shows a longer vegetative life span than wild-type cells. Our results suggest that Chlamydomonas is an excellent model system to study NAD+ metabolism and cell longevity.

  2. E151 (sym15), a pleiotropic mutant of pea (Pisum sativum L.), displays low nodule number, enhanced mycorrhizae, delayed lateral root emergence, and high root cytokinin levels.

    Science.gov (United States)

    Jones, James M C; Clairmont, Lindsey; Macdonald, Emily S; Weiner, Catherine A; Emery, R J Neil; Guinel, Frédérique C

    2015-07-01

    In legumes, the formation of rhizobial and mycorrhizal root symbioses is a highly regulated process which requires close communication between plant and microorganism. Plant mutants that have difficulties establishing symbioses are valuable tools for unravelling the mechanisms by which these symbioses are formed and regulated. Here E151, a mutant of Pisum sativum cv. Sparkle, was examined to characterize its root growth and symbiotic defects. The symbioses in terms of colonization intensity, functionality of micro-symbionts, and organ dominance were compared between the mutant and wild type. The endogenous cytokinin (CK) and abscisic acid (ABA) levels and the effect of the exogenous application of these two hormones were determined. E151 was found to be a low and delayed nodulator, exhibiting defects in both the epidermal and cortical programmes though a few mature and functional nodules develop. Mycorrhizal colonization of E151 was intensified, although the fungal functionality was impaired. Furthermore, E151 displayed an altered lateral root (LR) phenotype compared with that of the wild type whereby LR emergence is initially delayed but eventually overcome. No differences in ABA levels were found between the mutant and the wild type, but non-inoculated E151 exhibited significantly high CK levels. It is hypothesized that CK plays an essential role in differentially mediating the entry of the two micro-symbionts into the cortex; whereas it would inhibit the entry of the rhizobia in that tissue, it would promote that of the fungus. E151 is a developmental mutant which may prove to be a useful tool in further understanding the role of hormones in the regulation of beneficial root symbioses.

  3. Restriction fragment differential display of pediocin-resistant Listeria monocytogenes 412 mutants shows consistent overexpression of a putative beta-glucoside-specific PTS system.

    Science.gov (United States)

    Gravesen, A; Warthoe, P; Knochel, S; Thirstrup, K

    2000-06-01

    Pediocin PA-1, which is a bacteriocin produced by lactic acid bacteria, has potential as a biopreservative of food. However, such use may lead to the development of resistance in the target organism. Gene expression in two independent pediocin-resistant mutants of Listeria monocytogenes 412 was compared to the original isolate by restriction fragment differential display PCR (RFDD-PCR). This method amplifies cDNA restriction fragments under stringent PCR conditions, enabled by the use of specific primers complementary to ligated adaptor sequences. RFDD-PCR was very well suited for analysis of listerial gene expression, giving reproducible PCR product profiles. Three gene fragments having increased expression in both resistant mutants were identified. All three had homology to components of beta-glucoside-specific phosphoenolpyruvate-dependent phosphotransferase systems (PTS), one fragment having homology to enzyme II permeases, and the two others to phospho-beta-glucosidases. Overexpression of the putative PTS system was consistently observed in 10 additional pediocin-resistant mutants, isolated at different pH, salt content and temperature. The results suggest that RFDD-PCR is a strong approach for the analysis of prokaryotic gene expression and that the putative beta-glucoside-specific PTS system is involved in mediating pediocin resistance.

  4. Triclosan can select for an AdeIJK-overexpressing mutant of Acinetobacter baumannii ATCC 17978 that displays reduced susceptibility to multiple antibiotics.

    Science.gov (United States)

    Fernando, Dinesh M; Xu, Wayne; Loewen, Peter C; Zhanel, George G; Kumar, Ayush

    2014-11-01

    In order to determine if triclosan can select for mutants of Acinetobacter baumannii ATCC 17978 that display reduced susceptibilities to antibiotics, we isolated a triclosan-resistant mutant, A. baumannii AB042, by serial passaging of A. baumannii ATCC 17978 in growth medium supplemented with triclosan. The antimicrobial susceptibility of AB042 was analyzed by the 2-fold serial dilution method. Expression of five different resistance-nodulation-division (RND) pump-encoding genes (adeB, adeG, adeJ, A1S_2818, and A1S_3217), two outer membrane porin-encoding genes (carO and oprD), and the MATE family pump-encoding gene abeM was analyzed using quantitative reverse transcriptase (qRT) PCR. A. baumannii AB042 exhibited elevated resistance to multiple antibiotics, including piperacillin-tazobactam, doxycycline, moxifloxacin, ceftriaxone, cefepime, meropenem, doripenem, ertapenem, ciprofloxacin, aztreonam, tigecycline, and trimethoprim-sulfamethoxazole, in addition to triclosan. Genome sequencing of A. baumannii AB042 revealed a (116)G→V mutation in fabI, the gene encoding the target enzyme for triclosan. Expression analysis of efflux pumps showed overexpression of the AdeIJK pump, and sequencing of adeN, the gene that encodes the repressor of the adeIJK operon, revealed a 73-bp deletion which would cause a premature termination of translation, resulting in an inactive truncated AdeN protein. This work shows that triclosan can select for mutants of A. baumannii that display reduced susceptibilities to multiple antibiotics from chemically distinct classes in addition to triclosan resistance. This multidrug resistance can be explained by the overexpression of the AdeIJK efflux pump.

  5. Loss of CpSRP54 function leads to a truncated light-harvesting antenna size in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Jeong, Jooyeon; Baek, Kwangryul; Kirst, Henning; Melis, Anastasios; Jin, EonSeon

    2017-01-01

    The Chlamydomonas reinhardtii truncated light-harvesting antenna 4 (tla4) DNA transposon mutant has a pale green phenotype, a lower chlorophyll (Chl) per cell and a higher Chl a/b ratio in comparison with the wild type. It required a higher light intensity for the saturation of photosynthesis and displayed a greater per chlorophyll light-saturated rate of oxygen evolution than the wild type. The Chl antenna size of the photosystems in the tla4 mutant was only about 65% of that measured in the wild type. Molecular genetic analysis revealed that a single plasmid DNA insertion disrupted two genes on chromosome 11 of the mutant. A complementation study identified the "chloroplast signal recognition particle 54" gene (CpSRP54), as the lesion causing the tla4 phenotype. Disruption of this gene resulted in partial failure to assemble and, therefore, lower levels of light-harvesting Chl-binding proteins in the C. reinhardtii thylakoids. A comparative in silico 3-D structure-modeling analysis revealed that the M-domain of the CpSRP54 of C. reinhardtii possesses a more extended finger loop structure, due to different amino acid composition, as compared to that of the Arabidopsis CpSRP54. The work demonstrated that CpSRP54 deletion in microalgae can serve to generate tla mutants with a markedly smaller photosystem Chl antenna size, improved solar energy conversion efficiency, and photosynthetic productivity in high-density cultures under bright sunlight conditions.

  6. Drosophila roadblock and Chlamydomonas Lc7

    Science.gov (United States)

    Bowman, Aaron B.; Patel-King, Ramila S.; Benashski, Sharon E.; McCaffery, J. Michael; Goldstein, Lawrence S.B.; King, Stephen M.

    1999-01-01

    Eukaryotic organisms utilize microtubule-dependent motors of the kinesin and dynein superfamilies to generate intracellular movement. To identify new genes involved in the regulation of axonal transport in Drosophila melanogaster, we undertook a screen based upon the sluggish larval phenotype of known motor mutants. One of the mutants identified in this screen, roadblock (robl), exhibits diverse defects in intracellular transport including axonal transport and mitosis. These defects include intra-axonal accumulations of cargoes, severe axonal degeneration, and aberrant chromosome segregation. The gene identified by robl encodes a 97–amino acid polypeptide that is 57% identical (70% similar) to the 105–amino acid Chlamydomonas outer arm dynein–associated protein LC7, also reported here. Both robl and LC7 have homology to several other genes from fruit fly, nematode, and mammals, but not Saccharomyces cerevisiae. Furthermore, we demonstrate that members of this family of proteins are associated with both flagellar outer arm dynein and Drosophila and rat brain cytoplasmic dynein. We propose that roadblock/LC7 family members may modulate specific dynein functions. PMID:10402468

  7. Actin is required for IFT regulation in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Avasthi, Prachee; Onishi, Masayuki; Karpiak, Joel; Yamamoto, Ryosuke; Mackinder, Luke; Jonikas, Martin C; Sale, Winfield S; Shoichet, Brian; Pringle, John R; Marshall, Wallace F

    2014-09-01

    Assembly of cilia and flagella requires intraflagellar transport (IFT), a highly regulated kinesin-based transport system that moves cargo from the basal body to the tip of flagella [1]. The recruitment of IFT components to basal bodies is a function of flagellar length, with increased recruitment in rapidly growing short flagella [2]. The molecular pathways regulating IFT are largely a mystery. Because actin network disruption leads to changes in ciliary length and number, actin has been proposed to have a role in ciliary assembly. However, the mechanisms involved are unknown. In Chlamydomonas reinhardtii, conventional actin is found in both the cell body and the inner dynein arm complexes within flagella [3, 4]. Previous work showed that treating Chlamydomonas cells with the actin-depolymerizing compound cytochalasin D resulted in reversible flagellar shortening [5], but how actin is related to flagellar length or assembly remains unknown. Here we utilize small-molecule inhibitors and genetic mutants to analyze the role of actin dynamics in flagellar assembly in Chlamydomonas reinhardtii. We demonstrate that actin plays a role in IFT recruitment to basal bodies during flagellar elongation and that when actin is perturbed, the normal dependence of IFT recruitment on flagellar length is lost. We also find that actin is required for sufficient entry of IFT material into flagella during assembly. These same effects are recapitulated with a myosin inhibitor, suggesting that actin may act via myosin in a pathway by which flagellar assembly is regulated by flagellar length.

  8. Neutral lipid accumulation at elevated temperature in conditional mutants of two microalgae species

    DEFF Research Database (Denmark)

    Yao, Shuo; Brandt, Anders Bøving; Egsgaard, Helge

    2012-01-01

    mutants of Chlamydomonas reinhardtii and Chlorella vulgaris, nine out of fourty-one mutants in C. reinhardtii and eleven out of fifty-three mutants in C. vulgaris contained increased amounts of neutral lipids, predominantly as triacylglycerols. Upon temperature induced cell-cycle arrest, these mutants...

  9. Individual Flagellar Waveform Affects Collective Behavior of Chlamydomonas reinhardtii.

    Science.gov (United States)

    Kage, Azusa; Mogami, Yoshihiro

    2015-08-01

    Bioconvection is a form of collective motion that occurs spontaneously in the suspension of swimming microorganisms. In a previous study, we quantitatively described the "pattern transition," a phase transition phenomenon that so far has exclusively been observed in bioconvection of the unicellular green alga Chlamydomonas. We suggested that the transition could be induced by changes in the balance between the gravitational and shear-induced torques, both of which act to determine the orientation of the organism in the shear flow. As both of the torques should be affected by the geometry of the Chlamydomonas cell, alteration in the flagellar waveform might change the extent of torque generation by altering overall geometry of the cell. Based on this working hypothesis, we examined bioconvection behavior of two flagellar mutants of Chlamydomonas reinhardtii, ida1 and oda2, making reference to the wild type. Flagella of ida1 beat with an abnormal waveform, while flagella of oda2 show a normal waveform but lower beat frequency. As a result, both mutants had swimming speed of less than 50% of the wild type. ida1 formed bioconvection patterns with smaller spacing than those of wild type and oda2. Two-axis view revealed the periodic movement of the settling blobs of ida1, while oda2 showed qualitatively similar behavior to that of wild type. Unexpectedly, ida1 showed stronger negative gravitaxis than did wild type, while oda2 showed relatively weak gravitaxis. These findings suggest that flagellar waveform, not swimming speed or beat frequency, strongly affect bioconvection behavior in C. reinhardtii.

  10. Molecular techniques to interrogate and edit the Chlamydomonas nuclear genome.

    Science.gov (United States)

    Jinkerson, Robert E; Jonikas, Martin C

    2015-05-01

    The success of the green alga Chlamydomonas reinhardtii as a model organism is to a large extent due to the wide range of molecular techniques that are available for its characterization. Here, we review some of the techniques currently used to modify and interrogate the C. reinhardtii nuclear genome and explore several technologies under development. Nuclear mutants can be generated with ultraviolet (UV) light and chemical mutagens, or by insertional mutagenesis. Nuclear transformation methods include biolistic delivery, agitation with glass beads, and electroporation. Transforming DNA integrates into the genome at random sites, and multiple strategies exist for mapping insertion sites. A limited number of studies have demonstrated targeted modification of the nuclear genome by approaches such as zinc-finger nucleases and homologous recombination. RNA interference is widely used to knock down expression levels of nuclear genes. A wide assortment of transgenes has been successfully expressed in the Chlamydomonas nuclear genome, including transformation markers, fluorescent proteins, reporter genes, epitope tagged proteins, and even therapeutic proteins. Optimized expression constructs and strains help transgene expression. Emerging technologies such as the CRISPR/Cas9 system, high-throughput mutant identification, and a whole-genome knockout library are being developed for this organism. We discuss how these advances will propel future investigations.

  11. New thioredoxin targets in the unicellular photosynthetic eukaryote Chlamydomonas reinhardtii

    Science.gov (United States)

    Lemaire, Stéphane D.; Guillon, Blanche; Le Maréchal, Pierre; Keryer, Eliane; Miginiac-Maslow, Myroslawa; Decottignies, Paulette

    2004-01-01

    Proteomics were used to identify the proteins from the eukaryotic unicellular green alga Chlamydomonas reinhardtii that can be reduced by thioredoxin. These proteins were retained specifically on a thioredoxin affinity column made of a monocysteinic thioredoxin mutant able to form mixed disulfides with its targets. Of a total of 55 identified targets, 29 had been found previously in higher plants or Synechocystis, but 26 were new targets. Biochemical tests were performed on three of them, showing a thioredoxin-dependent activation of isocitrate lyase and isopropylmalate dehydrogenase and a thioredoxin-dependent deactivation of catalase that is redox insensitive in Arabidopsis. In addition, we identified a Ran protein, a previously uncharacterized nuclear target in a photosynthetic organism. The metabolic and evolutionary implications of these findings are discussed. PMID:15123830

  12. Selenoprotein-Transgenic Chlamydomonas reinhardtii

    Directory of Open Access Journals (Sweden)

    Jiazuan Ni

    2013-02-01

    Full Text Available Selenium (Se deficiency is associated with the occurrence of many diseases. However, excessive Se supplementation, especially with inorganic Se, can result in toxicity. Selenoproteins are the major forms of Se in vivo to exert its biological function. Expression of those selenoproteins, especially with the application of a newly developed system, is thus very important for studying the mechanism of Se in nutrition. The use of Chlamydomonas reinhardtii (C. reinhardtii as a biological vector to express an heterogeneous protein is still at the initial stages of development. In order to investigate the possibility of using this system to express selenoproteins, human 15-KDa selenoprotein (Sep15, a small but widely distributed selenoprotein in mammals, was chosen for the expression platform test. Apart from the wild-type human Sep15 gene fragment, two Sep15 recombinants were constructed containing Sep15 open reading frame (ORF and the selenocysteine insertion sequence (SECIS element from either human Sep15 or C. reinhardtii selenoprotein W1, a highly expressed selenoprotein in this alga. Those Sep15-containing plasmids were transformed into C. reinhardtii CC-849 cells. Results showed that Sep15 fragments were successfully inserted into the nuclear genome and expressed Sep15 protein in the cells. The transgenic and wild-type algae demonstrated similar growth curves in low Se culture medium. To our knowledge, this is the first report on expressing human selenoprotein in green alga.

  13. NPM/ALK mutants resistant to ASP3026 display variable sensitivity to alternative ALK inhibitors but succumb to the novel compound PF-06463922.

    Science.gov (United States)

    Mologni, Luca; Ceccon, Monica; Pirola, Alessandra; Chiriano, Gianpaolo; Piazza, Rocco; Scapozza, Leonardo; Gambacorti-Passerini, Carlo

    2015-03-20

    ALK is involved in the onset of several tumors. Crizotinib (XalkoriTM), a potent ALK inhibitor, represents the current front-line treatment for ALK+ NSCLC and shows great clinical efficacy. However, resistant disease often develops after initial response. ASP3026 is a novel second-generation ALK inhibitor with activity on crizotinib-resistant ALK-L1196M gatekeeper mutant. As resistance is likely to be a relevant hurdle for any drug, we sought to determine the resistance profile of ASP3026 in the context of NPM/ALK+ ALCL. We selected six ASP3026-resistant cell lines by culturing human ALCL cells in the presence of increasing concentrations of drug. The established resistant cell lines carry several point mutations in the ALK kinase domain (G1128S, C1156F, I1171N/T, F1174I, N1178H, E1210K and C1156F/D1203N were the most frequent) that are shown to confer resistance to ASP3026 in the Ba/F3 cell model. All mutants were profiled for cross-resistance against a panel of clinically relevant inhibitors including ceritinib, alectinib, crizotinib, AP26113 and PF-06463922. Finally, a genetically heterogeneous ASP3026-resistant cell line was exposed to second-line treatment simulations with all inhibitors. The population evolved according to relative sensitivity of its mutant subclones to the various drugs. Compound PF-06463922 did not allow the outgrowth of any resistant clone, at non-toxic doses.

  14. Cyclic AMP functions as a primary sexual signal in gametes of Chlamydomonas reinhardtii.

    Science.gov (United States)

    Pasquale, S M; Goodenough, U W

    1987-11-01

    When Chlamydomonas reinhardtii gametes of opposite mating type are mixed together, they adhere by a flagella-mediated agglutination that triggers three rapid mating responses: flagellar tip activation, cell wall loss, and mating structure activation accompanied by actin polymerization. Here we show that a transient 10-fold elevation of intracellular cAMP levels is also triggered by sexual agglutination. We further show that gametes of a single mating type can be induced to undergo all three mating responses when presented with exogenous dibutyryl-cAMP (db-cAMP). These events are also induced by cyclic nucleotide phosphodiesterase inhibitors, which elevate endogenous cAMP levels and act synergistically with db-cAMP. Non-agglutinating mutants of opposite mating type will fuse efficiently in the presence of db-cAMP. No activation of mating events is induced by calcium plus ionophores, 8-bromo-cGMP, dibutyryl-cGMP, nigericin at alkaline pH, phorbol esters, or forskolin. H-8, an inhibitor of cyclic nucleotide-dependent protein kinase, inhibits mating events in agglutinating cells and antagonizes the effects of cAMP on non-agglutinating cells. Adenylate cyclase activity was detected in both the gamete cell body and flagella, with the highest specific activity displayed in flagellar membrane fractions. The flagellar membrane adenylate cyclase is preferentially stimulated by Mn++, unresponsive to NaF, GTP, GTP gamma S, AlF4-, and forskolin, and is inhibited by trifluoperazine. Cyclic nucleotide phosphodiesterase activity is also present in flagella. Our observations indicate that cAMP is a sufficient initial signal for all of the known mating reaction events in C. reinhardtii, and suggest that the flagellar cyclase and/or phosphodiesterase may be important loci of control for the agglutination-stimulated production of this signal.

  15. Phototaxis beyond turning: persistent accumulation and response acclimation of the microalga Chlamydomonas reinhardtii

    CERN Document Server

    Arrieta, Jorge; Chioccioli, Maurizio; Polin, Marco; Tuval, Idan

    2016-01-01

    Phototaxis is an important reaction to light displayed by a wide range of motile microorganisms. Flagellated eukaryotic microalgae in particular, like the model organism Chlamydomonas reinhardtii, steer either towards or away from light by a rapid and precisely timed modulation of their flagellar activity. Cell steering, however, is only the beginning of a much longer process which ultimately allows cells to determine their light exposure history. This process is not well understood. Here we present a first quantitative study of the long timescale phototactic motility of Chlamydomonas at both single cell and population levels. Our results reveal that the phototactic strategy adopted by these microorganisms leads to an efficient exposure to light, and that the phototactic response is modulated over typical timescales of tens of seconds. The adaptation dynamics for phototaxis and chlorophyll fluorescence show a striking quantitative agreement, suggesting that photosynthesis controls quantitatively how cells nav...

  16. Ras-mutant cancer cells display B-Raf binding to Ras that activates extracellular signal-regulated kinase and is inhibited by protein kinase A phosphorylation.

    Science.gov (United States)

    Li, Yanping; Takahashi, Maho; Stork, Philip J S

    2013-09-20

    The small G protein Ras regulates proliferation through activation of the mitogen-activated protein (MAP) kinase (ERK) cascade. The first step of Ras-dependent activation of ERK signaling is Ras binding to members of the Raf family of MAP kinase kinase kinases, C-Raf and B-Raf. Recently, it has been reported that in melanoma cells harboring oncogenic Ras mutations, B-Raf does not bind to Ras and does not contribute to basal ERK activation. For other types of Ras-mutant tumors, the relative contributions of C-Raf and B-Raf are not known. We examined non-melanoma cancer cell lines containing oncogenic Ras mutations and express both C-Raf and B-Raf isoforms, including the lung cancer cell line H1299 cells. Both B-Raf and C-Raf were constitutively bound to oncogenic Ras and contributed to Ras-dependent ERK activation. Ras binding to B-Raf and C-Raf were both subject to inhibition by the cAMP-dependent protein kinase PKA. cAMP inhibited the growth of H1299 cells and Ras-dependent ERK activation via PKA. PKA inhibited the binding of Ras to both C-Raf and B-Raf through phosphorylations of C-Raf at Ser-259 and B-Raf at Ser-365, respectively. These studies demonstrate that in non-melanocytic Ras-mutant cancer cells, Ras signaling to B-Raf is a significant contributor to ERK activation and that the B-Raf pathway, like that of C-Raf, is a target for inhibition by PKA. We suggest that cAMP and hormones coupled to cAMP may prove useful in dampening the effects of oncogenic Ras in non-melanocytic cancer cells through PKA-dependent actions on B-Raf as well as C-Raf.

  17. An Exopolysaccharide-Deficient Mutant of Lactobacillus rhamnosus GG Efficiently Displays a Protective Llama Antibody Fragment against Rotavirus on Its Surface.

    Science.gov (United States)

    Álvarez, Beatriz; Krogh-Andersen, Kasper; Tellgren-Roth, Christian; Martínez, Noelia; Günaydın, Gökçe; Lin, Yin; Martín, M Cruz; Álvarez, Miguel A; Hammarström, Lennart; Marcotte, Harold

    2015-09-01

    Rotavirus is the leading cause of infantile diarrhea in developing countries, where it causes a high number of deaths among infants. Two vaccines are available, being highly effective in developed countries although markedly less efficient in developing countries. As a complementary treatment to the vaccines, a Lactobacillus strain producing an anti-rotavirus antibody fragment in the gastrointestinal tract could potentially be used. In order to develop such an alternative therapy, the effectiveness of Lactobacillus rhamnosus GG to produce and display a VHH antibody fragment (referred to as anti-rotavirus protein 1 [ARP1]) on the surface was investigated. L. rhamnosus GG is one of the best-characterized probiotic bacteria and has intrinsic antirotavirus activity. Among four L. rhamnosus GG strains [GG (CMC), GG (ATCC 53103), GG (NCC 3003), and GG (UT)] originating from different sources, only GG (UT) was able to display ARP1 on the bacterial surface. The genomic analysis of strain GG (UT) showed that the genes welE and welF of the EPS cluster are inactivated, which causes a defect in exopolysaccharide (EPS) production, allowing efficient display of ARP1 on its surface. Finally, GG (UT) seemed to confer a level of protection against rotavirus-induced diarrhea similar to that of wild-type GG (NCC 3003) in a mouse pup model, indicating that the EPS may not be involved in the intrinsic antirotavirus activity. Most important, GG (EM233), a derivative of GG (UT) producing ARP1, was significantly more protective than the control strain L. casei BL23.

  18. Kinesin-II is not essential for mitosis and cell growth in Chlamydomonas.

    Science.gov (United States)

    Matsuura, Kumi; Lefebvre, Paul A; Kamiya, Ritsu; Hirono, Masafumi

    2002-08-01

    The FLA10 gene product (Fla10p) in Chlamydomonas, a heterotrimeric kinesin-II, plays a crucial role in flagellar assembly as a motor protein driving intraflagellar transport. This protein has also been suggested to play a role in mitosis based on its localization to mitotic spindle. A role for Fla10p in mitosis has been difficult to test because to date only conditional (temperature-sensitive) mutant alleles were available, and it is not known whether these retain residual function for mitosis at the non-permissive temperature. In this report, we describe a null allele of fla10 produced by insertional mutagenesis. This mutant does not assemble flagella, but proliferates at a rate identical to that of wild type cells. Observation of microtubule organization in the cell body revealed that normal mitotic spindles are formed in dividing mutant cells. Thus, we conclude that FLA10 kinesin plays no significant roles in mitosis.

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

    Science.gov (United States)

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

    2013-03-01

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

  20. Client Proteins and Small Molecule Inhibitors Display Distinct Binding Preferences for Constitutive and Stress-Induced HSP90 Isoforms and Their Conformationally Restricted Mutants.

    Directory of Open Access Journals (Sweden)

    Thomas L Prince

    Full Text Available The two cytosolic/nuclear isoforms of the molecular chaperone HSP90, stress-inducible HSP90α and constitutively expressed HSP90β, fold, assemble and maintain the three-dimensional structure of numerous client proteins. Because many HSP90 clients are important in cancer, several HSP90 inhibitors have been evaluated in the clinic. However, little is known concerning possible unique isoform or conformational preferences of either individual HSP90 clients or inhibitors. In this report, we compare the relative interaction strength of both HSP90α and HSP90β with the transcription factors HSF1 and HIF1α, the kinases ERBB2 and MET, the E3-ubiquitin ligases KEAP1 and RHOBTB2, and the HSP90 inhibitors geldanamycin and ganetespib. We observed unexpected differences in relative client and drug preferences for the two HSP90 isoforms, with HSP90α binding each client protein with greater apparent affinity compared to HSP90β, while HSP90β bound each inhibitor with greater relative interaction strength compared to HSP90α. Stable HSP90 interaction was associated with reduced client activity. Using a defined set of HSP90 conformational mutants, we found that some clients interact strongly with a single, ATP-stabilized HSP90 conformation, only transiently populated during the dynamic HSP90 chaperone cycle, while other clients interact equally with multiple HSP90 conformations. These data suggest different functional requirements among HSP90 clientele that, for some clients, are likely to be ATP-independent. Lastly, the two inhibitors examined, although sharing the same binding site, were differentially able to access distinct HSP90 conformational states.

  1. The Chlamydomonas genome project: a decade on

    Science.gov (United States)

    Blaby, Ian K.; Blaby-Haas, Crysten; Tourasse, Nicolas; Hom, Erik F. Y.; Lopez, David; Aksoy, Munevver; Grossman, Arthur; Umen, James; Dutcher, Susan; Porter, Mary; King, Stephen; Witman, George; Stanke, Mario; Harris, Elizabeth H.; Goodstein, David; Grimwood, Jane; Schmutz, Jeremy; Vallon, Olivier; Merchant, Sabeeha S.; Prochnik, Simon

    2014-01-01

    The green alga Chlamydomonas reinhardtii is a popular unicellular organism for studying photosynthesis, cilia biogenesis and micronutrient homeostasis. Ten years since its genome project was initiated, an iterative process of improvements to the genome and gene predictions has propelled this organism to the forefront of the “omics” era. Housed at Phytozome, the Joint Genome Institute’s (JGI) plant genomics portal, the most up-to-date genomic data include a genome arranged on chromosomes and high-quality gene models with alternative splice forms supported by an abundance of RNA-Seq data. Here, we present the past, present and future of Chlamydomonas genomics. Specifically, we detail progress on genome assembly and gene model refinement, discuss resources for gene annotations, functional predictions and locus ID mapping between versions and, importantly, outline a standardized framework for naming genes. PMID:24950814

  2. Filling Knowledge Gaps in Biological Networks: integrating global approaches to understand H2 metabolism in Chlamydomonas reinhardtii - Final Report

    Energy Technology Data Exchange (ETDEWEB)

    Posewitz, Matthew C

    2011-06-30

    The green alga Chlamydomonas reinhardtii (Chlamydomonas) has numerous genes encoding enzymes that function in fermentative pathways. Among these genes, are the [FeFe]-hydrogenases, pyruvate formate lyase, pyruvate ferredoxin oxidoreductase, acetate kinase, and phosphotransacetylase. We have systematically undertaken a series of targeted mutagenesis approaches to disrupt each of these key genes and omics techniques to characterize alterations in metabolic flux. Funds from DE-FG02-07ER64423 were specifically leveraged to generate mutants with disruptions in the genes encoding the [FeFe]-hydrogenases HYDA1 and HYDA2, pyruvate formate lyase (PFL1), and in bifunctional alcohol/aldehyde alcohol dehydrogenase (ADH1). Additionally funds were used to conduct global transcript profiling experiments of wildtype Chlamydomonas cells, as well as of the hydEF-1 mutant, which is unable to make H2 due to a lesion in the [FeFe]-hydrogenase biosynthetic pathway. In the wildtype cells, formate, acetate and ethanol are the dominant fermentation products with traces of CO2 and H2 also being produced. In the hydEF-1 mutant, succinate production is increased to offset the loss of protons as a terminal electron acceptor. In the pfl-1 mutant, lactate offsets the loss of formate production, and in the adh1-1 mutant glycerol is made instead of ethanol. To further probe the system, we generated a double mutant (pfl1-1 adh1) that is unable to synthesize both formate and ethanol. This strain, like the pfl1 mutants, secreted lactate, but also exhibited a significant increase in the levels of extracellular glycerol, acetate, and intracellular reduced sugars, and a decline in dark, fermentative H2 production. Whereas wild-type Chlamydomonas fermentation primarily produces formate and ethanol, the double mutant performs a complete rerouting of the glycolytic carbon to lactate and glycerol. Lastly, transcriptome data have been analysed for both the wildtype and hydEF-1, that correlate with our

  3. AarF Domain Containing Kinase 3 (ADCK3 Mutant Cells Display Signs of Oxidative Stress, Defects in Mitochondrial Homeostasis and Lysosomal Accumulation.

    Directory of Open Access Journals (Sweden)

    Jason K Cullen

    Full Text Available Autosomal recessive ataxias are a clinically diverse group of syndromes that in some cases are caused by mutations in genes with roles in the DNA damage response, transcriptional regulation or mitochondrial function. One of these ataxias, known as Autosomal Recessive Cerebellar Ataxia Type-2 (ARCA-2, also known as SCAR9/COQ10D4; OMIM: #612016, arises due to mutations in the ADCK3 gene. The product of this gene (ADCK3 is an atypical kinase that is thought to play a regulatory role in coenzyme Q10 (CoQ10 biosynthesis. Although much work has been performed on the S. cerevisiae orthologue of ADCK3, the cellular and biochemical role of its mammalian counterpart, and why mutations in this gene lead to human disease is poorly understood. Here, we demonstrate that ADCK3 localises to mitochondrial cristae and is targeted to this organelle via the presence of an N-terminal localisation signal. Consistent with a role in CoQ10 biosynthesis, ADCK3 deficiency decreased cellular CoQ10 content. In addition, endogenous ADCK3 was found to associate in vitro with recombinant Coq3, Coq5, Coq7 and Coq9, components of the CoQ10 biosynthetic machinery. Furthermore, cell lines derived from ARCA-2 patients display signs of oxidative stress, defects in mitochondrial homeostasis and increases in lysosomal content. Together, these data shed light on the possible molecular role of ADCK3 and provide insight into the cellular pathways affected in ARCA-2 patients.

  4. MEETING: Chlamydomonas Annotation Jamboree - October 2003

    Energy Technology Data Exchange (ETDEWEB)

    Grossman, Arthur R

    2007-04-13

    Shotgun sequencing of the nuclear genome of Chlamydomonas reinhardtii (Chlamydomonas throughout) was performed at an approximate 10X coverage by JGI. Roughly half of the genome is now contained on 26 scaffolds, all of which are at least 1.6 Mb, and the coverage of the genome is ~95%. There are now over 200,000 cDNA sequence reads that we have generated as part of the Chlamydomonas genome project (Grossman, 2003; Shrager et al., 2003; Grossman et al. 2007; Merchant et al., 2007); other sequences have also been generated by the Kasuza sequence group (Asamizu et al., 1999; Asamizu et al., 2000) or individual laboratories that have focused on specific genes. Shrager et al. (2003) placed the reads into distinct contigs (an assemblage of reads with overlapping nucleotide sequences), and contigs that group together as part of the same genes have been designated ACEs (assembly of contigs generated from EST information). All of the reads have also been mapped to the Chlamydomonas nuclear genome and the cDNAs and their corresponding genomic sequences have been reassembled, and the resulting assemblage is called an ACEG (an Assembly of contiguous EST sequences supported by genomic sequence) (Jain et al., 2007). Most of the unique genes or ACEGs are also represented by gene models that have been generated by the Joint Genome Institute (JGI, Walnut Creek, CA). These gene models have been placed onto the DNA scaffolds and are presented as a track on the Chlamydomonas genome browser associated with the genome portal (http://genome.jgi-psf.org/Chlre3/Chlre3.home.html). Ultimately, the meeting grant awarded by DOE has helped enormously in the development of an annotation pipeline (a set of guidelines used in the annotation of genes) and resulted in high quality annotation of over 4,000 genes; the annotators were from both Europe and the USA. Some of the people who led the annotation initiative were Arthur Grossman, Olivier Vallon, and Sabeeha Merchant (with many individual

  5. Identification of the Elusive Pyruvate Reductase of Chlamydomonas reinhardtii Chloroplasts

    Science.gov (United States)

    Burgess, Steven J.; Taha, Hussein; Yeoman, Justin A.; Iamshanova, Oksana; Chan, Kher Xing; Boehm, Marko; Behrends, Volker; Bundy, Jacob G.; Bialek, Wojciech; Murray, James W.; Nixon, Peter J.

    2016-01-01

    Under anoxic conditions the green alga Chlamydomonas reinhardtii activates various fermentation pathways leading to the creation of formate, acetate, ethanol and small amounts of other metabolites including d-lactate and hydrogen. Progress has been made in identifying the enzymes involved in these pathways and their subcellular locations; however, the identity of the enzyme involved in reducing pyruvate to d-lactate has remained unclear. Based on sequence comparisons, enzyme activity measurements, X-ray crystallography, biochemical fractionation and analysis of knock-down mutants, we conclude that pyruvate reduction in the chloroplast is catalyzed by a tetrameric NAD+-dependent d-lactate dehydrogenase encoded by Cre07.g324550. Its expression during aerobic growth supports a possible function as a ‘lactate valve’ for the export of lactate to the mitochondrion for oxidation by cytochrome-dependent d-lactate dehydrogenases and by glycolate dehydrogenase. We also present a revised spatial model of fermentation based on our immunochemical detection of the likely pyruvate decarboxylase, PDC3, in the cytoplasm. PMID:26574578

  6. Ultraviolet modification of Chlamydomonas reinhardtii for carbon capture

    Directory of Open Access Journals (Sweden)

    Gopal NS

    2016-04-01

    Full Text Available Nikhil S Gopal,1 K Sudhakar2 1The Lawrenceville School, Lawrenceville, NJ, USA; 2Bioenergy Laboratory, Malauna Azad National Institute of Technology, Bhopal, India Purpose: Carbon dioxide (CO2 levels have been rising rapidly. Algae are single-cell organisms with highly efficient CO2 uptake mechanisms. Algae yield two to ten times more biomass versus terrestrial plants and can grow nearly anywhere. Large scale CO2 sequestration is not yet sustainable due to high amounts of nitrogen (N and phosphate (P needed to grow algae in media. Methods: Mutant strains of Chlamydomonas reinhardtii were created using ultraviolet light (2.2–3 K J/m2 and natural selection using media with 20%–80% lower N and P compared to standard Sueoka's high salt medium. Strains were selected based upon growth in media concentrations varying from 20% to 80% less N/P compared to control. Biomass was compared to wild-type control (CC-125 using direct counts, optical density dry weight, and mean doubling time. Results: Mean doubling time was 20 and 25 hours in the low N and N/P strains, respectively (vs 66 hours in control. Using direct counts, growth rates of mutant strains of low N and N/P cultures were not statistically different from control (P=0.37 and 0.70, respectively. Conclusion: Two new strains of algae, as well as wild-type control, were able to grow while using 20%–40% less N and P. Ultraviolet light-based modification of algae is an inexpensive and alternative option to genetic engineering techniques. This technique might make larger scale biosequestration possible. Keywords: biosequestration, ultraviolet, carbon sequestration, carbon capture, algae

  7. Light stress and photoprotection in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Erickson, Erika; Wakao, Setsuko; Niyogi, Krishna K

    2015-05-01

    Plants and algae require light for photosynthesis, but absorption of too much light can lead to photo-oxidative damage to the photosynthetic apparatus and sustained decreases in the efficiency and rate of photosynthesis (photoinhibition). Light stress can adversely affect growth and viability, necessitating that photosynthetic organisms acclimate to different environmental conditions in order to alleviate the detrimental effects of excess light. The model unicellular green alga, Chlamydomonas reinhardtii, employs diverse strategies of regulation and photoprotection to avoid, minimize, and repair photo-oxidative damage in stressful light conditions, allowing for acclimation to different and changing environments.

  8. [An experiment with Chlamydomonas reinhardtii on the Kosmos-2044 biosatellite].

    Science.gov (United States)

    Gavrilova, O V; Gabova, A V; Goriainova, L N; Filatova, E V

    1992-01-01

    Space experiment with Chlamydomonas reinhardtii demonstrated that the microgravity effects were noted in Chlamydomonas at both cellular and population levels: in space the cell size is increased, stage of active growth of the culture is extended, it contains the juvenile vegetative motile cells in greater quantities. Ultrastructural analysis indicated that in microgravity the changes in shape, structure and distribution of intracellular organelles and in volume ratio of organelles and cytoplasma are absent. Chlamydomonas data are in line with the results of the Infusoria and Chlorella experiments.

  9. Paternal inheritance of mitochondria in Chlamydomonas.

    Science.gov (United States)

    Nakamura, Soichi

    2010-03-01

    To analyze mitochondrial DNA (mtDNA)inheritance, differences in mtDNA between Chlamydomonas reinhardtii and Chlamydomonas smithii, respiration deficiency and antibiotic resistance were used to distinguish mtDNA origins. The analyses indicated paternal inheritance. However, these experiments raised questions regarding whether paternal inheritance occurred normally.Mitochondrial nucleoids were observed in living zygotes from mating until 3 days after mating and then until progeny formation. However, selective disappearance of nucleoids was not observed. Subsequently, experimental serial backcrosses between the two strains demonstrated strict paternal inheritance. The fate of mt+ and mt- mtDNA was followed using the differences in mtDNA between the two strains. The slow elimination of mt+ mtDNA through zygote maturation in darkness was observed, and later the disappearance of mt+ mtDNA was observed at the beginning of meiosis. To explain the different fates of mtDNA, methylation status was investigated; however, no methylation was detected. Variously constructed diploid cells showed biparental inheritance. Thus, when the mating process occurs normally, paternal inheritance occurs. Mutations disrupting mtDNA inheritance have not yet been isolated. Mutations that disrupt maternal inheritance of chloroplast DNA (cpDNA) do not disrupt inheritance of mtDNA. The genes responsible for mtDNA inheritance are different from those of chloroplasts.

  10. Enzymatic modification by point mutation and functional analysis of an omega-6 fatty acid desaturase from Arctic Chlamydomonas sp.

    Science.gov (United States)

    Jung, Woongsic; Kim, Eun Jae; Han, Se Jong; Kang, Sung-Ho; Choi, Han-Gu; Kim, Sanghee

    2017-02-07

    Arctic Chlamydomonas sp. is a dominant microalgal strain in cold or frozen freshwater in the Arctic region. The full-length open reading frame of the omega-6 fatty acid desaturase gene (AChFAD6) was obtained from the transcriptomic database of Arctic Chlamydomonas sp. from the KOPRI culture collection of polar micro-organisms. Amino acid sequence analysis indicated the presence of three conserved histidine-rich segments as unique characteristics of omega-6 fatty acid desaturases, and three transmembrane regions transported to plastidic membranes by chloroplast transit peptides in the N-terminal region. The AChFAD6 desaturase activity was examined by expressing wild-type and V254A mutant (Mut-AChFAD6) heterologous recombinant proteins. Quantitative gas chromatography indicated that the concentration of linoleic acids in AChFAD6-transformed cells increased more than 3-fold [6.73 ± 0.13 mg g(-1) dry cell weight (DCW)] compared with cells transformed with vector alone. In contrast, transformation with Mut-AChFAD6 increased the concentration of oleic acid to 9.23 ± 0.18 mg g(-1) DCW, indicating a change in enzymatic activity to mimic that of stearoyl-CoA desaturase. These results demonstrate that AChFAD6 of Arctic Chlamydomonas sp. increases membrane fluidity by enhancing denaturation of C18 fatty acids and facilitates production of large quantities of linoleic fatty acids in prokaryotic expression systems.

  11. CSL encodes a leucine-rich-repeat protein implicated in red/violet light signaling to the circadian clock in Chlamydomonas

    Science.gov (United States)

    Kinoshita, Ayumi; Niwa, Yoshimi; Onai, Kiyoshi; Fukuzawa, Hideya; Ishiura, Masahiro

    2017-01-01

    The green alga Chlamydomonas reinhardtii shows various light responses in behavior and physiology. One such photoresponse is the circadian clock, which can be reset by external light signals to entrain its oscillation to daily environmental cycles. In a previous report, we suggested that a light-induced degradation of the clock protein ROC15 is a trigger to reset the circadian clock in Chlamydomonas. However, light signaling pathways of this process remained unclear. Here, we screened for mutants that show abnormal ROC15 diurnal rhythms, including the light-induced protein degradation at dawn, using a luciferase fusion reporter. In one mutant, ROC15 degradation and phase resetting of the circadian clock by light were impaired. Interestingly, the impairments were observed in response to red and violet light, but not to blue light. We revealed that an uncharacterized gene encoding a protein similar to RAS-signaling-related leucine-rich repeat (LRR) proteins is responsible for the mutant phenotypes. Our results indicate that a previously uncharacterized red/violet light signaling pathway is involved in the phase resetting of circadian clock in Chlamydomonas. PMID:28333924

  12. A brief introduction to the model microswimmer {\\it Chlamydomonas reinhardtii}

    CERN Document Server

    Jeanneret, Raphaël; Polin, Marco

    2016-01-01

    The unicellular biflagellate green alga {\\it Chlamydomonas reinhardtii} has been an important model system in biology for decades, and in recent years it has started to attract growing attention also within the biophysics community. Here we provide a concise review of some of the aspects of {\\it Chlamydomonas} biology and biophysics most immediately relevant to physicists that might be interested in starting to work with this versatile microorganism.

  13. Efficient expression of nuclear transgenes in the green alga Chlamydomonas: synthesis of an HIV antigen and development of a new selectable marker.

    Science.gov (United States)

    Barahimipour, Rouhollah; Neupert, Juliane; Bock, Ralph

    2016-03-01

    The unicellular green alga Chlamydomonas reinhardtii has become an invaluable model system in plant biology. There is also considerable interest in developing this microalga into an efficient production platform for biofuels, pharmaceuticals, green chemicals and industrial enzymes. However, the production of foreign proteins in the nucleocytosolic compartment of Chlamydomonas is greatly hampered by the inefficiency of transgene expression from the nuclear genome. We have recently addressed this limitation by isolating mutant algal strains that permit high-level transgene expression and by determining the contributions of GC content and codon usage to gene expression efficiency. Here we have applied these new tools and explored the potential of Chlamydomonas to produce a recombinant biopharmaceutical, the HIV antigen P24. We show that a codon-optimized P24 gene variant introduced into our algal expression strains give rise to recombinant protein accumulation levels of up to 0.25% of the total cellular protein. Moreover, in combination with an expression strain, a resynthesized nptII gene becomes a highly efficient selectable marker gene that facilitates the selection of transgenic algal clones at high frequency. By establishing simple principles of successful transgene expression, our data open up new possibilities for biotechnological research in Chlamydomonas.

  14. Oil accumulation in the model green alga Chlamydomonas reinhardtii: characterization, variability between common laboratory strains and relationship with starch reserves

    Directory of Open Access Journals (Sweden)

    Carrier Patrick

    2011-01-01

    Full Text Available Abstract Background When cultivated under stress conditions, many microalgae species accumulate both starch and oil (triacylglycerols. The model green microalga Chlamydomonas reinhardtii has recently emerged as a model to test genetic engineering or cultivation strategies aiming at increasing lipid yields for biodiesel production. Blocking starch synthesis has been suggested as a way to boost oil accumulation. Here, we characterize the triacylglycerol (TAG accumulation process in Chlamydomonas and quantify TAGs in various wild-type and starchless strains. Results In response to nitrogen deficiency, Chlamydomonas reinhardtii produced TAGs enriched in palmitic, oleic and linoleic acids that accumulated in oil-bodies. Oil synthesis was maximal between 2 and 3 days following nitrogen depletion and reached a plateau around day 5. In the first 48 hours of oil deposition, a ~80% reduction in the major plastidial membrane lipids occurred. Upon nitrogen re-supply, mobilization of TAGs started after starch degradation but was completed within 24 hours. Comparison of oil content in five common laboratory strains (CC124, CC125, cw15, CC1690 and 11-32A revealed a high variability, from 2 μg TAG per million cell in CC124 to 11 μg in 11-32A. Quantification of TAGs on a cell basis in three mutants affected in starch synthesis (cw15sta1-2, cw15sta6 and cw15sta7-1 showed that blocking starch synthesis did not result in TAG over-accumulation compared to their direct progenitor, the arginine auxotroph strain 330. Moreover, no significant correlation was found between cellular oil and starch levels among the twenty wild-type, mutants and complemented strains tested. By contrast, cellular oil content was found to increase steeply with salt concentration in the growth medium. At 100 mM NaCl, oil level similar to nitrogen depletion conditions could be reached in CC124 strain. Conclusion A reference basis for future genetic studies of oil metabolism in Chlamydomonas

  15. Synergism between Inositol Polyphosphates and TOR Kinase Signaling in Nutrient Sensing, Growth Control, and Lipid Metabolism in Chlamydomonas[OPEN

    Science.gov (United States)

    Evans, Bradley S.; Li, Jia; Liu, Yu; Diamond, Spencer

    2016-01-01

    The networks that govern carbon metabolism and control intracellular carbon partitioning in photosynthetic cells are poorly understood. Target of Rapamycin (TOR) kinase is a conserved growth regulator that integrates nutrient signals and modulates cell growth in eukaryotes, though the TOR signaling pathway in plants and algae has yet to be completely elucidated. We screened the unicellular green alga Chlamydomonas reinhardtii using insertional mutagenesis to find mutants that conferred hypersensitivity to the TOR inhibitor rapamycin. We characterized one mutant, vip1-1, that is predicted to encode a conserved inositol hexakisphosphate kinase from the VIP family that pyrophosphorylates phytic acid (InsP6) to produce the low abundance signaling molecules InsP7 and InsP8. Unexpectedly, the rapamycin hypersensitive growth arrest of vip1-1 cells was dependent on the presence of external acetate, which normally has a growth-stimulatory effect on Chlamydomonas. vip1-1 mutants also constitutively overaccumulated triacylglycerols (TAGs) in a manner that was synergistic with other TAG inducing stimuli such as starvation. vip1-1 cells had reduced InsP7 and InsP8, both of which are dynamically modulated in wild-type cells by TOR kinase activity and the presence of acetate. Our data uncover an interaction between the TOR kinase and inositol polyphosphate signaling systems that we propose governs carbon metabolism and intracellular pathways that lead to storage lipid accumulation. PMID:27600537

  16. Flagellar coordination in Chlamydomonas cells held on micropipettes.

    Science.gov (United States)

    Rüffer, U; Nultsch, W

    1998-01-01

    The two flagella of Chlamydomonas are known to beat synchronously: During breaststroke beating they are generally coordinated in a bilateral way while in shock responses during undulatory beating coordination is mostly parallel [Rüffer and Nultsch, 1995: Botanica Acta 108:169-276]. Analysis of a great number of shock responses revealed that in undulatory beats also periods of bilateral coordination are found and that the coordination type may change several times during a shock response, without concomitant changes of the beat envelope and the beat period. In normal wt cells no coordination changes are found during breaststroke beating, but only short temporary asynchronies: During 2 or 3 normal beats of the cis flagellum, the trans flagellum performs 3 or 4 flat beats with a reduced beat envelope and a smaller beat period, resulting in one additional trans beat. Long periods with flat beats of the same shape and beat period are found in both flagella of the non-phototactic mutant ptx1 and in defective wt 622E cells. During these periods, the coordination is parallel, the two flagella beat alternately. A correlation between normal asynchronous trans beats and the parallel-coordinated beats in the presumably cis defective cells and also the undulatory beats is discussed. In the cis defective cells, a perpetual spontaneous change between parallel beats with small beat periods (higher beat frequency) and bilateral beats with greater beat periods (lower beat frequency) are observed and render questionable the existence of two different intrinsic beat frequencies of the two flagella cis and trans. Asynchronies occur spontaneously but may also be induced by light changes, either step-up or step-down, but not by both stimuli in turn as breaststroke flagellar photoresponses (BFPRs). Asynchronies are not involved in phototaxis. They are independent of the BFPRs, which are supposed to be the basis of phototaxis. Both types of coordination must be assumed to be regulated

  17. Dynamic curvature regulation accounts for the symmetric and asymmetric beats of Chlamydomonas flagella

    CERN Document Server

    Sartori, Pablo; Scholich, Andre; Jülicher, Frank; Howard, Jonathon

    2015-01-01

    Axonemal dyneins are the molecular motors responsible for the beating of cilia and flagella. These motors generate sliding forces between adjacent microtubule doublets within the axoneme, the motile cytoskeletal structure inside the flagellum. To create regular, oscillatory beating patterns, the activities of the axonemal dyneins must be coordinated both spatially and temporally. It is thought that coordination is mediated by stresses or strains that build up within the moving axoneme, but it is not known which components of stress or strain are involved, nor how they feed back on the dyneins. To answer this question, we used isolated, reactivate axonemes of the unicellular alga Chlamydomonas as a model system. We derived a theory for beat regulation in a two-dimensional model of the axoneme. We then tested the theory by measuring the beat waveforms of wild type axonemes, which have asymmetric beats, and mutant axonemes, in which the beat is nearly symmetric, using high-precision spatial and temporal imaging....

  18. Investigating the link between fermentative metabolism and hydrogen production in the unicellular green alga Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Burgess, S.J.; Nixon, P.J. [Imperial College London (United Kingdom)

    2010-07-01

    In the model green alga Chlamydomonas reinhardtii, the electrons required for hydrogen production can come from both the biophotolysis of water and from the fermentation of carbohydrate reserves. Anoxia leads to the activation of several fermentative pathways, which produce a number of end products including formic, malic and acetic acid along with ethanol, carbon dioxide and hydrogen. It has been proposed that by switching off competing fermentative pathways hydrogen production can be increased. Therefore the aim of this study was to devise an experimental strategy to down-regulate the expression of enzymes thought to control C. reinhardtii's fermentative metabolism. We demonstrate here that it is possible to use artificial microRNA (amiRNA) technology to generate knock-down mutants with reduced expression of pyruvate formate lyase (PFL1), a key fermentative enzyme in C. reinhardtii. This work opens up new possibilities to improve hydrogen yields through metabolic engineering. (orig.)

  19. Introducing Dunaliella LIP promoter containing light-inducible motifs improves transgenic expression in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Baek, Kwangryul; Lee, Yew; Nam, Onyou; Park, Seunghye; Sim, Sang Jun; Jin, EonSeon

    2016-03-01

    Promoter of the light-inducible protein gene (LIP) of Dunaliella was recently isolated in our laboratory. The aim of this work is to find the light-inducible motif in the Dunaliella LIP promoter and verify its regulatory motif with a Gaussia luciferase reporter gene transformed in Chlamydomonas reinhardtii. 400 bp upstream to the translational start site of the Dunaliella LIP gene was gradually truncated and analyzed for the luciferase expression. Furthermore, this promoter comprising duplicated or triplicated light-responsive motifs was tested for its augmentation of light response. Two putative light-responsive motifs, GT-1 binding motif and sequences over-represented in light-repressed promoters (SORLIP) located in the 200 bp LIP promoter fragment were analyzed for their light responsibility. It is turned out that SORLIP was responsible for the light-inducible activity. With the copy number of SORLIP up to three showed stronger high light response compared with the native LIP promoter fragment. Therefore, we found a light-responsive DNA motif operating in Chlamydomonas and confirm a synthetic promoter including this motif displayed light inducibility in heterologously transformed green algae for the first time. This light-inducible expression system will be applied to various area of algal research including algal biotechnology.

  20. Limiting steps of hydrogen production in Chlamydomonas reinhardtii and Synechocystis PCC 6803 as analysed by light-induced gas exchange transients

    Energy Technology Data Exchange (ETDEWEB)

    Cournac, L.; Mus, F.; Bernard, L.; Guedeney, G.; Peltier, G. [CEA Cadarache, Lab. d' Ecophysiologie de la Photosynthese, Saint Paul lez Durance (France); Vignais, P. [CEA Grenoble, Lab. de Biochimie er Biophysique des Systemes Integres (France)

    2002-12-01

    In the cyanobacterium Synechocystis PCC6803 and in the microalga Chlamydomonas reinhardtii, transient hydrogen photo-production is observed when cells are exposed to light in anoxia. We measured changes in H{sub 2}, O{sub 2}, and CO{sub 2} concentrations using time-resolved mass spectrometry in wild-type and mutant strains of Chlamydomonas and Synechocystis. In both organisms, non-photochemical reduction of the plastoquinone pool is shown to contribute to the initial H{sub 2} photo-production. This pathway, which does not produce O{sub 2}, exhibits a low rate in normal conditions. From the effect of the uncoupler FCCP, we conclude that PS II-independent H{sub 2} production in Chlamydomonas is limited by the trans-thylakoidal proton gradient. In Synechocystis, from the study of a mutant deficient in the NDH-1 complex (M55), we conclude that PS II-independent H{sub 2} production is limited by recycling of NAD(P)H through the NDH-1 complex. Based on these conclusions, we propose strategies for optimising H{sub 2} photo-production in these organisms. (Author)

  1. Three Acyltransferases and Nitrogen-responsive Regulator Are Implicated in Nitrogen Starvation-induced Triacylglycerol Accumulation in Chlamydomonas*

    Science.gov (United States)

    Boyle, Nanette R.; Page, Mark Dudley; Liu, Bensheng; Blaby, Ian K.; Casero, David; Kropat, Janette; Cokus, Shawn J.; Hong-Hermesdorf, Anne; Shaw, Johnathan; Karpowicz, Steven J.; Gallaher, Sean D.; Johnson, Shannon; Benning, Christoph; Pellegrini, Matteo; Grossman, Arthur; Merchant, Sabeeha S.

    2012-01-01

    Algae have recently gained attention as a potential source for biodiesel; however, much is still unknown about the biological triggers that cause the production of triacylglycerols. We used RNA-Seq as a tool for discovering genes responsible for triacylglycerol (TAG) production in Chlamydomonas and for the regulatory components that activate the pathway. Three genes encoding acyltransferases, DGAT1, DGTT1, and PDAT1, are induced by nitrogen starvation and are likely to have a role in TAG accumulation based on their patterns of expression. DGAT1 and DGTT1 also show increased mRNA abundance in other TAG-accumulating conditions (minus sulfur, minus phosphorus, minus zinc, and minus iron). Insertional mutants, pdat1-1 and pdat1-2, accumulate 25% less TAG compared with the parent strain, CC-4425, which demonstrates the relevance of the trans-acylation pathway in Chlamydomonas. The biochemical functions of DGTT1 and PDAT1 were validated by rescue of oleic acid sensitivity and restoration of TAG accumulation in a yeast strain lacking all acyltransferase activity. Time course analyses suggest than a SQUAMOSA promoter-binding protein domain transcription factor, whose mRNA increases precede that of lipid biosynthesis genes like DGAT1, is a candidate regulator of the nitrogen deficiency responses. An insertional mutant, nrr1-1, accumulates only 50% of the TAG compared with the parental strain in nitrogen-starvation conditions and is unaffected by other nutrient stresses, suggesting the specificity of this regulator for nitrogen-deprivation conditions. PMID:22403401

  2. Genetic tools and techniques for Chlamydomonas reinhardtii.

    Science.gov (United States)

    Mussgnug, Jan H

    2015-07-01

    The development of tools has always been a major driving force for the advancement of science. Optical microscopes were the first instruments that allowed discovery and descriptive studies of the subcellular features of microorganisms. Although optical and electron microscopes remained at the forefront of microbiological research tools since their inventions, the advent of molecular genetics brought about questions which had to be addressed with new "genetic tools". The unicellular green microalgal genus Chlamydomonas, especially the most prominent species C. reinhardtii, has become a frequently used model organism for many diverse fields of research and molecular genetic analyses of C. reinhardtii, as well as the available genetic tools and techniques, have become increasingly sophisticated throughout the last decades. The aim of this review is to provide an overview of the molecular key features of C. reinhardtii and summarize the progress related to the development of tools and techniques for genetic engineering of this organism, from pioneering DNA transformation experiments to state-of-the-art techniques for targeted nuclear genome editing and high-throughput screening approaches.

  3. High quality metabolomic data for Chlamydomonas reinhardtii

    Directory of Open Access Journals (Sweden)

    Fiehn Oliver

    2008-04-01

    Full Text Available Abstract The green eukaryote alga Chlamydomonas reinhardtii is a unicellular model to study control of metabolism in a photosynthetic organism. We here present method improvements for metabolite profiling based on GC-TOF mass spectrometry focusing on three parameters: quenching and cell disruption, extract solvent composition and metabolite annotation. These improvements facilitate using smaller cell numbers and hence, smaller culture volumes which enable faster and more precise sampling techniques that eventually lead to a higher number of samples that can be processed, e.g. for time course experiments. Quenching of metabolism was achieved by mixing 1 ml of culture to 1 ml of -70°C cold 70% methanol. After centrifugation, cells were lyophilized and disrupted by milling using 2-6E6 lyophilized cells, around 500-fold less than previously reported. Glass beads were compared to metal balls for milling, and five different extraction solvents were tested. Additionally, all peaks were annotated in an automated way using the GC-TOF database BinBase instead of manual investigation of a single reference chromatogram. Median precision of analysis was used to decide for the eventual procedure which was applied to a proof-of-principle study of time dependent changes of metabolism under standard conditions.

  4. Radial spoke proteins of Chlamydomonas flagella

    Science.gov (United States)

    Yang, Pinfen; Diener, Dennis R.; Yang, Chun; Kohno, Takahiro; Pazour, Gregory J.; Dienes, Jennifer M.; Agrin, Nathan S.; King, Stephen M.; Sale, Winfield S.; Kamiya, Ritsu; Rosenbaum, Joel L.; Witman, George B.

    2007-01-01

    Summary The radial spoke is a ubiquitous component of ‘9+2’ cilia and flagella, and plays an essential role in the control of dynein arm activity by relaying signals from the central pair of microtubules to the arms. The Chlamydomonas reinhardtii radial spoke contains at least 23 proteins, only 8 of which have been characterized at the molecular level. Here, we use mass spectrometry to identify 10 additional radial spoke proteins. Many of the newly identified proteins in the spoke stalk are predicted to contain domains associated with signal transduction, including Ca2+-, AKAP- and nucleotide-binding domains. This suggests that the spoke stalk is both a scaffold for signaling molecules and itself a transducer of signals. Moreover, in addition to the recently described HSP40 family member, a second spoke stalk protein is predicted to be a molecular chaperone, implying that there is a sophisticated mechanism for the assembly of this large complex. Among the 18 spoke proteins identified to date, at least 12 have apparent homologs in humans, indicating that the radial spoke has been conserved throughout evolution. The human genes encoding these proteins are candidates for causing primary ciliary dyskinesia, a severe inherited disease involving missing or defective axonemal structures, including the radial spokes. PMID:16507594

  5. Dynamic regulation of photosynthesis in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Minagawa, Jun; Tokutsu, Ryutaro

    2015-05-01

    Plants and algae have acquired the ability to acclimatize to ever-changing environments to survive. During photosynthesis, light energy is converted by several membrane protein supercomplexes into electrochemical energy, which is eventually used to assimilate CO2 . The efficiency of photosynthesis is modulated by many environmental factors, including temperature, drought, CO2 concentration, and the quality and quantity of light. Recently, our understanding of such regulators of photosynthesis and the underlying molecular mechanisms has increased considerably. The photosynthetic supercomplexes undergo supramolecular reorganizations within a short time after receiving environmental cues. These reorganizations include state transitions that balance the excitation of the two photosystems: qE quenching, which thermally dissipates excess energy at the level of the light-harvesting antenna, and cyclic electron flow, which supplies the increased ATP demanded by CO2 assimilation and the pH gradient to activate qE quenching. This review focuses on the recent findings regarding the environmental regulation of photosynthesis in model organisms, paying particular attention to the unicellular green alga Chlamydomonas reinhardtii, which offer a glimpse into the dynamic behavior of photosynthetic machinery in nature.

  6. Protein-Protein Interactions between Intermediate Chains and the Docking Complex of Chlamydomonas Flagellar Outer Arm Dynein

    Science.gov (United States)

    Ide, Takahiro; Owa, Mikito; King, Stephen M.; Kamiya, Ritsu; Wakabayashi, Ken-ichi

    2013-01-01

    Outer arm dynein (OAD) is bound to specific loci on outer-doublet-microtubules by interactions at two sites: via intermediate chain 1 (IC1) and the outer dynein arm docking complex (ODA-DC). Studies using Chlamydomonas mutants have suggested that the individual sites have rather weak affinities for microtubules, and therefore strong OAD attachment to microtubules is achieved by their cooperation. To test this idea, we examined interactions between IC1, IC2 (another intermediate chain) and ODA-DC using recombinant proteins. Recombinant IC1 and IC2 were found to form a 1:1 complex, and this complex associated with ODA-DC in vitro. Binding of IC1 to mutant axonemes revealed that there are specific binding sites for IC1. From these data, we propose a novel model of OAD-outer doublet association. PMID:23747306

  7. Mastoparan-Induced Cell Death Signalling in Chlamydomonas Reinhardtii

    NARCIS (Netherlands)

    Yordanova, Z.P.; Kapchina-Toteva, V.M.; Woltering, E.J.; Cristescu, S.M.; Harren, F.J.M.; Yakimova, E.T.

    2009-01-01

    The present study was focused on the elucidation of stress-induced cell death signaling events in the unicellular alga Chlamydomonas reinhardtii exposed to treatment with wasp venom mastoparan. By applying pharmacological approach with specific inhibitors, we have investigated the involvement of eth

  8. Copper response regulator1-dependent and -independent responses of the Chlamydomonas reinhardtii transcriptome to dark anoxia.

    Science.gov (United States)

    Hemschemeier, Anja; Casero, David; Liu, Bensheng; Benning, Christoph; Pellegrini, Matteo; Happe, Thomas; Merchant, Sabeeha S

    2013-09-01

    Anaerobiosis is a stress condition for aerobic organisms and requires extensive acclimation responses. We used RNA-Seq for a whole-genome view of the acclimation of Chlamydomonas reinhardtii to anoxic conditions imposed simultaneously with transfer to the dark. Nearly 1.4 × 10(3) genes were affected by hypoxia. Comparing transcript profiles from early (hypoxic) with those from late (anoxic) time points indicated that cells activate oxidative energy generation pathways before employing fermentation. Probable substrates include amino acids and fatty acids (FAs). Lipid profiling of the C. reinhardtii cells revealed that they degraded FAs but also accumulated triacylglycerols (TAGs). In contrast with N-deprived cells, the TAGs in hypoxic cells were enriched in desaturated FAs, suggesting a distinct pathway for TAG accumulation. To distinguish transcriptional responses dependent on copper response regulator1 (CRR1), which is also involved in hypoxic gene regulation, we compared the transcriptomes of crr1 mutants and complemented strains. In crr1 mutants, ~40 genes were aberrantly regulated, reaffirming the importance of CRR1 for the hypoxic response, but indicating also the contribution of additional signaling strategies to account for the remaining differentially regulated transcripts. Based on transcript patterns and previous results, we conclude that nitric oxide-dependent signaling cascades operate in anoxic C. reinhardtii cells.

  9. Mutagenesis of Ser24 of cytochrome b559 α subunit affects PSⅡ activities in Chlamydomonas reinhardtii

    Institute of Scientific and Technical Information of China (English)

    MA JingJing; LI LiangBi; JING YuXiang; KUANG TingYun

    2007-01-01

    In order to study the functions of cytochrome b559 (Cyt b559) in photosystem two (PSⅡ) activity, mutant S24F of Chlamydomonas reinhardtii was constructed using site directed mutagenesis, in which Serine24 (Ser24) locating downstream of Histidine23 (His23) in c subunit of Cyt b559 was replaced by Phenylalanine (Phe). Physiological and biochemical analysis showed that mutant S24F could be grown photoautotrophically or photoheterotrophically. However, their growth rate was slower either on HSM or TAP medium than that of the control; Analysis of PSⅡ activity revealed that its oxygen evolution was about 71% of wild type (WT); The Photochemical efficiency of PSll (Fv/Fm) of S24F was reduced 0.23 compared with WT; S24F was more sensitive to strong light irradiance than the wild type; Furthermore,SDS-PAGE and Western-blotting analysis indicated that the expression levels of c subunit of Cyt b559,LHCⅡ and PsbO of S24F were a little less than those of the wild type. Overall, these data suggests that Ser24 plays a significant role in making Cyt b559 structure maintain PSⅡ complex activity of oxygen evolution although it is not directly bound to heme group.

  10. Integration of carbon assimilation modes with photosynthetic light capture in the green alga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Berger, Hanna; Blifernez-Klassen, Olga; Ballottari, Matteo; Bassi, Roberto; Wobbe, Lutz; Kruse, Olaf

    2014-10-01

    The unicellular green alga Chlamydomonas reinhardtii is capable of using organic and inorganic carbon sources simultaneously, which requires the adjustment of photosynthetic activity to the prevailing mode of carbon assimilation. We obtained novel insights into the regulation of light-harvesting at photosystem II (PSII) following altered carbon source availability. In C. reinhardtii, synthesis of PSII-associated light-harvesting proteins (LHCBMs) is controlled by the cytosolic RNA-binding protein NAB1, which represses translation of particular LHCBM isoform transcripts. This mechanism is fine-tuned via regulation of the nuclear NAB1 promoter, which is activated when linear photosynthetic electron flow is restricted by CO(2)-limitation in a photoheterotrophic context. In the wild-type, accumulation of NAB1 reduces the functional PSII antenna size, thus preventing a harmful overexcited state of PSII, as observed in a NAB1-less mutant. We further demonstrate that translation control as a newly identified long-term response to prolonged CO(2)-limitation replaces LHCII state transitions as a fast response to PSII over-excitation. Intriguingly, activation of the long-term response is perturbed in state transition mutant stt7, suggesting a regulatory link between the long- and short-term response. We depict a regulatory circuit operating on distinct timescales and in different cellular compartments to fine-tune light-harvesting in photoheterotrophic eukaryotes.

  11. Genetic Analysis of the Chlamydomonas Reinhardtii I-Crei Mobile Intron Homing System in Escherichia Coli

    Science.gov (United States)

    Seligman, L. M.; Stephens, K. M.; Savage, J. H.; Monnat-Jr., R. J.

    1997-01-01

    We have developed and used a genetic selection system in Escherichia coli to study functional requirements for homing site recognition and cleavage by a representative eukaryotic mobile intron endonuclease. The homing endonuclease, I-CreI, was originally isolated from the chloroplast of the unicellular green alga Chlamydomonas reinhardtii. I-CreI homing site mutants contained base pair substitutions or single base deletions that altered the rate of homing site cleavage and/or product release. I-CreI endonuclease mutants fell into six phenotypic classes that differed in in vivo activity, toxicity or genetic dominance. Inactivating mutations clustered in the N-terminal 60% of the I-CreI amino acid sequence, and two frameshift mutations were isolated that resulted in premature translation termination though retained partial activity. These mutations indicate that the N-terminal two-thirds of the I-CreI endonuclease is sufficient for homing site recognition and cleavage. Substitution mutations altered in four potential active site residues were examined: D20N, Q47H or R70A substitutions inactivated endonuclease activity, whereas S22A did not. The genetic approach we have taken complements phylogenetic and structural studies of mobile intron endonucleases and has provided new information on the mechanistic basis of I-CreI homing site recognition and cleavage. PMID:9409828

  12. LC2, the Chlamydomonas Homologue of the t Complex-encoded Protein Tctex2, Is Essential for Outer Dynein Arm Assembly

    Science.gov (United States)

    Pazour, Gregory J.; Koutoulis, Anthony; Benashski, Sharon E.; Dickert, Bethany L.; Sheng, Hong; Patel-King, Ramila S.; King, Stephen M.; Witman, George B.

    1999-01-01

    Tctex2 is thought to be one of the distorter genes of the mouse t haplotype. This complex greatly biases the segregation of the chromosome that carries it such that in heterozygous +/t males, the t haplotype is transmitted to >95% of the offspring, a phenomenon known as transmission ratio distortion. The LC2 outer dynein arm light chain of Chlamydomonas reinhardtii is a homologue of the mouse protein Tctex2. We have identified Chlamydomonas insertional mutants with deletions in the gene encoding LC2 and demonstrate that the LC2 gene is the same as the ODA12 gene, the product of which had not been identified previously. Complete deletion of the LC2/ODA12 gene causes loss of all outer arms and a slow jerky swimming phenotype. Transformation of the deletion mutant with the cloned LC2/ODA12 gene restores the outer arms and rescues the motility phenotype. Therefore, LC2 is required for outer arm assembly. The fact that LC2 is an essential subunit of flagellar outer dynein arms allows us to propose a detailed mechanism whereby transmission ratio distortion is explained by the differential binding of mutant (t haplotype encoded) and wild-type dyneins to the axonemal microtubules of t-bearing or wild-type sperm, with resulting differences in their motility. PMID:10512883

  13. An optimized, chemically regulated gene expression system for Chlamydomonas.

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    Paola Ferrante

    Full Text Available BACKGROUND: Chlamydomonas reinhardtii is a model system for algal and cell biology and is used for biotechnological applications, such as molecular farming or biological hydrogen production. The Chlamydomonas metal-responsive CYC6 promoter is repressed by copper and induced by nickel ions. However, induction by nickel is weak in some strains, poorly reversible by chelating agents like EDTA, and causes, at high concentrations, toxicity side effects on Chlamydomonas growth. Removal of these bottlenecks will encourage the wide use of this promoter as a chemically regulated gene expression system. METHODOLOGY: Using a codon-optimized Renilla luciferase as a reporter gene, we explored several strategies to improve the strength and reversibility of CYC6 promoter induction. Use of the first intron of the RBCS2 gene or of a modified TAP medium increases the strength of CYC6 induction up to 20-fold. In the modified medium, induction is also obtained after addition of specific copper chelators, like TETA. At low concentrations (up to 10 microM TETA is a more efficient inducer than Ni, which becomes a very efficient inducer at higher concentrations (50 microM. Neither TETA nor Ni show toxicity effects at the concentrations used. Unlike induction by Ni, induction by TETA is completely reversible by micromolar copper concentrations, thus resulting in a transient "wave" in luciferase activity, which can be repeated in subsequent growth cycles. CONCLUSIONS: We have worked out a chemically regulated gene expression system that can be finely tuned to produce temporally controlled "waves" in gene expression. The use of cassettes containing the CYC6 promoter, and of modified growth media, is a reliable and economically sustainable system for the temporally controlled expression of foreign genes in Chlamydomonas.

  14. Regulation by glutathionylation of isocitrate lyase from Chlamydomonas reinhardtii.

    Science.gov (United States)

    Bedhomme, Mariette; Zaffagnini, Mirko; Marchand, Christophe H; Gao, Xing-Huang; Moslonka-Lefebvre, Mathieu; Michelet, Laure; Decottignies, Paulette; Lemaire, Stéphane D

    2009-12-25

    Post-translational modification of protein cysteine residues is emerging as an important regulatory and signaling mechanism. We have identified numerous putative targets of redox regulation in the unicellular green alga Chlamydomonas reinhardtii. One enzyme, isocitrate lyase (ICL), was identified both as a putative thioredoxin target and as an S-thiolated protein in vivo. ICL is a key enzyme of the glyoxylate cycle that allows growth on acetate as a sole source of carbon. The aim of the present study was to clarify the molecular mechanism of the redox regulation of Chlamydomonas ICL using a combination of biochemical and biophysical methods. The results clearly show that purified C. reinhardtii ICL can be inactivated by glutathionylation and reactivated by glutaredoxin, whereas thioredoxin does not appear to regulate ICL activity, and no inter- or intramolecular disulfide bond could be formed under any of the conditions tested. Glutathionylation of the protein was investigated by mass spectrometry analysis, Western blotting, and site-directed mutagenesis. The enzyme was found to be protected from irreversible oxidative inactivation by glutathionylation of its catalytic Cys(178), whereas a second residue, Cys(247), becomes artifactually glutathionylated after prolonged incubation with GSSG. The possible functional significance of this post-translational modification of ICL in Chlamydomonas and other organisms is discussed.

  15. Two types of chloroplast gene promoters in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Klein, U; De Camp, J D; Bogorad, L

    1992-04-15

    Structures of the promoters of Chlamydomonas reinhardtii plastid atpB and 16S rRNA-encoding genes were analyzed in vivo. Chimeric constructs, containing the Chlamydomonas chloroplast atpB or 16S rRNA-encoding gene promoter coupled to the Escherichia coli uidA (beta-glucuronidase, GUS) reporter gene and bordered by C. reinhardtii chloroplast sequences, were stably introduced into the chloroplast of Chlamydomonas by microprojectile bombardment. Activity of the promoters in the chloroplast of GUS gene-positive transformants was assayed by measuring the abundance of GUS transcripts and determining the relative rates of GUS transcription in vivo. Deletion analyses of the 16S rRNA gene and atpB promoter fragments showed that the two promoters differ structurally. The 16S rRNA gene promoter resembles the bacterial sigma 70 type with typical -10 and -35 elements. The atpB promoter, on the other hand, lacks a conserved motif in the -35 region but contains, in the -10 region, a characteristic octameric palindrome (TATAATAT) that is conserved in the promoter sequences of some other C. reinhardtii chloroplast genes. For maximum activity, the atpB promoter requires sequences of approximately 22 base pairs upstream and approximately 60 base pairs downstream of the transcription start site.

  16. A C2H2 zinc finger protein FEMU2 is required for fox1 expression in Chlamydomonas reinhardtii.

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    Xiaodong Deng

    Full Text Available Chlamydomonas reinhardtii fox1 gene encodes a ferroxidase that is involved in cellular Fe uptake and highly induced during Fe deficient conditions. In an effort to identify fox1 promoter regulatory elements, an insertional library was generated in a transgenic Chlamydomonas strain (2A38 harboring an arylsulfatase (ARS reporter gene driven by the fox1 promoter. Mutants with a defective response to low iron conditions were selected for further study. Among these, a strain containing a disrupted femu2 gene was identified. Activation of the fox1 promoter by the femu2 gene product was confirmed by silencing the femu2 gene using RNA interference. In three femu2 RNAi transgenic lines (IR3, IR6, and IR7, ARS reporter gene activities declined by 84.3%, 86.4%, and 88.8%, respectively under Fe deficient conditions. Furthermore, RT-PCR analysis of both the femu2 mutant and the RNAi transgenic lines showed significantly decreased transcript abundance of the endogenous fox1 gene under Fe deficient conditions. Amino acid sequence analysis of the femu2 gene product identified three potential C2H2 zinc finger (ZF motifs and a nuclear localization study suggests that FEMU2 is localized to the nucleus. In addition, a potential FEMU2 binding site ((G/TTTGG(G/T(G/TT was identified using PCR-mediated random binding site selection. Taken together, this evidence suggests that FEMU2 is involved in up-regulation of the fox1 gene in Fe deficient cells.

  17. Functional analysis of Photosystem I light-harvesting complexes (Lhca) gene products of Chlamydomonas reinhardtii

    NARCIS (Netherlands)

    Mozzo, Milena; Mantelli, Manuela; Passarini, Francesca; Caffarri, Stefano; Croce, Roberta; Bassi, Roberto

    2010-01-01

    The outer antenna system of Chlamydomonas reinhardtii Photosystem I is composed of nine gene products, but due to difficulty in purification their individual properties are not known. In this work, the functional properties of the nine Lhca antennas of Chlamydomonas, have been investigated upon expr

  18. Omics in Chlamydomonas for Biofuel Production.

    Science.gov (United States)

    Aucoin, Hanna R; Gardner, Joseph; Boyle, Nanette R

    2016-01-01

    In response to demands for sustainable domestic fuel sources, research into biofuels has become increasingly important. Many challenges face biofuels in their effort to replace petroleum fuels, but rational strain engineering of algae and photosynthetic organisms offers a great deal of promise. For decades, mutations and stress responses in photosynthetic microbiota were seen to result in production of exciting high-energy fuel molecules, giving hope but minor capability for design. However, '-omics' techniques for visualizing entire cell processing has clarified biosynthesis and regulatory networks. Investigation into the promising production behaviors of the model organism C. reinhardtii and its mutants with these powerful techniques has improved predictability and understanding of the diverse, complex interactions within photosynthetic organisms. This new equipment has created an exciting new frontier for high-throughput, predictable engineering of photosynthetically produced carbon-neutral biofuels.

  19. Neurobehavioral Mutants Identified in an ENU Mutagenesis Project

    Energy Technology Data Exchange (ETDEWEB)

    Cook, Melloni N. [University of Memphis; Dunning, Jonathan P [University of Memphis; Wiley, Ronald G [Vanderbilt University and Veterans Administration, Nashville, TN; Chesler, Elissa J [ORNL; Johnson, Dabney K [ORNL; Goldowitz, Daniel [University of Tennessee Health Science Center, Memphis

    2007-01-01

    We report on a behavioral screening test battery that successfully identified several neurobehavioral mutants among a large-scale ENU-mutagenized mouse population. Large numbers of ENU mutagenized mice were screened for abnormalities in central nervous system function based on abnormal performance in a series of behavior tasks. We developed and employed a high-throughput screen of behavioral tasks to detect behavioral outliers. Twelve mutant pedigrees, representing a broad range of behavioral phenotypes, have been identified. Specifically, we have identified two open field mutants (one displaying hyper-locomotion, the other hypo-locomotion), four tail suspension mutants (all displaying increased immobility), one nociception mutant (displaying abnormal responsiveness to thermal pain), two prepulse inhibition mutants (displaying poor inhibition of the startle response), one anxiety-related mutant (displaying decreased anxiety in the light/dark test), and one learning and memory mutant (displaying reduced response to the conditioned stimulus) These findings highlight the utility of a set of behavioral tasks used in a high throughput screen to identify neurobehavioral mutants. Further analysis (i.e., behavioral and genetic mapping studies) of mutants is in progress with the ultimate goal of identification of novel genes and mouse models relevant to human disorders as well as the identification of novel therapeutic targets.

  20. Phosphoprotein SAK1 is a regulator of acclimation to singlet oxygen in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Wakao, Setsuko; Chin, Brian L; Ledford, Heidi K; Dent, Rachel M; Casero, David; Pellegrini, Matteo; Merchant, Sabeeha S; Niyogi, Krishna K

    2014-05-23

    Singlet oxygen is a highly toxic and inevitable byproduct of oxygenic photosynthesis. The unicellular green alga Chlamydomonas reinhardtii is capable of acclimating specifically to singlet oxygen stress, but the retrograde signaling pathway from the chloroplast to the nucleus mediating this response is unknown. Here we describe a mutant, singlet oxygen acclimation knocked-out 1 (sak1), that lacks the acclimation response to singlet oxygen. Analysis of genome-wide changes in RNA abundance during acclimation to singlet oxygen revealed that SAK1 is a key regulator of the gene expression response during acclimation. The SAK1 gene encodes an uncharacterized protein with a domain conserved among chlorophytes and present in some bZIP transcription factors. The SAK1 protein is located in the cytosol, and it is induced and phosphorylated upon exposure to singlet oxygen, suggesting that it is a critical intermediate component of the retrograde signal transduction pathway leading to singlet oxygen acclimation.DOI: http://dx.doi.org/10.7554/eLife.02286.001.

  1. Phosphoprotein SAK1 is a regulator of acclimation to singlet oxygen in Chlamydomonas reinhardtii

    Science.gov (United States)

    Wakao, Setsuko; Chin, Brian L; Ledford, Heidi K; Dent, Rachel M; Casero, David; Pellegrini, Matteo; Merchant, Sabeeha S; Niyogi, Krishna K

    2014-01-01

    Singlet oxygen is a highly toxic and inevitable byproduct of oxygenic photosynthesis. The unicellular green alga Chlamydomonas reinhardtii is capable of acclimating specifically to singlet oxygen stress, but the retrograde signaling pathway from the chloroplast to the nucleus mediating this response is unknown. Here we describe a mutant, singlet oxygen acclimation knocked-out 1 (sak1), that lacks the acclimation response to singlet oxygen. Analysis of genome-wide changes in RNA abundance during acclimation to singlet oxygen revealed that SAK1 is a key regulator of the gene expression response during acclimation. The SAK1 gene encodes an uncharacterized protein with a domain conserved among chlorophytes and present in some bZIP transcription factors. The SAK1 protein is located in the cytosol, and it is induced and phosphorylated upon exposure to singlet oxygen, suggesting that it is a critical intermediate component of the retrograde signal transduction pathway leading to singlet oxygen acclimation. DOI: http://dx.doi.org/10.7554/eLife.02286.001 PMID:24859755

  2. An organelle K+ channel is required for osmoregulation in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Xu, Feifei; Wu, Xiaoan; Jiang, Lin-Hua; Zhao, Hucheng; Pan, Junmin

    2016-08-01

    Fresh water protozoa and algae face hypotonic challenges in their living environment. Many of them employ a contractile vacuole system to uptake excessive water from the cytoplasm and expel it to the environment to achieve cellular homeostasis. K(+), a major osmolyte in contractile vacuole, is predicted to create higher osmolarity for water influx. Molecular mechanisms for K(+) permeation through the plasma membrane have been well studied. However, how K(+) permeates organelles such as the contractile vacuole is not clear. Here, we show that the six-transmembrane K(+) channel KCN11 in Chlamydomonas is exclusively localized to contractile vacuole. Ectopic expression of KCN11 in HEK293T cells results in voltage-gated K(+) channel activity. Disruption of the gene or mutation of key residues for K(+) permeability of the channel leads to dysfunction of cell osmoregulation in very hypotonic conditions. The contractile cycle is inhibited in the mutant cells with a slower rate of contractile vacuole swelling, leading to cell death. These data demonstrate a new role for six-transmembrane K(+) channels in contractile vacuole functioning and provide further insights into osmoregulation mediated by the contractile vacuole.

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

    Science.gov (United States)

    Nickelsen, J.; Kück, U.

    Chloroplasts are typical organelles of photoautotrophic eukaryotic cells which drive a variety of functions, including photosynthesis. For many years the unicellular green alga Chlamydomonas reinhardtii has served as an experimental organism for studying photosynthetic processes. The recent development of molecular tools for this organism together with efficient methods of genetic analysis and the availability of many photosynthesis mutants has now made this alga a powerful model system for the analysis of chloroplast biogenesis. For example, techniques have been developed to transfer recombinant DNA into both the nuclear and the chloroplast genome. This allows both complementation tests and analyses of gene functions in vivo. Moreover, site-specific DNA recombinations in the chloroplast allow targeted gene disruption experiments which enable a "reverse genetics" to be performed. The potential of the algal system for the study of chloroplast biogenesis is illustrated in this review by the description of regulatory systems of gene expression involved in organelle biogenesis. One example concerns the regulation of trans-splicing of chloroplast mRNAs, a process which is controlled by both multiple nuclear- and chloroplast-encoded factors. The second example involves the stabilization of chloroplast mRNAs. The available data lead us predict distinct RNA elements, which interact with trans-acting factors to protect the RNA against nucleolytic attacks.

  4. Phosphoribulokinase from Chlamydomonas reinhardtii: a Benson-Calvin cycle enzyme enslaved to its cysteine residues.

    Science.gov (United States)

    Thieulin-Pardo, Gabriel; Remy, Thérèse; Lignon, Sabrina; Lebrun, Régine; Gontero, Brigitte

    2015-04-01

    Phosphoribulokinase (PRK) in the green alga Chlamydomonas reinhardtii is a finely regulated and well-studied enzyme of the Benson-Calvin cycle. PRK can form a complex with glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and the small chloroplast protein CP12. This study aimed to determine the molecular determinants on PRK involved in the complex and the mechanism of action of a recently described novel regulation of PRK that involves glutathionylation. A combination of mass spectrometry, mutagenesis and activity analyses showed that Cys16, besides its role as the binding site of ATP, was also the site for S-glutathionylation. Previous kinetic analysis of the C55S mutant showed that in the oxidized inactive form of PRK, this residue formed a disulfide bridge with the Cys16 residue. This is the only bridge reported for PRK in the literature. Our data show for the first time that a disulfide bridge between Cys243 and Cys249 on PRK is required to form the PRK-GAPDH-CP12 complex. These results uncover a new mechanism for the PRK-GAPDH-CP12 formation involving a thiol disulfide exchange reaction with CP12 and identify Cys16 of PRK as a target of glutathionylation acting against oxidative stress. Although Cys16 is the key residue involved in binding ATP and acting as a defense against oxidative damage, the formation of the algal ternary complex requires the formation of another disulfide bridge on PRK involving Cys243 and Cys249.

  5. Thioredoxin Ch1 of Chlamydomonas reinhardtii displays an unusual resistance toward one-electron oxidation.

    Science.gov (United States)

    Sicard-Roselli, Cécile; Lemaire, Stéphane; Jacquot, Jean-Pierre; Favaudon, Vincent; Marchand, Christophe; Houée-Levin, Chantal

    2004-09-01

    To test thioredoxin resistance to oxidizing free radicals, we have studied the one-electron oxidation of wild-type thioredoxin and of two forms with the point mutations D30A and W35A, using azide radicals generated by gamma-ray or pulse radiolysis. The oxidation patterns of wild-type thioredoxin and D30A are similar. In these forms, Trp35 is the primary target and is 'repaired' by one-electron reduction; first by intramolecular electron transfer from tyrosine, and then from other residues. Conversely, during oxidation of W35A, Trp13 is poorly reactive. For all proteins, activity is conserved showing an unusual resistance toward oxidation.

  6. A new F131V mutation in Chlamydomonas phytoene desaturase locates a cluster of norflurazon resistance mutations near the FAD-binding site in 3D protein models.

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    Julio V Suarez

    Full Text Available The green alga Chlamydomonas reinhardtii provides a tractable genetic model to study herbicide mode of action using forward genetics. The herbicide norflurazon inhibits phytoene desaturase, which is required for carotenoid synthesis. Locating amino acid substitutions in mutant phytoene desaturases conferring norflurazon resistance provides a genetic approach to map the herbicide binding site. We isolated a UV-induced mutant able to grow in very high concentrations of norflurazon (150 µM. The phytoene desaturase gene in the mutant strain contained the first resistance mutation to be localised to the dinucleotide-binding Rossmann-likedomain. A highly conserved phenylalanine amino acid at position 131 of the 564 amino acid precursor protein was changed to a valine in the mutant protein. F131, and two other amino acids whose substitution confers norflurazon resistance in homologous phytoene desaturase proteins, map to distant regions in the primary sequence of the C. reinhardtii protein (V472, L505 but in tertiary models these residues cluster together to a region close to the predicted FAD binding site. The mutant gene allowed direct 5 µM norflurazon based selection of transformants, which were tolerant to other bleaching herbicides including fluridone, flurtamone, and diflufenican but were more sensitive to beflubutamid than wild type cells. Norflurazon resistance and beflubutamid sensitivity allow either positive or negative selection against transformants expressing the mutant phytoene desaturase gene.

  7. A novel screening protocol for the isolation of hydrogen producing Chlamydomonas reinhardtii strains

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    Happe Thomas

    2008-10-01

    Full Text Available Abstract Background Sealed Chlamydomonas reinhardtii cultures evolve significant amounts of hydrogen gas under conditions of sulfur depletion. However, the eukaryotic green alga goes through drastic metabolic changes during this nutritional stress resulting in cell growth inhibition and eventually cell death. This study aimed at isolating C. reinhardtii transformants which produce hydrogen under normal growth conditions to allow a continuous hydrogen metabolism without the stressful impact of nutrient deprivation. Results To achieve a steady photobiological hydrogen production, a screening protocol was designed to identify C. reinhardtii DNA insertional mutagenesis transformants with an attenuated photosynthesis to respiration capacity ratio (P/R ratio. The screening protocol entails a new and fast method for mutant strain selection altered in their oxygen production/consumption balance. Out of 9000 transformants, four strains with P/R ratios varying from virtually zero to three were isolated. Strain apr1 was found to have a slightly higher respiration rate and a significantly lower photosynthesis rate than the wild type. Sealed cultures of apr1 became anaerobic in normal growth medium (TAP under moderate light conditions and induced [FeFe]-hydrogenase activity, yet without significant hydrogen gas evolution. However, Calvin-Benson cycle inactivation of anaerobically adapted apr1 cells in the light led to a 2-3-fold higher in vivo hydrogen production than previously reported for the sulfur-deprived C. reinhardtii wild type. Conclusion Attenuated P/R capacity ratio in microalgal mutants constitutes a platform for achieving steady state photobiological hydrogen production. Using this platform, algal hydrogen metabolism can be analyzed without applying nutritional stress. Furthermore, these strains promise to be useful for biotechnological hydrogen generation, since high in vivo hydrogen production rates are achievable under normal growth

  8. RNA-Seq Analysis of Sulfur-Deprived Chlamydomonas Cells Reveals Aspects of Acclimation Critical for Cell Survival[W

    Science.gov (United States)

    González-Ballester, David; Casero, David; Cokus, Shawn; Pellegrini, Matteo; Merchant, Sabeeha S.; Grossman, Arthur R.

    2010-01-01

    The Chlamydomonas reinhardtii transcriptome was characterized from nutrient-replete and sulfur-depleted wild-type and snrk2.1 mutant cells. This mutant is null for the regulatory Ser-Thr kinase SNRK2.1, which is required for acclimation of the alga to sulfur deprivation. The transcriptome analyses used microarray hybridization and RNA-seq technology. Quantitative RT-PCR evaluation of the results obtained by these techniques showed that RNA-seq reports a larger dynamic range of expression levels than do microarray hybridizations. Transcripts responsive to sulfur deprivation included those encoding proteins involved in sulfur acquisition and assimilation, synthesis of sulfur-containing metabolites, Cys degradation, and sulfur recycling. Furthermore, we noted potential modifications of cellular structures during sulfur deprivation, including the cell wall and complexes associated with the photosynthetic apparatus. Moreover, the data suggest that sulfur-deprived cells accumulate proteins with fewer sulfur-containing amino acids. Most of the sulfur deprivation responses are controlled by the SNRK2.1 protein kinase. The snrk2.1 mutant exhibits a set of unique responses during both sulfur-replete and sulfur-depleted conditions that are not observed in wild-type cells; the inability of this mutant to acclimate to S deprivation probably leads to elevated levels of singlet oxygen and severe oxidative stress, which ultimately causes cell death. The transcriptome results for wild-type and mutant cells strongly suggest the occurrence of massive changes in cellular physiology and metabolism as cells become depleted for sulfur and reveal aspects of acclimation that are likely critical for cell survival. PMID:20587772

  9. Excitation energy transfer in Chlamydomonas reinhardtii deficient in the PSI core or the PSII core under conditions mimicking state transitions.

    Science.gov (United States)

    Wlodarczyk, Lucyna M; Dinc, Emine; Croce, Roberta; Dekker, Jan P

    2016-06-01

    The efficient use of excitation energy in photosynthetic membranes is achieved by a dense network of pigment-protein complexes. These complexes fulfill specific functions and interact dynamically with each other in response to rapidly changing environmental conditions. Here, we studied how in the intact cells of Chlamydomonas reinhardtii (C.r.) the lack of the photosystem I (PSI) core or the photosystem II (PSII) core affects these interactions. To that end the mutants F15 and M18 (both PSI-deficient) and FUD7 (PSII-deficient) were incubated under conditions known to promote state transitions in wild-type. The intact cells were then instantly frozen to 77K and the full-spectrum time-resolved fluorescence emission of the cells was measured by means of streak camera. In the PSI-deficient mutants excitation energy transfer (EET) towards light-harvesting complexes of PSI (Lhca) occurs in less than 0.5 ns, and fluorescence from Lhca decays in 3.1 ns. Decreased trapping by PSII and increased fluorescence of Lhca upon state 1 (S1)→state 2 (S2) transition appears in the F15 and less in the M18 mutant. In the PSII-deficient mutant FUD7, quenched (0.5 ns) and unquenched (2 ns) light-harvesting complexes of PSII (LHCII) are present in both states, with the quenched form more abundant in S2 than in S1. Moreover, EET of 0.4 ns from the remaining LHCII to PSI increases upon S1→S2 transition. We relate the excitation energy kinetics observed in F15, M18 and FUD7 to the remodeling of the photosynthetic apparatus in these mutants under S1 and S2 conditions.

  10. Genome-wide analysis of alternative splicing in Chlamydomonas reinhardtii

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    Thomas Julie

    2010-02-01

    Full Text Available Abstract Background Genome-wide computational analysis of alternative splicing (AS in several flowering plants has revealed that pre-mRNAs from about 30% of genes undergo AS. Chlamydomonas, a simple unicellular green alga, is part of the lineage that includes land plants. However, it diverged from land plants about one billion years ago. Hence, it serves as a good model system to study alternative splicing in early photosynthetic eukaryotes, to obtain insights into the evolution of this process in plants, and to compare splicing in simple unicellular photosynthetic and non-photosynthetic eukaryotes. We performed a global analysis of alternative splicing in Chlamydomonas reinhardtii using its recently completed genome sequence and all available ESTs and cDNAs. Results Our analysis of AS using BLAT and a modified version of the Sircah tool revealed AS of 498 transcriptional units with 611 events, representing about 3% of the total number of genes. As in land plants, intron retention is the most prevalent form of AS. Retained introns and skipped exons tend to be shorter than their counterparts in constitutively spliced genes. The splice site signals in all types of AS events are weaker than those in constitutively spliced genes. Furthermore, in alternatively spliced genes, the prevalent splice form has a stronger splice site signal than the non-prevalent form. Analysis of constitutively spliced introns revealed an over-abundance of motifs with simple repetitive elements in comparison to introns involved in intron retention. In almost all cases, AS results in a truncated ORF, leading to a coding sequence that is around 50% shorter than the prevalent splice form. Using RT-PCR we verified AS of two genes and show that they produce more isoforms than indicated by EST data. All cDNA/EST alignments and splice graphs are provided in a website at http://combi.cs.colostate.edu/as/chlamy. Conclusions The extent of AS in Chlamydomonas that we observed is much

  11. Effects of chromium on photosynthetic and photoreceptive apparatus of the alga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Rodríguez, M Cecilia; Barsanti, Laura; Passarelli, Vincenzo; Evangelista, Valter; Conforti, Visitacion; Gualtieri, Paolo

    2007-10-01

    Chromium is a highly toxic non-essential metal for microorganisms and plants. Due to its widespread industrial use, chromium (Cr) has become a serious pollutant in diverse environmental settings. The presence of Cr leads to the selection of algal populations able to tolerate high levels of Cr compounds. The diverse Cr-resistance mechanisms displayed by microorganisms include biosorption, diminished accumulation, precipitation, reduction of Cr(6+) to Cr(3+), and chromate efflux. In this paper we describe the effects of Cr(6+) (the more toxic species) on the photosynthetic and photoreceptive apparatus of the fresh water unicellular alga Chlamydomonas reinhardtii. We measured the effect of the heavy metal by means of in vivo absorption microspectroscopy of both the thylakoid compartments and the eyespot. The decomposition of the overall absorption spectra in pigment constituents indicates that Cr(6+) induced a complete pheophinitization of the chrorophylls and a modification of the carotenoids present in the eyespot only when its concentration is equal or greater than 10 microM. Due to this low tolerance level, C. reinhardtii could be used as indicator of Cr pollution, but it is not feasible for bioremediation purposes.

  12. The Chlamydomonas reinhardtii alternative oxidase 1 is regulated by heat stress.

    Science.gov (United States)

    Zalutskaya, Zhanneta; Lapina, Tatiana; Ermilova, Elena

    2015-12-01

    The alternative oxidase (AOX) is a non-energy conserving terminal oxidase that has emerged as an important mitochondrial component of the cell stress responses. Although the most studied abiotic condition in relation to Chlamydomonas reinhardtii is high temperature, changes in AOX capacity of the alga were studied only under oxidative stress and cold. To examine whether elevated temperatures affected AOX1 expression, we applied quantitative real-time PCR and pharmaceutical approaches. In this work, we demonstrated a sharp increase in AOX1 transcript and protein abundance under heat stress. Furthermore, C. reinhardtii cells displayed a large increase in alternative respiration in response to high temperature. Feeding with the protein kinase inhibitor staurosporine strongly retarded the AOX1 transcription. Finally, the addition of the calcium chelator EGTA prevented heat-induced AOX1 expression. Together, our results imply that heat-inducible Ca(2+) influx and protein kinase(s) may mediate AOX1 expression at elevated temperatures. Characterization of heat-induced AOX1 regulation in the green alga C. reinhardtii provides a framework for a more complete understanding of the function of this conserved protein.

  13. Coordinated Regulation of Gene Expression for Carotenoid Metabolism in Chlamydomonas reinhardtii

    Institute of Scientific and Technical Information of China (English)

    Tian-Hu Sun; Cheng-Qian Liu; Yuan-Yuan Hui; Wen-Kai Wu; Zhi-Gang Zhou; Shan Lu

    2010-01-01

    Carotenoids are important plant pigments for both light harvesting and photooxidation protection.Using the model system of the unicellular green alga Chlamydomonas reinhardtii,we characterized the regulation of gene expression for carotenoid metabolism by quantifying changes in the transcript abundance of dxs,dxr and ipi in the plastidic methylerythritol phosphate pathway and of ggps,psy,pds,lcyb and bchy,directly involved in carotenoid metabolism,under different photoperiod,light and metabolite treatments.The expression of these genes fluctuated with light/dark shifting.Light treatment also promoted the accumulation of transcripts of all these genes.Of the genes studied,dxs,ggps and lcyb displayed the typical circadian pattern by retaining a rhythmic fluctuation of transcript abundance under both constant light and constant dark entrainments.The expression of these genes could also be regulated by metabolic intermediates.For example,ggps was significantly suppressed by a geranylgeranyl pyrophosphate supplement and ipi was upregulated by isopentenyl pyrophosphate.Furthermore,CrOr,a C.reinhardtii homolog of the recently characterized Or gene that accounts for carotenoid accumulation,also showed co-expression with carotenoid biosynthetic genes such as pds and lcyb.Our data suggest a coordinated regulation on carotenoid metabolism in C.reinhardtii at the transcriptional level.

  14. Development of the light-harvesting chlorophyll antenna in the green alga Chlamydomonas reinhardtii is regulated by the novel Tla1 gene.

    Science.gov (United States)

    Tetali, Sarada D; Mitra, Mautusi; Melis, Anastasios

    2007-03-01

    The Chlamydomonas reinhardtii tla1 (truncated light-harvesting chlorophyll antenna size) mutant was generated upon DNA insertional mutagenesis and shown to specifically possess a smaller than wild type (WT) chlorophyll antenna size in both photosystems. Molecular and genetic analysis revealed that the exogenous plasmid DNA was inserted at the end of the 5' UTR and just prior to the ATG start codon of a hitherto unknown nuclear gene (termed Tla1), which encodes a protein of 213 amino acids. The Tla1 gene in the mutant is transcribed with a new 5' UTR sequence, derived from the 3' end of the transforming plasmid. This replacement of the native 5' UTR and promoter regions resulted in enhanced transcription of the tla1 gene in the mutant but inhibition in the translation of the respective tla1 mRNA. Transformation of the tla1 mutant with WT Tla1 genomic DNA successfully rescued the mutant. These results are evidence that polymorphism in the 5' UTR of the Tla1 transcripts resulted in the tla1 phenotype and that expression of the Tla1 gene is a prerequisite for the development/assembly of the Chl antenna in C. reinhardtii. A blast search with the Tla1 deduced amino acid sequence

  15. Analysis of the nucleus-encoded and chloroplast-targeted rieske protein by classic and site-directed mutagenesis of Chlamydomonas.

    Science.gov (United States)

    de Vitry, C; Finazzi, G; Baymann, F; Kallas, T

    1999-10-01

    Three mutants of the alga Chlamydomonas reinhardtii affected in the nuclear PETC gene encoding the Rieske iron-sulfur protein 2Fe-2S subunit of the chloroplast cytochrome b(6)f complex have been characterized. One has a stable deletion that eliminates the protein; two others carry substitutions Y87D and W163R that result in low accumulation of the protein. Attenuated expression of the stromal protease ClpP increases accumulation and assembly into b(6)f complexes of the Y87D and W163R mutant Rieske proteins in quantities sufficient for analysis. Electron-transfer kinetics of these complexes were 10- to 20-fold slower than those for the wild type. The deletion mutant was used as a recipient for site-directed mutant petC alleles. Six glycine residues were replaced by alanine residues (6G6A) in the flexible hinge that is critical for domain movement; substitutions were created near the 2Fe-2S cluster (S128 and W163); and seven C-terminal residues were deleted (G171och). Although the 6G6A and G171och mutations affect highly conserved segments in the chloroplast Rieske protein, photosynthesis in the mutants was similar to that of the wild type. These results establish the basis for mutational analysis of the nuclear-encoded and chloroplast-targeted Rieske protein of photosynthesis.

  16. A role for the membrane in regulating Chlamydomonas flagellar length.

    Directory of Open Access Journals (Sweden)

    William Dentler

    Full Text Available Flagellar assembly requires coordination between the assembly of axonemal proteins and the assembly of the flagellar membrane and membrane proteins. Fully grown steady-state Chlamydomonas flagella release flagellar vesicles from their tips and failure to resupply membrane should affect flagellar length. To study vesicle release, plasma and flagellar membrane surface proteins were vectorially pulse-labeled and flagella and vesicles were analyzed for biotinylated proteins. Based on the quantity of biotinylated proteins in purified vesicles, steady-state flagella appeared to shed a minimum of 16% of their surface membrane per hour, equivalent to a complete flagellar membrane being released every 6 hrs or less. Brefeldin-A destroyed Chlamydomonas Golgi, inhibited the secretory pathway, inhibited flagellar regeneration, and induced full-length flagella to disassemble within 6 hrs, consistent with flagellar disassembly being induced by a failure to resupply membrane. In contrast to membrane lipids, a pool of biotinylatable membrane proteins was identified that was sufficient to resupply flagella as they released vesicles for 6 hrs in the absence of protein synthesis and to support one and nearly two regenerations of flagella following amputation. These studies reveal the importance of the secretory pathway to assemble and maintain full-length flagella.

  17. The LC7 Light Chains of Chlamydomonas Flagellar Dyneins Interact with Components Required for Both Motor Assembly and Regulation

    Science.gov (United States)

    DiBella, Linda M.; Sakato, Miho; Patel-King, Ramila S.; Pazour, Gregory J.; King, Stephen M.

    2004-01-01

    Members of the LC7/Roadblock family of light chains (LCs) have been found in both cytoplasmic and axonemal dyneins. LC7a was originally identified within Chlamydomonas outer arm dynein and associates with this motor's cargo-binding region. We describe here a novel member of this protein family, termed LC7b that is also present in the Chlamydomonas flagellum. Levels of LC7b are reduced ∼20% in axonemes isolated from strains lacking inner arm I1 and are ∼80% lower in the absence of the outer arms. When both dyneins are missing, LC7b levels are diminished to <10%. In oda9 axonemal extracts that completely lack outer arms, LC7b copurifies with inner arm I1, whereas in ida1 extracts that are devoid of I1 inner arms it associates with outer arm dynein. We also have observed that some LC7a is present in both isolated axonemes and purified 18S dynein from oda1, suggesting that it is also a component of both the outer arm and inner arm I1. Intriguingly, in axonemal extracts from the LC7a null mutant, oda15, which assembles ∼30% of its outer arms, LC7b fails to copurify with either dynein, suggesting that it interacts with LC7a. Furthermore, both the outer arm γ heavy chain and DC2 from the outer arm docking complex completely dissociate after salt extraction from oda15 axonemes. EDC cross-linking of purified dynein revealed that LC7b interacts with LC3, an outer dynein arm thioredoxin; DC2, an outer arm docking complex component; and also with the phosphoprotein IC138 from inner arm I1. These data suggest that LC7a stabilizes both the outer arms and inner arm I1 and that both LC7a and LC7b are involved in multiple intradynein interactions within both dyneins. PMID:15304520

  18. Mutations of photosystem II D1 protein that empower efficient phenotypes of Chlamydomonas reinhardtii under extreme environment in space.

    Science.gov (United States)

    Giardi, Maria Teresa; Rea, Giuseppina; Lambreva, Maya D; Antonacci, Amina; Pastorelli, Sandro; Bertalan, Ivo; Johanningmeier, Udo; Mattoo, Autar K

    2013-01-01

    Space missions have enabled testing how microorganisms, animals and plants respond to extra-terrestrial, complex and hazardous environment in space. Photosynthetic organisms are thought to be relatively more prone to microgravity, weak magnetic field and cosmic radiation because oxygenic photosynthesis is intimately associated with capture and conversion of light energy into chemical energy, a process that has adapted to relatively less complex and contained environment on Earth. To study the direct effect of the space environment on the fundamental process of photosynthesis, we sent into low Earth orbit space engineered and mutated strains of the unicellular green alga, Chlamydomonas reinhardtii, which has been widely used as a model of photosynthetic organisms. The algal mutants contained specific amino acid substitutions in the functionally important regions of the pivotal Photosystem II (PSII) reaction centre D1 protein near the QB binding pocket and in the environment surrounding Tyr-161 (YZ) electron acceptor of the oxygen-evolving complex. Using real-time measurements of PSII photochemistry, here we show that during the space flight while the control strain and two D1 mutants (A250L and V160A) were inefficient in carrying out PSII activity, two other D1 mutants, I163N and A251C, performed efficient photosynthesis, and actively re-grew upon return to Earth. Mimicking the neutron irradiation component of cosmic rays on Earth yielded similar results. Experiments with I163N and A251C D1 mutants performed on ground showed that they are better able to modulate PSII excitation pressure and have higher capacity to reoxidize the QA (-) state of the primary electron acceptor. These results highlight the contribution of D1 conformation in relation to photosynthesis and oxygen production in space.

  19. Whole Genome Re-Sequencing Identifies a Quantitative Trait Locus Repressing Carbon Reserve Accumulation during Optimal Growth in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Goold, Hugh Douglas; Nguyen, Hoa Mai; Kong, Fantao; Beyly-Adriano, Audrey; Légeret, Bertrand; Billon, Emmanuelle; Cuiné, Stéphan; Beisson, Fred; Peltier, Gilles; Li-Beisson, Yonghua

    2016-05-04

    Microalgae have emerged as a promising source for biofuel production. Massive oil and starch accumulation in microalgae is possible, but occurs mostly when biomass growth is impaired. The molecular networks underlying the negative correlation between growth and reserve formation are not known. Thus isolation of strains capable of accumulating carbon reserves during optimal growth would be highly desirable. To this end, we screened an insertional mutant library of Chlamydomonas reinhardtii for alterations in oil content. A mutant accumulating five times more oil and twice more starch than wild-type during optimal growth was isolated and named constitutive oil accumulator 1 (coa1). Growth in photobioreactors under highly controlled conditions revealed that the increase in oil and starch content in coa1 was dependent on light intensity. Genetic analysis and DNA hybridization pointed to a single insertional event responsible for the phenotype. Whole genome re-sequencing identified in coa1 a >200 kb deletion on chromosome 14 containing 41 genes. This study demonstrates that, 1), the generation of algal strains accumulating higher reserve amount without compromising biomass accumulation is feasible; 2), light is an important parameter in phenotypic analysis; and 3), a chromosomal region (Quantitative Trait Locus) acts as suppressor of carbon reserve accumulation during optimal growth.

  20. Evidence for Nuclear Control of the Expression of the atpA and atpB Chloroplast Genes in Chlamydomonas.

    Science.gov (United States)

    Drapier, D.; Girard-Bascou, J.; Wollman, F. A.

    1992-03-01

    We analyzed three nuclear mutants of Chlamydomonas reinhardtii altered in the expression of the chloroplast genes atpA or atpB coding for the [alpha] or [beta] subunit of the chloroplast ATP synthase. These mutants revealed the existence of three nuclear products controlling the expression of the two chloroplast genes: the first one acts on the translation of the atpA transcript, and the two others act specifically on the stability of either the atpB or the atpA mRNAs. The nuclear mutation responsible for the decreased stability of the atpB mRNA prevented translation of the corresponding polypeptide. In contrast, the mutation responsible for the decreased stability of the atpA mRNA had limited effect on the translation of the [alpha] subunit, thereby allowing its accumulation and assembly in an active ATP synthase. Although acting originally on the expression of only one of the two main coupling factor 1 subunits, the three mutations caused a change in the translation rate of the other subunit, as viewed in 5-min pulse labeling experiments. This is indicative of a concerted expression of the [alpha] and [beta] subunits at an early post-translational step, or during translation, that may be critical for the assembly of the chloroplast ATP synthase.

  1. Whole Genome Re-Sequencing Identifies a Quantitative Trait Locus Repressing Carbon Reserve Accumulation during Optimal Growth in Chlamydomonas reinhardtii

    Science.gov (United States)

    Goold, Hugh Douglas; Nguyen, Hoa Mai; Kong, Fantao; Beyly-Adriano, Audrey; Légeret, Bertrand; Billon, Emmanuelle; Cuiné, Stéphan; Beisson, Fred; Peltier, Gilles; Li-Beisson, Yonghua

    2016-01-01

    Microalgae have emerged as a promising source for biofuel production. Massive oil and starch accumulation in microalgae is possible, but occurs mostly when biomass growth is impaired. The molecular networks underlying the negative correlation between growth and reserve formation are not known. Thus isolation of strains capable of accumulating carbon reserves during optimal growth would be highly desirable. To this end, we screened an insertional mutant library of Chlamydomonas reinhardtii for alterations in oil content. A mutant accumulating five times more oil and twice more starch than wild-type during optimal growth was isolated and named constitutive oil accumulator 1 (coa1). Growth in photobioreactors under highly controlled conditions revealed that the increase in oil and starch content in coa1 was dependent on light intensity. Genetic analysis and DNA hybridization pointed to a single insertional event responsible for the phenotype. Whole genome re-sequencing identified in coa1 a >200 kb deletion on chromosome 14 containing 41 genes. This study demonstrates that, 1), the generation of algal strains accumulating higher reserve amount without compromising biomass accumulation is feasible; 2), light is an important parameter in phenotypic analysis; and 3), a chromosomal region (Quantitative Trait Locus) acts as suppressor of carbon reserve accumulation during optimal growth. PMID:27141848

  2. X-Ray Fiber Diffraction Recordings from Oriented Demembranated Chlamydomonas Flagellar Axonemes.

    Science.gov (United States)

    Toba, Shiori; Iwamoto, Hiroyuki; Kamimura, Shinji; Oiwa, Kazuhiro

    2015-06-16

    The high homology of its axonemal components with humans and a large repertoire of axonemal mutants make Chlamydomonas a useful model system for experiments on the structure and function of eukaryotic cilia and flagella. Using this organism, we explored the spatial arrangement of axonemal components under physiological conditions by small-angle x-ray fiber diffraction. Axonemes were oriented in physiological solution by continuous shear flow and exposed to intense and stable x rays generated in the synchrotron radiation facility SPring-8, BL45XU. We compared diffraction patterns from axonemes isolated from wild-type and mutant strains lacking the whole outer arm (oda1), radial spoke (pf14), central apparatus (pf18), or the α-chain of the outer arm dynein (oda11). Diffraction of the axonemes showed a series of well-defined meridional/layer-line and equatorial reflections. Diffraction patterns from mutant axonemes exhibited a systematic loss/attenuation of meridional/layer-line reflections, making it possible to determine the origin of various reflections. The 1/24 and 1/12 nm(-1) meridional reflections of oda1 and oda11 were much weaker than those of the wild-type, suggesting that the outer dynein arms are the main contributor to these reflections. The weaker 1/32 and 1/13.7 nm(-1) meridional reflections from pf14 compared with the wild-type suggest that these reflections come mainly from the radial spokes. The limited contribution of the central pair apparatus to the diffraction patterns was confirmed by the similarity between the patterns of the wild-type and pf18. The equatorial reflections were complex, but a comparison with electron micrograph-based models allowed the density of each axonemal component to be estimated. Addition of ATP to rigor-state axonemes also resulted in subtle changes in equatorial intensity profiles, which could report nucleotide-dependent structural changes of the dynein arms. The first detailed description of axonemal reflections

  3. Universal Numeric Segmented Display

    CERN Document Server

    Azad, Md Abul kalam; Kamruzzaman, S M

    2010-01-01

    Segmentation display plays a vital role to display numerals. But in today's world matrix display is also used in displaying numerals. Because numerals has lots of curve edges which is better supported by matrix display. But as matrix display is costly and complex to implement and also needs more memory, segment display is generally used to display numerals. But as there is yet no proposed compact display architecture to display multiple language numerals at a time, this paper proposes uniform display architecture to display multiple language digits and general mathematical expressions with higher accuracy and simplicity by using a 18-segment display, which is an improvement over the 16 segment display.

  4. Transcriptome-Wide Changes in Chlamydomonas reinhardtii Gene Expression Regulated by Carbon Dioxide and the CO2-Concentrating Mechanism Regulator CIA5/CCM1[W][OA

    Science.gov (United States)

    Fang, Wei; Si, Yaqing; Douglass, Stephen; Casero, David; Merchant, Sabeeha S.; Pellegrini, Matteo; Ladunga, Istvan; Liu, Peng; Spalding, Martin H.

    2012-01-01

    We used RNA sequencing to query the Chlamydomonas reinhardtii transcriptome for regulation by CO2 and by the transcription regulator CIA5 (CCM1). Both CO2 and CIA5 are known to play roles in acclimation to low CO2 and in induction of an essential CO2-concentrating mechanism (CCM), but less is known about their interaction and impact on the whole transcriptome. Our comparison of the transcriptome of a wild type versus a cia5 mutant strain under three different CO2 conditions, high CO2 (5%), low CO2 (0.03 to 0.05%), and very low CO2 (<0.02%), provided an entry into global changes in the gene expression patterns occurring in response to the interaction between CO2 and CIA5. We observed a massive impact of CIA5 and CO2 on the transcriptome, affecting almost 25% of all Chlamydomonas genes, and we discovered an array of gene clusters with distinctive expression patterns that provide insight into the regulatory interaction between CIA5 and CO2. Several individual clusters respond primarily to either CIA5 or CO2, providing access to genes regulated by one factor but decoupled from the other. Three distinct clusters clearly associated with CCM-related genes may represent a rich source of candidates for new CCM components, including a small cluster of genes encoding putative inorganic carbon transporters. PMID:22634760

  5. Transcriptome-Wide Changes in Chlamydomonas reinhardtii Gene Expression Regulated by Carbon Dioxide and the CO2-Concentrating Mechanism Regulator CIA5/CCM1

    Energy Technology Data Exchange (ETDEWEB)

    Fang, W; Si, YQ; Douglass, S; Casero, D; Merchant, SS; Pellegrini, M; Ladunga, I; Liu, P; Spalding, MH

    2012-06-26

    We used RNA sequencing to query the Chlamydomonas reinhardtii transcriptome for regulation by CO2 and by the transcription regulator CIA5 (CCM1). Both CO2 and CIA5 are known to play roles in acclimation to low CO2 and in induction of an essential CO2-concentrating mechanism (CCM), but less is known about their interaction and impact on the whole transcriptome. Our comparison of the transcriptome of a wild type versus a cia5 mutant strain under three different CO2 conditions, high CO2 (5%), low CO2 (0.03 to 0.05%), and very low CO2 (< 0.02%), provided an entry into global changes in the gene expression patterns occurring in response to the interaction between CO2 and CIA5. We observed a massive impact of CIA5 and CO2 on the transcriptome, affecting almost 25% of all Chlamydomonas genes, and we discovered an array of gene clusters with distinctive expression patterns that provide insight into the regulatory interaction between CIA5 and CO2. Several individual clusters respond primarily to either CIA5 or CO2, providing access to genes regulated by one factor but decoupled from the other. Three distinct clusters clearly associated with CCM-related genes may represent a rich source of candidates for new CCM components, including a small cluster of genes encoding putative inorganic carbon transporters.

  6. Genetic and biochemical analysis of the TLA1 gene in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Mitra, Mautusi; Melis, Anastasios

    2010-02-01

    The Chlamydomonas reinhardtii genomic DNA database contains a predicted open reading frame (ORF-P) without an apparent stop-codon and unknown coding sequence, located in close proximity and immediately upstream of the TLA1 gene (GenBank Accession No. AF534570). The latter was implicated in the regulation of the light-harvesting chlorophyll antenna size of photosynthesis (Tetali et al. Planta 225:813-829, 2007). To provide currently lacking information on ORF-P and its potential participation in TLA1 gene expression, thus in the regulation of the chlorophyll antenna size, genetic and biochemical analyses were undertaken. The coding and UTR regions of the ORF-P were defined and delineated from those of the adjacent TLA1 gene. ORF-P is shown to encode a protein with a distinct RING-like zinc finger domain that is present in numerous eukaryotic proteins, believed to play a role in cellular ubiquitination, leading to regulation of cellular processes like signaling, growth, transcription, and DNA repair. It is further shown that the two genes share a 74-bp overlap between the 3' UTR region of ORF-P and the 5' UTR region of TLA1. However, they possess distinct start and stop codons and separate coding sequences, and transcribed as separate mRNAs without any trans-splicing between them. Complementation experiments showed that the TLA1 gene alone is sufficient to rescue the truncated chlorophyll antenna size phenotype of the tla1 mutant. Protein sequence alignments in C. reinhardtii and the colorless microalga Polytomella parva suggested that TLA1 defines the relationship between nucleus and organelle in microalgae, indirectly affecting the development of the chlorophyll antenna size.

  7. Phosphorylation controls the localization and activation of the lumenal carbonic anhydrase in Chlamydomonas reinhardtii.

    Directory of Open Access Journals (Sweden)

    Amaya Blanco-Rivero

    Full Text Available BACKGROUND: Cah3 is the only carbonic anhydrase (CA isoform located in the thylakoid lumen of Chlamydomonas reinhardtii. Previous studies demonstrated its association with the donor side of the photosystem II (PSII where it is required for the optimal function of the water oxidizing complex. However this enzyme has also been frequently proposed to perform a critical function in inorganic carbon acquisition and CO(2 fixation and all mutants lacking Cah3 exhibit very poor growth after transfer to low CO(2 conditions. RESULTS/CONCLUSIONS: In the present work we demonstrate that after transfer to low CO(2, Cah3 is phosphorylated and that phosphorylation is correlated to changes in its localization and its increase in activity. When C. reinhardtii wild-type cells were acclimated to limiting CO(2 conditions, the Cah3 activity increased about 5-6 fold. Under these conditions, there were no detectable changes in the level of the Cah3 polypeptide. The increase in activity was specifically inhibited in the presence of Staurosporine, a protein kinase inhibitor, suggesting that the Cah3 protein was post-translationally regulated via phosphorylation. Immunoprecipitation and in vitro dephosphorylation experiments confirm this hypothesis. In vivo phosphorylation analysis of thylakoid polypeptides indicates that there was a 3-fold increase in the phosphorylation signal of the Cah3 polypeptide within the first two hours after transfer to low CO(2 conditions. The increase in the phosphorylation signal was correlated with changes in the intracellular localization of the Cah3 protein. Under high CO(2 conditions, the Cah3 protein was only associated with the donor side of PSII in the stroma thylakoids. In contrast, in cells grown at limiting CO(2 the protein was partly concentrated in the thylakoids crossing the pyrenoid, which did not contain PSII and were surrounded by Rubisco molecules. SIGNIFICANCE: This is the first report of a CA being post

  8. A deoxyribonuclease from Chlamydomonas reinhardii. 1. Purification and properties.

    Science.gov (United States)

    Tait, G C; Harris, W J

    1977-05-16

    A deoxyribonuclease has been purified more than 2000-fold from the green algae, Chlamydomonas reinhardii. The enzyme is most active on denatured DNA. Optimum activity is at pH 8.5, in 80 mM Tris-HCl buffer and 2 mM CaCl2. Other divalent cations can replace Ca2+ with varying lower efficiency. EDTA and inorganic phosphate are strongly inhibitory, while ATP and high concentrations of 2-mercaptoethanol are slightly inhibitory. The molecular weight is approximately 35 000, the Stokes radius is 2.7 nm, and the sedimentation coefficient 2.8 S. It is a single polypeptide chain, and the frictional ratio of 1.27 suggests it is only slightly asymetrical. The isoelectric point is 9.5. This enzyme has been termed exonuclease 1.

  9. Cilia and ciliopathies:From Chlamydomonas and beyond

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    The biological function of motile cilia/flagella has long been recognized. The non-motile primary cilium, once regarded as a vestigial organelle, however, has been found recently to play unexpected roles in mammalian physiology and development. Defects in cilia have profound impact on human health. Diseases related to cilia, collectively called ciliopathies include male infertility, primary cilia dyskinesia, renal cyst formation, blindness, polydactyly, obesity, hypertension, and even mental retardation. Our current understanding of cilia and ciliopathies has been fueled by basic research employing various model organisms including Chlamydomonas, a unicellular green alga. This review article provides a general introduction to the cell biology of cilia and an overview of various cilia-related diseases.

  10. Real-time monitoring of genetically modified Chlamydomonas reinhardtii during the Foton M3 space mission

    Science.gov (United States)

    Lambreva, M.; Rea, G.; Antonacci, A.; Serafini, A.; Damasso, M.; Pastorelli, S.; Margonelli, A.; Johanningmeier, U.; Bertalan, I.; Pezzotti, G.; Giardi, M. T.

    2008-09-01

    Long-term space exploration, colonization or habitation requires biological life support systems capable to cope with the deleterious space environment. The use of oxygenic photosynthetic microrganisms is an intriguing possibility mainly for food, O2 and nutraceutical compounds production. The critical points of utilizing plants- or algae-based life support systems are the microgravity and the ionizing radiation, which can influence the performance of these organisms. The aim of the present study was to assess the effects of space environment on the photosynthetic activity of various microrganisms and to select space stresstolerant strains. Photosystem II D1 protein sitedirected and random mutants of the unicellular green alga Chlamydomonas reinhardtii [1] were used as a model system to test and select the amino acid substitutions capable to account for space stress tolerance. We focussed our studies also on the accumulation of the Photosystem II photoprotective carotenoids (the xantophylls violaxanthin, anteraxanthin and zeaxanthin), powerful antioxidants that epidemiological studies demonstrated to be human vision protectors. For this purpose some mutants modified at the level of enzymes involved in the biosynthesis of xanthophylls were included in the study [2]. To identify the consequences of the space environment on the photosynthetic apparatus the changes in the Photosystem II efficiency were monitored in real time during the ESA-Russian Foton- M3 mission in September 2007. For the space flight a high-tech, multicell fluorescence detector, Photo-II, was designed and built by the Centre for Advanced Research in Space Optics in collaboration with Kayser-Italy, Biosensor and DAS. Photo-II is an automatic device developed to measure the chlorophyll fluorescence and to provide a living conditions for several different algae strains (Fig.1). Twelve different C. reinhardti strains were analytically selected and two replications for each strain were brought to space

  11. Preliminary study on a gravity-insensitive rice mutant

    Institute of Scientific and Technical Information of China (English)

    金静; 朱诚; 张红心; 孙宗修

    2004-01-01

    A gravity-insensitive mutant was isolated from rice (Oryza sativa L. Cv. Zhonghua 11) transformed by Agrobacterium tumefaciens. The mutant's shoot growth (prostrate growth) was insensitive to gravity; whereas root growth displayed a normal positive gravitropism.Histological observation of root caps and leaf sheaths indicated that there was no significant difference in the number and size of amyloplasts in cells of the mutant and cells of the wild type

  12. Preliminary study on a gravity-insensitive rice mutant

    Institute of Scientific and Technical Information of China (English)

    金静; 朱诚; 张红心; 孙宗修

    2004-01-01

    A gravity-insensitive mutant was isolated from rice (Oryza sativa L. cv. Zhonghua 11) transformed by Agrobacterium tumefaciens. The mutant's shoot growth (prostrate growth) was insensitive to gravity; whereas root growth displayed a normal positive gravitropism. Histological observation of root caps and leaf sheaths indicated that there was no significant difference in the number and size of amyloplasts in cells of the mutant and cells of the wild type.

  13. LHCSR3 affects de-coupling and re-coupling of LHCII to PSII during state transitions in Chlamydomonas reinhardtii

    Science.gov (United States)

    Roach, Thomas; Na, Chae Sun

    2017-01-01

    Photosynthetic organisms have to tolerate rapid changes in light intensity, which is facilitated by non-photochemical quenching (NPQ) and involves modification of energy transfer from light-harvesting complexes (LHC) to the photosystem reaction centres. NPQ includes dissipating excess light energy to heat (qE) and the reversible coupling of LHCII to photosystems (state transitions/qT), which are considered separate NPQ mechanisms. In the model alga Chlamydomonas reinhardtii the LHCSR3 protein has a well characterised role in qE. Here, it is shown in the npq4 mutant, deficient in LHCSR3, that energy coupling to photosystem II (PSII) more akin to qT is also disrupted, but no major differences in LHC phosphorylation or LHC compositions were found in comparison to wild-type cells. The qT of wild-type cells possessed two kinetically distinguishable phases, with LHCSR3 participating in the more rapid (<2 min) phase. This LHCSR3-mediated qT was sensitive to physiological levels of H2O2, which accelerated qE induction, revealing a way that may help C. reinhardtii tolerate a sudden increase in light intensity. Overall, a clear mechanistic overlap between qE and qT is shown. PMID:28233792

  14. A Chlamydomonas Homologue of the Putative Murine t Complex Distorter Tctex-2 Is an Outer Arm Dynein Light Chain

    Science.gov (United States)

    Patel-King, Ramila S.; Benashski, Sharon E.; Harrison, Alistair; King, Stephen M.

    1997-01-01

    Molecular analysis of a 19,000-Mr protein from the Chlamydomonas flagellum reveals that it is homologous to the t complex–encoded protein Tctex-2, which is a candidate for one of the distorter products that cause the extreme transmission ratio distortion (meiotic drive) of the murine t complex. The 19,000-Mr protein is extracted from the axoneme with 0.6 M NaCl and comigrates with the outer dynein arm in sucrose density gradients. This protein also is specifically missing in axonemes prepared from a mutant that does not assemble the outer arm. These data raise the possibility that Tctex-2 is a sperm flagellar dynein component. Combined with the recent identification of Tctex-1 (another distorter candidate) as a light chain of cytoplasmic dynein, these results lead to a biochemical model for how differential defects in spermiogenesis that result in the phenomenon of meiotic drive might be generated in wild-type vs t-bearing sperm. PMID:9166408

  15. Chlamydomonas axonemal dynein assembly locus ODA8 encodes a conserved flagellar protein needed for cytoplasmic maturation of outer dynein arm complexes.

    Science.gov (United States)

    Desai, Paurav B; Freshour, Judy R; Mitchell, David R

    2015-01-01

    The Chlamydomonas reinhardtii oda8 mutation blocks assembly of flagellar outer dynein arms (ODAs), and interacts genetically with ODA5 and ODA10, which encode axonemal proteins thought to aid dynein binding onto axonemal docking sites. We positionally cloned ODA8 and identified the gene product as the algal homolog of vertebrate LRRC56. Its flagellar localization depends on ODA5 and ODA10, consistent with genetic interaction studies, but phylogenomics suggests that LRRC56 homologs play a role in intraflagellar transport (IFT)-dependent assembly of outer row dynein arms, not axonemal docking. ODA8 distribution between cytoplasm and flagella is similar to that of IFT proteins and about half of flagellar ODA8 is in the soluble matrix fraction. Dynein extracted in vitro from wild type axonemes will rebind efficiently to oda8 mutant axonemes, without re-binding of ODA8, further supporting a role in dynein assembly or transport, not axonemal binding. Assays comparing preassembled ODA complexes from the cytoplasm of wild type and mutant strains show that dynein in oda8 mutant cytoplasm has not properly preassembled and cannot bind normally onto oda axonemes. We conclude that ODA8 plays an important role in formation and transport of mature dynein complexes during flagellar assembly.

  16. The Chlamydomonas Genome Reveals the Evolution of Key Animal and Plant Functions

    Energy Technology Data Exchange (ETDEWEB)

    Merchant, Sabeeha S

    2007-04-09

    Chlamydomonas reinhardtii is a unicellular green alga whose lineage diverged from land plants over 1 billion years ago. It is a model system for studying chloroplast-based photosynthesis, as well as the structure, assembly, and function of eukaryotic flagella (cilia), which were inherited from the common ancestor of plants and animals, but lost in land plants. We sequenced the 120-megabase nuclear genome of Chlamydomonas and performed comparative phylogenomic analyses, identifying genes encoding uncharacterized proteins that are likely associated with the function and biogenesis of chloroplasts or eukaryotic flagella. Analyses of the Chlamydomonas genome advance our understanding of the ancestral eukaryotic cell, reveal previously unknown genes associated with photosynthetic and flagellar functions, and establish links between ciliopathy and the composition and function of flagella.

  17. Absence of lutein, violaxanthin and neoxanthin affects the functional chlorophyll antenna size of photosystem-II but not that of photosystem-I in the green alga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Polle, J E; Niyogi, K K; Melis, A

    2001-05-01

    Chlamydomonas reinhardtii double mutant npq2 lor1 lacks the beta, epsilon-carotenoids lutein and loroxanthin as well as all beta,beta-epoxycarotenoids derived from zeaxanthin (e.g. violaxanthin and neoxanthin). Thus, the only carotenoids present in the thylakoid membranes of the npq2 lor1 cells are beta-carotene and zeaxanthin. The effect of these mutations on the photochemical apparatus assembly and function was investigated. In cells of the mutant strain, the content of photosystem-II (PSII) and photosystem-I (PSI) was similar to that of the wild type, but npq2 lor1 had a significantly smaller PSII light-harvesting Chl antenna size. In contrast, the Chl antenna size of PSI was not truncated in the mutant. SDS-PAGE and Western blot analysis qualitatively revealed the presence of all LHCII and LHCI apoproteins in the thylakoid membrane of the mutant. The results showed that some of the LHCII and most of the LHCI were assembled and functionally connected with PSII and PSI, respectively. Photon conversion efficiency measurements, based on the initial slope of the light-saturation curve of photosynthesis and on the yield of Chl a fluorescence in vivo, showed similar efficiencies. However, a significantly greater light intensity was required for the saturation of photosynthesis in the mutant than in the wild type. It is concluded that zeaxanthin can successfully replace lutein and violaxanthin in most of the functional light-harvesting antenna of the npq2 lor1 mutant.

  18. Retrograde bilin signaling enables Chlamydomonas greening and phototrophic survival

    Science.gov (United States)

    Duanmu, Deqiang; Casero, David; Dent, Rachel M.; Gallaher, Sean; Yang, Wenqiang; Rockwell, Nathan C.; Martin, Shelley S.; Pellegrini, Matteo; Niyogi, Krishna K.; Merchant, Sabeeha S.; Grossman, Arthur R.; Lagarias, J. Clark

    2013-01-01

    The maintenance of functional chloroplasts in photosynthetic eukaryotes requires real-time coordination of the nuclear and plastid genomes. Tetrapyrroles play a significant role in plastid-to-nucleus retrograde signaling in plants to ensure that nuclear gene expression is attuned to the needs of the chloroplast. Well-known sites of synthesis of chlorophyll for photosynthesis, plant chloroplasts also export heme and heme-derived linear tetrapyrroles (bilins), two critical metabolites respectively required for essential cellular activities and for light sensing by phytochromes. Here we establish that Chlamydomonas reinhardtii, one of many chlorophyte species that lack phytochromes, can synthesize bilins in both plastid and cytosol compartments. Genetic analyses show that both pathways contribute to iron acquisition from extracellular heme, whereas the plastid-localized pathway is essential for light-dependent greening and phototrophic growth. Our discovery of a bilin-dependent nuclear gene network implicates a widespread use of bilins as retrograde signals in oxygenic photosynthetic species. Our studies also suggest that bilins trigger critical metabolic pathways to detoxify molecular oxygen produced by photosynthesis, thereby permitting survival and phototrophic growth during the light period. PMID:23345435

  19. Lipidomic Analysis of Chlamydomonas reinhardtii under Nitrogen and Sulfur Deprivation.

    Directory of Open Access Journals (Sweden)

    Dawei Yang

    Full Text Available Chlamydomonas reinhardtii accumulates lipids under complete nutrient starvation conditions while overall growth in biomass stops. In order to better understand biochemical changes under nutrient deprivation that maintain production of algal biomass, we used a lipidomic assay for analyzing the temporal regulation of the composition of complex lipids in C. reinhardtii in response to nitrogen and sulfur deprivation. Using a chip-based nanoelectrospray direct infusion into an ion trap mass spectrometer, we measured a diversity of lipid species reported for C. reinhardtii, including PG phosphatidylglycerols, PI Phosphatidylinositols, MGDG monogalactosyldiacylglycerols, DGDG digalactosyldiacylglycerols, SQDG sulfoquinovosyldiacylglycerols, DGTS homoserine ether lipids and TAG triacylglycerols. Individual lipid species were annotated by matching mass precursors and MS/MS fragmentations to the in-house LipidBlast mass spectral database and MS2Analyzer. Multivariate statistics showed a clear impact on overall lipidomic phenotypes on both the temporal and the nutrition stress level. Homoserine-lipids were found up-regulated at late growth time points and higher cell density, while triacyclglycerols showed opposite regulation of unsaturated and saturated fatty acyl chains under nutritional deprivation.

  20. Response of Chlamydomonas reinhardtii to naphthenic acid exposure

    Energy Technology Data Exchange (ETDEWEB)

    Goff, K.; Wilson, K. [Saskatchewan Univ., Saskatoon, SK (Canada); Headley, J. [Environment Canada, Ottawa, ON (Canada)

    2010-07-01

    This study examined the feasibility of using a model organism for the algal bioremediation of oil sands process water (OSPW), a highly toxic mixture of sediments, bitumen, ions, and organic and inorganic compounds. Naphthenic acids (NAs) are a contaminant class of particular concern. Bioremediation techniques may mitigate toxicity of OSPW in general, and NAs in particular. Although most studies on the biodegradation of NAs focus on the role of bacteria, fungi, and emergent macrophytes, studies have indicated that algae may also play a key role through direct degradation, biosequestration, or photosynthetic aeration of waters to promote other biological reactions. Chlamydomonas frigida is of particular interest, but no cultures are currently available. Therefore, this study used C. reinhardtii, a well-characterized model organism, to begin analysis of potential algal bioremediation of OSPW. Cultures of C. reinhardtii were grown heterotrophically in nutrient media spiked with a dilution series of NAs. Culture densities were measured to compile growth curves over time, changes in rate of growth, and survivability. Negative ion electrospray mass spectrometry was used to determine the concentration of NAs in solution in relation to growth rate and culture density. The study determined the tolerance of C. reinhardtii to NAs. A mechanism for this tolerance was then proposed.

  1. Insecticides induced biochemical changes in freshwater microalga Chlamydomonas mexicana.

    Science.gov (United States)

    Kumar, Muthukannan Satheesh; Kabra, Akhil N; Min, Booki; El-Dalatony, Marwa M; Xiong, Jiuqiang; Thajuddin, Nooruddin; Lee, Dae Sung; Jeon, Byong-Hun

    2016-01-01

    The effect of insecticides (acephate and imidacloprid) on a freshwater microalga Chlamydomonas mexicana was investigated with respect to photosynthetic pigments, carbohydrate and protein contents, fatty acids composition and induction of stress indicators including proline, superoxide dismutase (SOD) and catalase (CAT). C. mexicana was cultivated with 1, 5, 10, 15, 20 and 25 mg L(-1) of acephate and imidacloprid. The microalga growth increased with increasing concentrations of both insecticides up to 15 mg L(-1), beyond which the growth declined compared to control condition (without insecticides). C. mexicana cultivated with 15 mg L(-1) of both insecticides for 12 days was used for further analysis. The accumulation of photosynthetic pigments (chlorophyll and carotenoids), carbohydrates and protein was decreased in the presence of both insecticides. Acephate and imidacloprid induced the activities of superoxide dismutase (SOD) and catalase (CAT) and increased the concentration of proline in the microalga, which play a defensive role against various environmental stresses. Fatty acid analysis revealed that the fraction of polyunsaturated fatty acids decreased on exposure to both insecticides. C. mexicana also promoted 25 and 21% removal of acephate and imidacloprid, respectively. The biochemical changes in C. mexicana on exposure to acephate and imidacloprid indicate that the microalga undergoes an adaptive change in response to the insecticide-induced oxidative stress.

  2. Establishing Chlamydomonas reinhardtii as an industrial biotechnology host.

    Science.gov (United States)

    Scaife, Mark A; Nguyen, Ginnie T D T; Rico, Juan; Lambert, Devinn; Helliwell, Katherine E; Smith, Alison G

    2015-05-01

    Microalgae constitute a diverse group of eukaryotic unicellular organisms that are of interest for pure and applied research. Owing to their natural synthesis of value-added natural products microalgae are emerging as a source of sustainable chemical compounds, proteins and metabolites, including but not limited to those that could replace compounds currently made from fossil fuels. For the model microalga, Chlamydomonas reinhardtii, this has prompted a period of rapid development so that this organism is poised for exploitation as an industrial biotechnology platform. The question now is how best to achieve this? Highly advanced industrial biotechnology systems using bacteria and yeasts were established in a classical metabolic engineering manner over several decades. However, the advent of advanced molecular tools and the rise of synthetic biology provide an opportunity to expedite the development of C. reinhardtii as an industrial biotechnology platform, avoiding the process of incremental improvement. In this review we describe the current status of genetic manipulation of C. reinhardtii for metabolic engineering. We then introduce several concepts that underpin synthetic biology, and show how generic parts are identified and used in a standard manner to achieve predictable outputs. Based on this we suggest that the development of C. reinhardtii as an industrial biotechnology platform can be achieved more efficiently through adoption of a synthetic biology approach.

  3. Modulation of Chlamydomonas reinhardtii flagellar motility by redox poise

    Science.gov (United States)

    Wakabayashi, Ken-ichi; King, Stephen M.

    2006-01-01

    Redox-based regulatory systems are essential for many cellular activities. Chlamydomonas reinhardtii exhibits alterations in motile behavior in response to different light conditions (photokinesis). We hypothesized that photokinesis is signaled by variations in cytoplasmic redox poise resulting from changes in chloroplast activity. We found that this effect requires photosystem I, which generates reduced NADPH. We also observed that photokinetic changes in beat frequency and duration of the photophobic response could be obtained by altering oxidative/reductive stress. Analysis of reactivated cell models revealed that this redox poise effect is mediated through the outer dynein arms (ODAs). Although the global redox state of the thioredoxin-related ODA light chains LC3 and LC5 and the redox-sensitive Ca2+-binding subunit of the docking complex DC3 did not change upon light/dark transitions, we did observe significant alterations in their interactions with other flagellar components via mixed disulfides. These data indicate that redox poise directly affects ODAs and suggest that it may act in the control of flagellar motility. PMID:16754958

  4. Expression of human soluble TRAIL in Chlamydomonas reinhardtii chloroplast

    Institute of Scientific and Technical Information of China (English)

    YANG Zongqi; LI yinü; CHEN Feng; LI Dong; ZHANG Zhifang; LIU Yanxin; ZHENG Dexian; WANG Yong; SHEN Guifang

    2006-01-01

    Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces selectively apoptosis in various tumor cells and virus-infected cells, but rarely in normal cells. A chloroplast expression vector, p64TRAIL, containing the cDNA coding for the soluble TRAIL (sTRAIL), was constructed with clpP-trnL-petB-chlL-rpl23-rpl2 as Chlamydomonas reinhardtii plastid homologous recombinant fragments and spectinomycin-resistant aadA gene as a select marker. The plasmid p64TRAIL was transferred into the chloroplast genome of C. reinhardtii by the biolistic method. Three independently transformed lines were obtained by 100 mg/L spectinomycin selection. PCR amplification, Southern blot analysis of the sTRAIL coding region DNA and cultivation cells in the dark all showed that the exogenous DNA had been integrated into chloroplast genome of C. reinhardtii. Western blot analysis showed that human soluble TRAIL was expressed in C. reinhardtii chloroplast. The densitometric analysis of Western blot indicated that the expressed human sTRAIL protein in the chloroplasts of C. reinhardtii accounted for about 0.43%-0.67% of the total soluble proteins.These experimental results demonstrated the possibility of using transgenic chloroplasts of green alga as bioreactors for production of biopharmaceuticals.

  5. Enhanced excision repair and lack of PSII activity contribute to higher UV survival of Chlamydomonas reinhardtii cells in dark.

    Science.gov (United States)

    Chaudhari, Vishalsingh R; Vyawahare, Aniket; Bhattacharjee, Swapan K; Rao, Basuthkar J

    2015-03-01

    Plant cells are known to differentiate their responses to stress depending up on the light conditions. We observed that UVC sensitive phenotype of light grown asynchronous Chlamydomonas reinhardtii culture (Light culture: LC) can be converted to relatively resistant form by transfer to dark condition (Dark culture: DC) before UVC exposure. The absence of photosystem II (PSII) function, by either atrazine treatment in wild type or in D1 (psbA) null mutant, conferred UV protection even in LC. We provide an indirect support for involvement of reactive oxygen species (ROS) signalling by showing higher UV survival on exposures to mild dose of H2O2 or Methyl Viologen. Circadian trained culture also showed a rhythmic variation in UV sensitivity in response to alternating light-dark (12 h:12 h) entrainment, with maximum UV survival at the end of 12 h dark and minimum at the end of 12 h light. This rhythm failed to maintain in "free running" conditions, making it a non-circadian phenotype. Moreover, atrazine strongly inhibited rhythmic UV sensitivity and conferred a constitutively high resistance, without affecting internal circadian rhythm marker expression. Dampening of UV sensitivity rhythm in Thymine-dimer excision repair mutant (cc-888) suggested the involvement of DNA repair in this phenomenon. DNA excision repair (ER) assays in cell-free extracts revealed that dark incubated cells exhibit higher ER compared to those growing in light, underscoring the role of ER in conferring differential UV sensitivity in dark versus light incubation. We suggest that multiple factors such as ROS changes triggered by differences in PSII activity, concomitant with differential ER efficiency collectively contribute to light-dark (12 h: 12 h) rhythmicity in C. reinhardtii UV sensitivity.

  6. Mutagenesis and phenotypic selection as a strategy toward domestication of Chlamydomonas reinhardtii strains for improved performance in photobioreactors.

    Science.gov (United States)

    Bonente, Giulia; Formighieri, Cinzia; Mantelli, Manuela; Catalanotti, Claudia; Giuliano, Giovanni; Morosinotto, Tomas; Bassi, Roberto

    2011-09-01

    Microalgae have a valuable potential for biofuels production. As a matter of fact, algae can produce different molecules with high energy content, including molecular hydrogen (H(2)) by the activity of a chloroplastic hydrogenase fueled by reducing power derived from water and light energy. The efficiency of this reaction, however, is limited and depends from an intricate relationships between oxygenic photosynthesis and mitochondrial respiration. The way toward obtaining algal strains with high productivity in photobioreactors requires engineering of their metabolism at multiple levels in a process comparable to domestication of crops that were derived from their wild ancestors through accumulation of genetic traits providing improved productivity under conditions of intensive cultivation as well as improved nutritional/industrial properties. This holds true for the production of any biofuels from algae: there is the need to isolate multiple traits to be combined and produce organisms with increased performances. Among the different limitations in H(2) productivity, we identified three with a major relevance, namely: (i) the light distribution through the mass culture; (ii) the strong sensitivity of the hydrogenase to even very low oxygen concentrations; and (iii) the presence of alternative pathways, such as the cyclic electron transport, competing for reducing equivalents with hydrogenase and H(2) production. In order to identify potentially favorable mutations, we generated a collection of random mutants in Chlamydomonas reinhardtii which were selected through phenotype analysis for: (i) a reduced photosynthetic antenna size, and thus a lower culture optical density; (ii) an altered photosystem II activity as a tool to manipulate the oxygen concentration within the culture; and (iii) State 1-State 2 transition mutants, for a reduced cyclic electron flow and maximized electrons flow toward the hydrogenase. Such a broad approach has been possible thanks to the

  7. Site Energies of Active and Inactive Pheophytins in the Reaction Center of Photosystem II from Chlamydomonas Reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Acharya, K.; Neupane, B.; Zazubovich, V.; Sayre, R. T.; Picorel, R.; Seibert, M.; Jankowiak, R.

    2012-03-29

    It is widely accepted that the primary electron acceptor in various Photosystem II (PSII) reaction center (RC) preparations is pheophytin {alpha} (Pheo {alpha}) within the D1 protein (Pheo{sub D1}), while Pheo{sub D2} (within the D2 protein) is photochemically inactive. The Pheo site energies, however, have remained elusive, due to inherent spectral congestion. While most researchers over the past two decades placed the Q{sub y}-states of Pheo{sub D1} and Pheo{sub D2} bands near 678-684 and 668-672 nm, respectively, recent modeling [Raszewski et al. Biophys. J. 2005, 88, 986-998; Cox et al. J. Phys. Chem. B 2009, 113, 12364-12374] of the electronic structure of the PSII RC reversed the assignment of the active and inactive Pheos, suggesting that the mean site energy of Pheo{sub D1} is near 672 nm, whereas Pheo{sub D2} ({approx}677.5 nm) and Chl{sub D1} ({approx}680 nm) have the lowest energies (i.e., the Pheo{sub D2}-dominated exciton is the lowest excited state). In contrast, chemical pigment exchange experiments on isolated RCs suggested that both pheophytins have their Q{sub y} absorption maxima at 676-680 nm [Germano et al. Biochemistry 2001, 40, 11472-11482; Germano et al. Biophys. J. 2004, 86, 1664-1672]. To provide more insight into the site energies of both Pheo{sub D1} and Pheo{sub D2} (including the corresponding Q{sub x} transitions, which are often claimed to be degenerate at 543 nm) and to attest that the above two assignments are most likely incorrect, we studied a large number of isolated RC preparations from spinach and wild-type Chlamydomonas reinhardtii (at different levels of intactness) as well as the Chlamydomonas reinhardtii mutant (D2-L209H), in which the active branch Pheo{sub D1} is genetically replaced with chlorophyll {alpha} (Chl {alpha}). We show that the Q{sub x}-/Q{sub y}-region site energies of Pheo{sub D1} and Pheo{sub D2} are {approx}545/680 nm and {approx}541.5/670 nm, respectively, in good agreement with our previous assignment

  8. Site energies of active and inactive pheophytins in the reaction center of Photosystem II from Chlamydomonas reinhardtii.

    Science.gov (United States)

    Acharya, K; Neupane, B; Zazubovich, V; Sayre, R T; Picorel, R; Seibert, M; Jankowiak, R

    2012-03-29

    It is widely accepted that the primary electron acceptor in various Photosystem II (PSII) reaction center (RC) preparations is pheophytin a (Pheo a) within the D1 protein (Pheo(D1)), while Pheo(D2) (within the D2 protein) is photochemically inactive. The Pheo site energies, however, have remained elusive, due to inherent spectral congestion. While most researchers over the past two decades placed the Q(y)-states of Pheo(D1) and Pheo(D2) bands near 678-684 and 668-672 nm, respectively, recent modeling [Raszewski et al. Biophys. J. 2005, 88, 986 - 998; Cox et al. J. Phys. Chem. B 2009, 113, 12364 - 12374] of the electronic structure of the PSII RC reversed the assignment of the active and inactive Pheos, suggesting that the mean site energy of Pheo(D1) is near 672 nm, whereas Pheo(D2) (~677.5 nm) and Chl(D1) (~680 nm) have the lowest energies (i.e., the Pheo(D2)-dominated exciton is the lowest excited state). In contrast, chemical pigment exchange experiments on isolated RCs suggested that both pheophytins have their Q(y) absorption maxima at 676-680 nm [Germano et al. Biochemistry 2001, 40, 11472 - 11482; Germano et al. Biophys. J. 2004, 86, 1664 - 1672]. To provide more insight into the site energies of both Pheo(D1) and Pheo(D2) (including the corresponding Q(x) transitions, which are often claimed to be degenerate at 543 nm) and to attest that the above two assignments are most likely incorrect, we studied a large number of isolated RC preparations from spinach and wild-type Chlamydomonas reinhardtii (at different levels of intactness) as well as the Chlamydomonas reinhardtii mutant (D2-L209H), in which the active branch Pheo(D1) is genetically replaced with chlorophyll a (Chl a). We show that the Q(x)-/Q(y)-region site energies of Pheo(D1) and Pheo(D2) are ~545/680 nm and ~541.5/670 nm, respectively, in good agreement with our previous assignment [Jankowiak et al. J. Phys. Chem. B 2002, 106, 8803 - 8814]. The latter values should be used to model excitonic

  9. Displaying gray shades in liquid crystal displays

    Indian Academy of Sciences (India)

    T N Ruckmongathan

    2003-08-01

    Quality of image in a display depends on the contrast, colour, resolution and the number of gray shades. A large number of gray shades is necessary to display images without any contour lines. These contours are due to limited number of gray shades in the display causing abrupt changes in grayness of the image, while the original image has a gradual change in brightness. Amplitude modulation has the capability to display a large number of gray shades with minimum number of time intervals [1,2]. This paper will cover the underlying principle of amplitude modulation, some variants and its extension to multi-line addressing. Other techniques for displaying gray shades in passive matrix displays are reviewed for the sake of comparison.

  10. Remodeling of Membrane Lipids in Iron-starved Chlamydomonas*

    Science.gov (United States)

    Urzica, Eugen I.; Vieler, Astrid; Hong-Hermesdorf, Anne; Page, M. Dudley; Casero, David; Gallaher, Sean D.; Kropat, Janette; Pellegrini, Matteo; Benning, Christoph; Merchant, Sabeeha S.

    2013-01-01

    Chlamydomonas reinhardtii cells exposed to abiotic stresses (e.g. nitrogen, zinc, or phosphorus deficiency) accumulate triacylglycerols (TAG), which are stored in lipid droplets. Here, we report that iron starvation leads to formation of lipid droplets and accumulation of TAGs. This occurs between 12 and 24 h after the switch to iron-starvation medium. C. reinhardtii cells deprived of iron have more saturated fatty acid (FA), possibly due to the loss of function of FA desaturases, which are iron-requiring enzymes with diiron centers. The abundance of a plastid acyl-ACP desaturase (FAB2) is decreased to the same degree as ferredoxin. Ferredoxin is a substrate of the desaturases and has been previously shown to be a major target of the iron deficiency response. The increase in saturated FA (C16:0 and C18:0) is concomitant with the decrease in unsaturated FA (C16:4, C18:3, or C18:4). This change was gradual for diacylglyceryl-N,N,N-trimethylhomoserine (DGTS) and digalactosyldiacylglycerol (DGDG), whereas the monogalactosyldiacylglycerol (MGDG) FA profile remained stable during the first 12 h, whereas MGDG levels were decreasing over the same period of time. These changes were detectable after only 2 h of iron starvation. On the other hand, DGTS and DGDG contents gradually decreased until a minimum was reached after 24–48 h. RNA-Seq analysis of iron-starved C. reinhardtii cells revealed notable changes in many transcripts coding for enzymes involved in FA metabolism. The mRNA abundances of genes coding for components involved in TAG accumulation (diacylglycerol acyltransferases or major lipid droplet protein) were increased. A more dramatic increase at the transcript level has been observed for many lipases, suggesting that major remodeling of lipid membranes occurs during iron starvation in C. reinhardtii. PMID:23983122

  11. Remodeling of membrane lipids in iron-starved Chlamydomonas.

    Science.gov (United States)

    Urzica, Eugen I; Vieler, Astrid; Hong-Hermesdorf, Anne; Page, M Dudley; Casero, David; Gallaher, Sean D; Kropat, Janette; Pellegrini, Matteo; Benning, Christoph; Merchant, Sabeeha S

    2013-10-18

    Chlamydomonas reinhardtii cells exposed to abiotic stresses (e.g. nitrogen, zinc, or phosphorus deficiency) accumulate triacylglycerols (TAG), which are stored in lipid droplets. Here, we report that iron starvation leads to formation of lipid droplets and accumulation of TAGs. This occurs between 12 and 24 h after the switch to iron-starvation medium. C. reinhardtii cells deprived of iron have more saturated fatty acid (FA), possibly due to the loss of function of FA desaturases, which are iron-requiring enzymes with diiron centers. The abundance of a plastid acyl-ACP desaturase (FAB2) is decreased to the same degree as ferredoxin. Ferredoxin is a substrate of the desaturases and has been previously shown to be a major target of the iron deficiency response. The increase in saturated FA (C16:0 and C18:0) is concomitant with the decrease in unsaturated FA (C16:4, C18:3, or C18:4). This change was gradual for diacylglyceryl-N,N,N-trimethylhomoserine (DGTS) and digalactosyldiacylglycerol (DGDG), whereas the monogalactosyldiacylglycerol (MGDG) FA profile remained stable during the first 12 h, whereas MGDG levels were decreasing over the same period of time. These changes were detectable after only 2 h of iron starvation. On the other hand, DGTS and DGDG contents gradually decreased until a minimum was reached after 24-48 h. RNA-Seq analysis of iron-starved C. reinhardtii cells revealed notable changes in many transcripts coding for enzymes involved in FA metabolism. The mRNA abundances of genes coding for components involved in TAG accumulation (diacylglycerol acyltransferases or major lipid droplet protein) were increased. A more dramatic increase at the transcript level has been observed for many lipases, suggesting that major remodeling of lipid membranes occurs during iron starvation in C. reinhardtii.

  12. Physiological characterization of cadmium-exposed Chlamydomonas reinhardtii.

    Science.gov (United States)

    Bräutigam, Anja; Schaumlöffel, Dirk; Preud'homme, Hugues; Thondorf, Iris; Wesenberg, Dirk

    2011-12-01

    Chlamydomonas reinhardtii is a common model organism for investigation of metal stress. This green alga produces phytochelatins in the presence of metal ions. The influence of cadmium is of main interest, because it is a strong activator of phytochelatin synthase. Cell wall bound and intracellular cadmium content was determined after exposition to 70 µm CdCl(2), showing the main portion of the metal outside the cell. Nevertheless, imported cadmium was sufficient to cause significant changes in thiolpeptide metabolism and its transcriptional regulation. Modern analytical approaches enable new insights into phytochelatin (PC) distribution. A new rapid and precise UPLC-MS method allowed high-throughput PC quantification in algal samples after 1, 4, 24 and 48 h cadmium stress. Initially, canonic PCs were synthesized in C. reinhardtii during cadmium exposition, but afterwards CysPCs became the major thiolpeptides. Thus, after 48 h the concentration of the PC-isoforms CysPC(2-3) and CysGSH attained between 105 and 199 nmol g(-1) fresh weight (FW), whereas the PC(2-3) concentrations were only 15 nmol g(-1) FW. The relative quantification of γ-glutamyl transpeptidase (γ-GT) mRNA suggests the generation of CysPCs by glutamate cleavage from canonic PCs by γ-GT. Furthermore, a homology model of C. reinhardtii phytochelatin synthase was constructed to verify the use of crystal structures from Anabaena sp. phytochelatin synthase (PCS) for docking studies with canonical PCs and CysPCs. From the difference in energy scores, we hypothesize that CysPC may prevent the synthesis of canonical PCs by blocking the binding pocket. Finally, possible physiological reasons for the high abundance of CysPC compared with their canonic precursors are discussed.

  13. Adaptation prevents the extinction of Chlamydomonas reinhardtii under toxic beryllium

    Directory of Open Access Journals (Sweden)

    Beatriz Baselga-Cervera

    2016-03-01

    Full Text Available The current biodiversity crisis represents a historic challenge for natural communities: the environmental rate of change exceeds the population’s adaptation capability. Integrating both ecological and evolutionary responses is necessary to make reliable predictions regarding the loss of biodiversity. The race against extinction from an eco-evolutionary perspective is gaining importance in ecological risk assessment. Here, we performed a classical study of population dynamics—a fluctuation analysis—and evaluated the results from an adaption perspective. Fluctuation analysis, widely used with microorganisms, is an effective empirical procedure to study adaptation under strong selective pressure because it incorporates the factors that influence demographic, genetic and environmental changes. The adaptation of phytoplankton to beryllium (Be is of interest because human activities are increasing the concentration of Be in freshwater reserves; therefore, predicting the effects of human-induced pollutants is necessary for proper risk assessment. The fluctuation analysis was performed with phytoplankton, specifically, the freshwater microalgae Chlamydomonas reinhardtii, under acute Be exposure. High doses of Be led to massive microalgae death; however, by conducting a fluctuation analysis experiment, we found that C. reinhardtii was able to adapt to 33 mg/l of Be due to pre-existing genetic variability. The rescuing adapting genotype presented a mutation rate of 9.61 × 10−6 and a frequency of 10.42 resistant cells per million wild-type cells. The genetic adaptation pathway that was experimentally obtained agreed with the theoretical models of evolutionary rescue (ER. Furthermore, the rescuing genotype presented phenotypic and physiologic differences from the wild-type genotype, was 25% smaller than the Be-resistant genotype and presented a lower fitness and quantum yield performance. The abrupt distinctions between the wild-type and the Be

  14. Rapid induction of lipid droplets in Chlamydomonas reinhardtii and Chlorella vulgaris by Brefeldin A.

    Directory of Open Access Journals (Sweden)

    Sangwoo Kim

    Full Text Available Algal lipids are the focus of intensive research because they are potential sources of biodiesel. However, most algae produce neutral lipids only under stress conditions. Here, we report that treatment with Brefeldin A (BFA, a chemical inducer of ER stress, rapidly triggers lipid droplet (LD formation in two different microalgal species, Chlamydomonas reinhardtii and Chlorella vulgaris. LD staining using Nile red revealed that BFA-treated algal cells exhibited many more fluorescent bodies than control cells. Lipid analyses based on thin layer chromatography and gas chromatography revealed that the additional lipids formed upon BFA treatment were mainly triacylglycerols (TAGs. The increase in TAG accumulation was accompanied by a decrease in the betaine lipid diacylglyceryl N,N,N-trimethylhomoserine (DGTS, a major component of the extraplastidic membrane lipids in Chlamydomonas, suggesting that at least some of the TAGs were assembled from the degradation products of membrane lipids. Interestingly, BFA induced TAG accumulation in the Chlamydomonas cells regardless of the presence or absence of an acetate or nitrogen source in the medium. This effect of BFA in Chlamydomonas cells seems to be due to BFA-induced ER stress, as supported by the induction of three homologs of ER stress marker genes by the drug. Together, these results suggest that ER stress rapidly triggers TAG accumulation in two green microalgae, C. reinhardtii and C. vulgaris. A further investigation of the link between ER stress and TAG synthesis may yield an efficient means of producing biofuel from algae.

  15. Effect of O2:CO2 Ratio on the Primary Metabolism of Chlamydomonas reinhardtii

    NARCIS (Netherlands)

    Kliphuis, A.M.J.; Martens, D.E.; Janssen, M.G.J.; Wijffels, R.H.

    2011-01-01

    High oxygen:carbon dioxide ratios may have a negative effect on growth and productivity of microalgae. To investigate the effect of O2 and CO2 concentrations and the ratio between these on the metabolism of Chlamydomonas reinhardtii we performed turbidostat experiments at different O2:CO2 ratios. Th

  16. Phytochelatins do not correlate with the level of Cd accumulation in Chlamydomonas spp.

    Science.gov (United States)

    Nishikawa, Kahoko; Onodera, Ai; Tominaga, Noriko

    2006-06-01

    Chlamydomonas acidophila KT-1 and Chlamydomonas acidophila DVB238 exhibit a strong heavy metal tolerance, but C. acidophila DVB238 can accumulate a much higher amount of Cadmium (Cd) than C. acidophila KT-1. Phytochelatins (PCs) are known to play an important role in the detoxification of several toxic heavy metals, but the relationship between PCs and Cd accumulation is not clear. PC metabolism and Cd accumulation were investigated by using three Chlamydomonas strains including Chlamydomonas reinhardtii C-9 as a standard alga. The results showed that the PC content did not correlate closely with the level of Cd accumulation, maintenance of a high GSH level seeming to be more important for Cd accumulation. The ultrastructure of C. acidophila KT-1 was extremely disrupted by a great increase in starch granules, which resulted in a moribund state, but hyper-accumulator C. acidophila DVB238 did not exhibit an increase in starch granules in its cells, in spite of Cd accumulation in its chloroplasts, cytosol and vacuoles. These results indicated that C. acidophila DVB238 probably has a developed detoxification system preventing such as destruction of the cells due to Cd toxicity.

  17. Mastoparan-induced programmed cell death in the unicellular alga Chlamydomonas reinhardtti

    NARCIS (Netherlands)

    Yordanova, Z.P.; Woltering, E.J.; Kapchina-Toteva, V.M.; Iakimova, E.T.

    2013-01-01

    The present study was focused on the elucidation of stress-induced cell death signaling events in the unicellular alga Chlamydomonas reinhardtii exposed to treatment with wasp venom mastoparan. By applying pharmacological approach with specific inhibitors, we have investigated the involvement of eth

  18. An automated GCxGC-TOF-MS protocol for batch-wise extraction and alignment of mass isotopomer matrixes from differential 13C-labelling experiments: a case study for photoautotrophic-mixotrophic grown Chlamydomonas reinhardtii cells.

    Science.gov (United States)

    Kempa, Stefan; Hummel, Jan; Schwemmer, Thorsten; Pietzke, Matthias; Strehmel, Nadine; Wienkoop, Stefanie; Kopka, Joachim; Weckwerth, Wolfram

    2009-02-01

    Two dimensional gas chromatography coupled to time-of-flight mass spectrometry (GCxGC-TOF-MS) is a promising technique to overcome limits of complex metabolome analysis using one dimensional GC-TOF-MS. Especially at the stage of data export and data mining, however, convenient procedures to cope with the complexity of GCxGC-TOF-MS data are still in development. Here, we present a high sample throughput protocol exploiting first and second retention index for spectral library search and subsequent construction of a high dimensional data matrix useful for statistical analysis. The method was applied to the analysis of (13)C-labelling experiments in the unicellular green alga Chlamydomonas reinhardtii. We developed a rapid sampling and extraction procedure for Chlamydomonas reinhardtii laboratory strain (CC503), a cell wall deficient mutant. By testing all published quenching protocols we observed dramatic metabolite leakage rates for certain metabolites. To circumvent metabolite leakage, samples were directly quenched and analyzed without separation of the medium. The growth medium was adapted to this rapid sampling protocol to avoid interference with GCxGC-TOF-MS analysis. To analyse batches of samples a new software tool, MetMax, was implemented which extracts the isotopomer matrix from stable isotope labelling experiments together with the first and second retention index (RI1 and RI2). To exploit RI1 and RI2 for metabolite identification we used the Golm metabolome database (GMD [1] with RI1/RI2-reference spectra and new search algorithms. Using those techniques we analysed the dynamics of (13)CO(2) and (13)C-acetate uptake in Chlamydomonas reinhardtii cells in two different steady states namely photoautotroph and mixotroph growth conditions.

  19. Forward genetic screen for auxin-deficient mutants by cytokinin.

    Science.gov (United States)

    Wu, Lei; Luo, Pan; Di, Dong-Wei; Wang, Li; Wang, Ming; Lu, Cheng-Kai; Wei, Shao-Dong; Zhang, Li; Zhang, Tian-Zi; Amakorová, Petra; Strnad, Miroslav; Novák, Ondřej; Guo, Guang-Qin

    2015-07-06

    Identification of mutants with impairments in auxin biosynthesis and dynamics by forward genetic screening is hindered by the complexity, redundancy and necessity of the pathways involved. Furthermore, although a few auxin-deficient mutants have been recently identified by screening for altered responses to shade, ethylene, N-1-naphthylphthalamic acid (NPA) or cytokinin (CK), there is still a lack of robust markers for systematically isolating such mutants. We hypothesized that a potentially suitable phenotypic marker is root curling induced by CK, as observed in the auxin biosynthesis mutant CK-induced root curling 1 / tryptophan aminotransferase of Arabidopsis 1 (ckrc1/taa1). Phenotypic observations, genetic analyses and biochemical complementation tests of Arabidopsis seedlings displaying the trait in large-scale genetic screens showed that it can facilitate isolation of mutants with perturbations in auxin biosynthesis, transport and signaling. However, unlike transport/signaling mutants, the curled (or wavy) root phenotypes of auxin-deficient mutants were significantly induced by CKs and could be rescued by exogenous auxins. Mutants allelic to several known auxin biosynthesis mutants were re-isolated, but several new classes of auxin-deficient mutants were also isolated. The findings show that CK-induced root curling provides an effective marker for discovering genes involved in auxin biosynthesis or homeostasis.

  20. Handbook of display technology

    CERN Document Server

    Castellano, Joseph A

    1992-01-01

    This book presents a comprehensive review of technical and commercial aspects of display technology. It provides design engineers with the information needed to select proper technology for new products. The book focuses on flat, thin displays such as light-emitting diodes, plasma display panels, and liquid crystal displays, but it also includes material on cathode ray tubes. Displays include a large number of products from televisions, auto dashboards, radios, and household appliances, to gasoline pumps, heart monitors, microwave ovens, and more.For more information on display tech

  1. COPPER RESPONSE REGULATOR1–Dependent and –Independent Responses of the Chlamydomonas reinhardtii Transcriptome to Dark Anoxia[W

    Science.gov (United States)

    Hemschemeier, Anja; Casero, David; Liu, Bensheng; Benning, Christoph; Pellegrini, Matteo; Happe, Thomas; Merchant, Sabeeha S.

    2013-01-01

    Anaerobiosis is a stress condition for aerobic organisms and requires extensive acclimation responses. We used RNA-Seq for a whole-genome view of the acclimation of Chlamydomonas reinhardtii to anoxic conditions imposed simultaneously with transfer to the dark. Nearly 1.4 × 103 genes were affected by hypoxia. Comparing transcript profiles from early (hypoxic) with those from late (anoxic) time points indicated that cells activate oxidative energy generation pathways before employing fermentation. Probable substrates include amino acids and fatty acids (FAs). Lipid profiling of the C. reinhardtii cells revealed that they degraded FAs but also accumulated triacylglycerols (TAGs). In contrast with N-deprived cells, the TAGs in hypoxic cells were enriched in desaturated FAs, suggesting a distinct pathway for TAG accumulation. To distinguish transcriptional responses dependent on COPPER RESPONSE REGULATOR1 (CRR1), which is also involved in hypoxic gene regulation, we compared the transcriptomes of crr1 mutants and complemented strains. In crr1 mutants, ∼40 genes were aberrantly regulated, reaffirming the importance of CRR1 for the hypoxic response, but indicating also the contribution of additional signaling strategies to account for the remaining differentially regulated transcripts. Based on transcript patterns and previous results, we conclude that nitric oxide–dependent signaling cascades operate in anoxic C. reinhardtii cells. PMID:24014546

  2. A Compressive Superresolution Display

    KAUST Repository

    Heide, Felix

    2014-06-22

    In this paper, we introduce a new compressive display architecture for superresolution image presentation that exploits co-design of the optical device configuration and compressive computation. Our display allows for superresolution, HDR, or glasses-free 3D presentation.

  3. Lunar Sample Display Locations

    Data.gov (United States)

    National Aeronautics and Space Administration — NASA provides a number of lunar samples for display at museums, planetariums, and scientific expositions around the world. Lunar displays are open to the public....

  4. plenty, a novel hypernodulation mutant in Lotus japonicus.

    Science.gov (United States)

    Yoshida, Chie; Funayama-Noguchi, Sachiko; Kawaguchi, Masayoshi

    2010-09-01

    Nitrogen fixation in nodules that contain symbiotic rhizobial bacteria enables legumes to thrive in nitrogen-poor soils. However, this symbiosis is energy consuming. Therefore, legumes strictly control nodulation at both local and systemic levels. Mutants deficient in such controls exhibit a range of phenotypes from non-nodulation to hypernodulation. Here, we isolated a novel hypernodulation mutant from the M(2) progeny derived from Lotus japonicus MG-20 seeds mutagenized by irradiation with a carbon ion beam. We named the mutant 'plenty' because it formed more nodules than the wild-type MG-20. The nodulation zone in the plenty mutant was wider than that in the wild type, but not as enhanced as those in other previously reported hypernodulation mutants such as har1, klv or tml of L. japonicus. Unlike these hypernodulation mutants, the plenty mutant developed nodules of the same size as MG-20. Overall, the plenty mutant exhibited a unique phenotype of moderate hypernodulation. However, a biomass assay indicated that this unique pattern of hypernodulation was a hindrance to host plant growth. The plenty mutant displayed some tolerance to external nitrates and a normal triple response to ethylene. Grafting experiments demonstrated that the root of plenty was responsible for its hypernodulation phenotype. Genetic mapping indicated that the PLENTY gene was located on chromosome 2.

  5. Invisible Display in Aluminum

    DEFF Research Database (Denmark)

    Prichystal, Jan Phuklin; Hansen, Hans Nørgaard; Bladt, Henrik Henriksen

    2005-01-01

    Bang & Olufsen a/s has been working with ideas for invisible integration of displays in metal surfaces. Invisible integration of information displays traditionally has been possible by placing displays behind transparent or semitransparent materials such as plastic or glass. The wish for an integ...... be obtained by shining light from the backside of the workpiece. When there is no light from the backside, the front surface seems totally untouched. This was achieved by laser ablation with ultra-short pulses.......Bang & Olufsen a/s has been working with ideas for invisible integration of displays in metal surfaces. Invisible integration of information displays traditionally has been possible by placing displays behind transparent or semitransparent materials such as plastic or glass. The wish...... for an integrated display in a metal surface is often ruled by design and functionality of a product. The integration of displays in metal surfaces requires metal removal in order to clear the area of the display to some extent. The idea behind an invisible display in Aluminum concerns the processing of a metal...

  6. The energy balance of the biomass generation of Chlamydomonas acidophila under acidic and neutral conditions and Chlamydomonas reinhardtii; Die Energiebilanz der Biomasseneubildung von Chlamydomonas acidophila unter sauren und neutralen Bedingungen und von Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Langner, Uwe

    2009-01-16

    In this study the influence of pH < 3 as an extreme environment has been investigated for the eukaryotic green alga Chlamydomonas (C.) acidophila. The limited number of trophic levels, consisting of bacteria, phytoplankton, zooplankton and macrophytes, is a special characteristic of extreme acidic water bodies. C. acidophila was isolated from an extreme acidic mining lake (RL 111) (Bissinger et al. 2000). A special feature of the examined algal species is its wide tolerance range of external pH values from 2 to 7 (Cassin 1874, Gerloff-Elias et al. 2005a). C. acidophila is a dominant species in the acidic mining lakes, it can grow up to chlorophyll maxima of 500 {mu}g L{sup -1} during the summer time (Nixdorf et al. 1998, 2003). The alga can be found elsewhere in extreme acidic water bodies around the world. The hydrochemistry of the acidic mining lakes in the central regions of Germany and Lusatia show clear differences compared to neutral water bodies. Some of the characteristics of acidic mining lakes are high metal and heavy metal (aluminum) concentrations, high ion concentrations, which lead to high conductivity, as well as low phosphate concentrations, ammonium as the only nitrogen source and CO{sub 2} as the only inorganic carbon source. Many eukaryotic microalgae in acidic water bodies including C. acidophila show a neutral cytosolic pH. This is provided by special adaptations of the organisms. Thus, organisms in extreme acidic environments have a positive cell surface charge, a very efficient H{sup +}-ATPase and high internal buffer capacities. These mechanisms work contrary to the proton influx and the acidification of the cytosol and are therefore proof for the physiological impact of osmoregulation by microalgae in extreme acidic environments (Sekler et al.1991, Weiss et al. 1999). Hence, these mechanisms also complicate the access to nutrients for the algal cell. The passive influx of positively charged ions such as potassium or ammonium is reduced by

  7. Truncated photosystem chlorophyll antenna size in the green microalga Chlamydomonas reinhardtii upon deletion of the TLA3-CpSRP43 gene.

    Science.gov (United States)

    Kirst, Henning; Garcia-Cerdan, Jose Gines; Zurbriggen, Andreas; Ruehle, Thilo; Melis, Anastasios

    2012-12-01

    The truncated light-harvesting antenna size3 (tla3) DNA insertional transformant of Chlamydomonas reinhardtii is a chlorophyll-deficient mutant with a lighter green phenotype, a lower chlorophyll (Chl) per cell content, and higher Chl a/b ratio than corresponding wild-type strains. Functional analyses revealed a higher intensity for the saturation of photosynthesis and greater light-saturated photosynthetic activity in the tla3 mutant than in the wild type and a Chl antenna size of the photosystems that was only about 40% of that in the wild type. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and western-blot analyses showed that the tla3 strain was deficient in the Chl a/b light-harvesting complex. Molecular and genetic analyses revealed a single plasmid insertion in chromosome 4 of the tla3 nuclear genome, causing deletion of predicted gene g5047 and plasmid insertion within the fourth intron of downstream-predicted gene g5046. Complementation studies defined that gene g5047 alone was necessary and sufficient to rescue the tla3 mutation. Gene g5047 encodes a C. reinhardtii homolog of the chloroplast-localized SRP43 signal recognition particle, whose occurrence and function in green microalgae has not hitherto been investigated. Biochemical analysis showed that the nucleus-encoded and chloroplast-localized CrCpSRP43 protein specifically operates in the assembly of the peripheral components of the Chl a/b light-harvesting antenna. This work demonstrates that cpsrp43 deletion in green microalgae can be employed to generate tla mutants with a substantially diminished Chl antenna size. The latter exhibit improved solar energy conversion efficiency and photosynthetic productivity under mass culture and bright sunlight conditions.

  8. Assembly of the light-harvesting chlorophyll antenna in the green alga Chlamydomonas reinhardtii requires expression of the TLA2-CpFTSY gene.

    Science.gov (United States)

    Kirst, Henning; García-Cerdán, Jose Gines; Zurbriggen, Andreas; Melis, Anastasios

    2012-02-01

    The truncated light-harvesting antenna2 (tla2) mutant of Chlamydomonas reinhardtii showed a lighter-green phenotype, had a lower chlorophyll (Chl) per-cell content, and higher Chl a/b ratio than corresponding wild-type strains. Physiological analyses revealed a higher intensity for the saturation of photosynthesis and greater P(max) values in the tla2 mutant than in the wild type. Biochemical analyses showed that the tla2 strain was deficient in the Chl a-b light-harvesting complex, and had a Chl antenna size of the photosystems that was only about 65% of that in the wild type. Molecular and genetic analyses showed a single plasmid insertion in the tla2 strain, causing a chromosomal DNA rearrangement and deletion/disruption of five nuclear genes. The TLA2 gene, causing the tla2 phenotype, was cloned by mapping the insertion site and upon complementation with each of the genes that were deleted. Successful complementation was achieved with the C. reinhardtii TLA2-CpFTSY gene, whose occurrence and function in green microalgae has not hitherto been investigated. Functional analysis showed that the nuclear-encoded and chloroplast-localized CrCpFTSY protein specifically operates in the assembly of the peripheral components of the Chl a-b light-harvesting antenna. In higher plants, a cpftsy null mutation inhibits assembly of both the light-harvesting complex and photosystem complexes, thus resulting in a seedling-lethal phenotype. The work shows that cpftsy deletion in green algae, but not in higher plants, can be employed to generate tla mutants. The latter exhibit improved solar energy conversion efficiency and photosynthetic productivity under mass culture and bright sunlight conditions.

  9. Polyplanar optic display

    Energy Technology Data Exchange (ETDEWEB)

    Veligdan, J.; Biscardi, C.; Brewster, C.; DeSanto, L. [Brookhaven National Lab., Upton, NY (United States). Dept. of Advanced Technology; Beiser, L. [Leo Beiser Inc., Flushing, NY (United States)

    1997-07-01

    The Polyplanar Optical Display (POD) is a unique display screen which can be used with any projection source. This display screen is 2 inches thick and has a matte black face which allows for high contrast images. The prototype being developed is a form, fit and functional replacement display for the B-52 aircraft which uses a monochrome ten-inch display. The new display uses a 100 milliwatt green solid state laser (532 nm) as its optical source. In order to produce real-time video, the laser light is being modulated by a Digital Light Processing (DLP{trademark}) chip manufactured by Texas Instruments, Inc. A variable astigmatic focusing system is used to produce a stigmatic image on the viewing face of the POD. In addition to the optical design, the authors discuss the electronic interfacing to the DLP{trademark} chip, the opto-mechanical design and viewing angle characteristics.

  10. OLED displays and lighting

    CERN Document Server

    Koden, Mitsuhiro

    2017-01-01

    Organic light-emitting diodes (OLEDs) have emerged as the leading technology for the new display and lighting market. OLEDs are solid-state devices composed of thin films of organic molecules that create light with the application of electricity. OLEDs can provide brighter, crisper displays on electronic devices and use less power than conventional light-emitting diodes (LEDs) or liquid crystal displays (LCDs) used today. This book covers both the fundamentals and practical applications of flat and flexible OLEDs.

  11. Scalable Resolution Display Walls

    KAUST Repository

    Leigh, Jason

    2013-01-01

    This article will describe the progress since 2000 on research and development in 2-D and 3-D scalable resolution display walls that are built from tiling individual lower resolution flat panel displays. The article will describe approaches and trends in display hardware construction, middleware architecture, and user-interaction design. The article will also highlight examples of use cases and the benefits the technology has brought to their respective disciplines. © 1963-2012 IEEE.

  12. Expression and molecular analysis of phbB gene in Chlamydomonas reinhardtii

    Institute of Scientific and Technical Information of China (English)

    WANG Chaogang; HU Zhangli; HU Wei; LEI Anping

    2004-01-01

    The expression vector containing phbB and ble genes was constructed and transformed into cell-walldeficient strain Chlamydomonas reinhardtii CC-849 by the glass-bead method. The transgenic alga was selected and maintained in the TAP agar plates containing 10 μg/mL Zeomycin. Transgenic alga, which could express phbB at the transcriptional level, was obtained and further confirmed with PCR, Southern blot and RT-PCR-DNA hybridization analysis.

  13. JAVA Stereo Display Toolkit

    Science.gov (United States)

    Edmonds, Karina

    2008-01-01

    This toolkit provides a common interface for displaying graphical user interface (GUI) components in stereo using either specialized stereo display hardware (e.g., liquid crystal shutter or polarized glasses) or anaglyph display (red/blue glasses) on standard workstation displays. An application using this toolkit will work without modification in either environment, allowing stereo software to reach a wider audience without sacrificing high-quality display on dedicated hardware. The toolkit is written in Java for use with the Swing GUI Toolkit and has cross-platform compatibility. It hooks into the graphics system, allowing any standard Swing component to be displayed in stereo. It uses the OpenGL graphics library to control the stereo hardware and to perform the rendering. It also supports anaglyph and special stereo hardware using the same API (application-program interface), and has the ability to simulate color stereo in anaglyph mode by combining the red band of the left image with the green/blue bands of the right image. This is a low-level toolkit that accomplishes simply the display of components (including the JadeDisplay image display component). It does not include higher-level functions such as disparity adjustment, 3D cursor, or overlays all of which can be built using this toolkit.

  14. Real-time monitoring of genetically modified Chlamydomonas reinhardtii during the Foton M3 space mission and ground irradiation experiment

    Science.gov (United States)

    Lambreva, Maya; Rea, Giuseppina; Antonacci, Amina; Serafini, Agnese; Damasso, Mario; Margonelli, Andrea; Johanningmeier, Udo; Bertalan, Ivo; Pezzotti, Gianni; Giardi, Maria Teresa

    Long-term space exploration, colonization or habitation requires biological life support systems capable to cope with the deleterious space environment. The use of oxygenic photosynthetic microrganisms is an intriguing possibility mainly for food, O2 and nutraceutical compounds production. The critical points of utilizing plantsor algae-based life support systems are the microgravity and the ionizing radiation, which can influence the performance of these organisms. The aim of the present study was to assess the effects of space environment on the photosynthetic activity of various microrganisms and to select space stress-tolerant strains. Site-directed and random mutants of the unicellular green alga Chlamydomonas reinhardtii of Photosystem II D1 protein were used as a model system to test and select the amino acid substitutions capable to account for space stress tolerance. We focussed our studies also on the accumulation of the Photosystem II photoprotective carotenoids (the xantophylls violaxanthin, anteraxanthin and zeaxanthin), powerful antioxidants that epidemiological studies demonstrated to be human vision protectors. Metabolite profiling by quantitative HPLC methods revealed the organisms and the stress conditions capable to accumulate the highest pigment levels. In order to develop a project for a rationale metabolic engineering of algal secondary metabolites overproduction, we are performing expression analyses on the carotenoid biosynthetic pathway under physiological and mimicked space conditions. To identify the consequences of the space environment on the photosynthetic apparatus the changes in the Photosystem II efficiency were monitored in real time during the ESA-Russian Foton-M3 mission in September 2007. For the space flight a high-tech, multicell fluorescence biosensor, Photo-II, was designed and built by the Centre for Advanced Research in Space Optics in collaboration with Kayser-Italy, Biosensor and DAS. Photo-II is an automatic device

  15. Survival of Human Metallothionein-2 Transplastomic Chlamydomonas reinhardtii to Ultraviolet B Exposure

    Institute of Scientific and Technical Information of China (English)

    Yu-Kun ZHANG; Gui-Fang SHEN; Bing-Gen RU

    2006-01-01

    Solar ultraviolet (UV) radiation has a great influence on green organisms, especially plankton like Chlamydomonas. A human metallothionein-2 gene, which is generally considered to have an anti-radiation function by its coding product, was transferred into the chloroplast genome of Chlamydomonas reinhardtii.To dynamically measure the UV effects on Chlamydomonas cells grown in liquid tris-acetate-phosphate medium, a new method was developed based on the relationship between the chlorophyll content of an algal culture and its absorbance at 570 nm after the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. In this experiment, both the wild-type and the transplastomic C. reinhardtii cells were cultivated in 96-well microplates containing liquid tris-acetate-phosphate medium in the absence or presence of zinc, copper,cadmium and cysteine. The transgenic C. reinhardtii showed a higher resistance than wild-type to UV-B exposure under all the examined conditions. Metals in the medium had positive impacts on both types of cells,but had significant influence only on the transplastomic cells. However, the high cell viability of the transgenic alga at the end of the 8 h UV-B treatment disappeared after a 20-h recovery culture. Cysteine did not protect cells from UV-B damage, but clearly enhanced the growth of both wild-type and transgenic C.reinhardtii.

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

    DEFF Research Database (Denmark)

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

    2016-01-01

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

  17. Connexin mutants and cataracts

    Directory of Open Access Journals (Sweden)

    Eric C Beyer

    2013-04-01

    Full Text Available The lens is a multicellular, but avascular tissue that must stay transparent to allow normal transmission of light and focusing of it on the retina. Damage to lens cells and/or proteins can cause cataracts, opacities that disrupt these processes. The normal survival of the lens is facilitated by an extensive network of gap junctions formed predominantly of connexin46 and connexin50. Mutations of the genes that encode these connexins (GJA3 and GJA8 have been identified and linked to inheritance of cataracts in human families and mouse lines. In vitro expression studies of several of these mutants have shown that they exhibit abnormalities that may lead to disease. Many of the mutants reduce or modify intercellular communication due to channel alterations (including loss of function or altered gating or due to impaired cellular trafficking which reduces the number of gap junction channels within the plasma membrane. However, the abnormalities detected in studies of other mutants suggest that they cause cataracts through other mechanisms including gain of hemichannel function (leading to cell injury and death and formation of cytoplasmic accumulations (that may act as light scattering particles. These observations and the anticipated results of ongoing studies should elucidate the mechanisms of cataract development due to mutations of lens connexins and abnormalities of other lens proteins. They may also contribute to our understanding of the mechanisms of disease due to connexin mutations in other tissues.

  18. Helmet-Mounted Displays (HMD)

    Data.gov (United States)

    Federal Laboratory Consortium — The Helmet-Mounted Display labis responsible for monocular HMD day display evaluations; monocular HMD night vision performance processes; binocular HMD day display...

  19. Standardizing visual display quality

    NARCIS (Netherlands)

    Besuijen, Ko; Spenkelink, Gerd P.J.

    1998-01-01

    The current ISO 9241–3 standard for visual display quality and the proposed user performance tests are reviewed. The standard is found to be more engineering than ergonomic and problems with system configuration, software applications, display settings, user behaviour, wear and physical environment

  20. Polyplanar optical display electronics

    Energy Technology Data Exchange (ETDEWEB)

    DeSanto, L.; Biscardi, C. [Brookhaven National Lab., Upton, NY (United States). Dept. of Advanced Technology

    1997-07-01

    The Polyplanar Optical Display (POD) is a unique display screen which can be used with any projection source. The prototype ten inch display is two inches thick and has a matte black face which allows for high contrast images. The prototype being developed is a form, fit and functional replacement display for the B-52 aircraft which uses a monochrome ten-inch display. In order to achieve a long lifetime, the new display uses a 100 milliwatt green solid-state laser (10,000 hr. life) at 532 nm as its light source. To produce real-time video, the laser light is being modulated by a Digital Light Processing (DLP{trademark}) chip manufactured by Texas Instruments. In order to use the solid-state laser as the light source and also fit within the constraints of the B-52 display, the Digital Micromirror Device (DMD{trademark}) circuit board is removed from the Texas Instruments DLP light engine assembly. Due to the compact architecture of the projection system within the display chassis, the DMD{trademark} chip is operated remotely from the Texas Instruments circuit board. The authors discuss the operation of the DMD{trademark} divorced from the light engine and the interfacing of the DMD{trademark} board with various video formats (CVBS, Y/C or S-video and RGB) including the format specific to the B-52 aircraft. A brief discussion of the electronics required to drive the laser is also presented.

  1. Visual merchandising window display

    Directory of Open Access Journals (Sweden)

    Opris (Cas. Stanila M.

    2013-12-01

    Full Text Available Window display plays a major part in the selling strategies; it does not only include the simple display of goods, nowadays it is a form of art, also having the purpose of sustaining the brand image. This article wants to reveal the tools that are essential in creating a fabulous window display. Being a window designer is not an easy job, you have to always think ahead trends, to have a sense of colour, to know how to use light to attract customers in the store after only one glance at the window. The big store window displays are theatre scenes: with expensive backgrounds, special effects and high fashion mannequins. The final role of the displays is to convince customers to enter the store and trigger the purchasing act which is the final goal of the retail activity.

  2. Multiple stressor effects in Chlamydomonas reinhardtii – Toward understanding mechanisms of interaction between effects of ultraviolet radiation and chemical pollutants

    Energy Technology Data Exchange (ETDEWEB)

    Korkaric, Muris [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Department of Environmental Toxicology, 8600, Duebendorf (Switzerland); ETH Zürich, Institute of Biogeochemistry and Pollutant Dynamics, 8092 Zürich (Switzerland); Behra, Renata; Fischer, Beat B. [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Department of Environmental Toxicology, 8600, Duebendorf (Switzerland); Junghans, Marion [Swiss Center for Applied Ecotoxicology Eawag-EPFL, 8600, Duebendorf (Switzerland); Eggen, Rik I.L., E-mail: rik.eggen@eawag.ch [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Department of Environmental Toxicology, 8600, Duebendorf (Switzerland); ETH Zürich, Institute of Biogeochemistry and Pollutant Dynamics, 8092 Zürich (Switzerland)

    2015-05-15

    Highlights: • Systematic study of multiple stressor effects of UVR and chemicals in C. reinhardtii. • UVR and chemicals did not act independently on algal photosynthesis and reproduction. • Multiple stressor effects of UVR and chemicals depended on chemical MOA. • Synergistic effect interactions not limited to oxidative stress inducing chemicals. • Multiple MOAs of UVR may limit applicability of current prediction models. - Abstract: The effects of chemical pollutants and environmental stressors, such as ultraviolet radiation (UVR), can interact when organisms are simultaneously exposed, resulting in higher (synergistic) or lower (antagonistic) multiple stressor effects than expected based on the effects of single stressors. Current understanding of interactive effects is limited due to a lack of mechanism-based multiple stressor studies. It has been hypothesized that effect interactions may generally occur if chemical and non-chemical stressors cause similar physiological effects in the organism. To test this hypothesis, we exposed the model green alga Chlamydomonas reinhardtii to combinations of UVR and single chemicals displaying modes of action (MOA) similar or dissimilar to the impact of UVR on photosynthesis. Stressor interactions were analyzed based on the independent action model. Effect interactions were found to depend on the MOA of the chemicals, and also on their concentrations, the exposure time and the measured endpoint. Indeed, only chemicals assumed to cause effects on photosynthesis similar to UVR showed interactions with UVR on photosynthetic yield: synergistic in case of Cd(II) and paraquat and antagonistic in case of diuron. No interaction on photosynthesis was observed for S-metolachlor, which acts dissimilarly to UVR. However, combined effects of S-metolachlor and UVR on algal reproduction were synergistic, highlighting the importance of considering additional MOA of UVR. Possible mechanisms of stressor effect interactions are

  3. An improved ARS2-derived nuclear reporter enhances the efficiency and ease of genetic engineering in Chlamydomonas

    DEFF Research Database (Denmark)

    Specht, Elizabeth A; Nour-Eldin, Hussam Hassan; Hoang, Kevin T D

    2015-01-01

    The model alga Chlamydomonas reinhardtii has been used to pioneer genetic engineering techniques for high-value protein and biofuel production from algae. To date, most studies of transgenic Chlamydomonas have utilized the chloroplast genome due to its ease of engineering, with a sizeable suite o...... nuclear promoters. This enhanced arylsulfatase reporter construct improves the efficiency and ease of genetic engineering within the Chlamydomonas nuclear genome, with potential application to other algal strains.......The model alga Chlamydomonas reinhardtii has been used to pioneer genetic engineering techniques for high-value protein and biofuel production from algae. To date, most studies of transgenic Chlamydomonas have utilized the chloroplast genome due to its ease of engineering, with a sizeable suite...... of reporters and well-characterized expression constructs. The advanced manipulation of algal nuclear genomes has been hampered by limited strong expression cassettes, and a lack of high-throughput reporters. We have improved upon an endogenous reporter gene - the ARS2 gene encoding an arylsulfatase enzyme...

  4. Panoramic projection avionics displays

    Science.gov (United States)

    Kalmanash, Michael H.

    2003-09-01

    Avionics projection displays are entering production in advanced tactical aircraft. Early adopters of this technology in the avionics community used projection displays to replace or upgrade earlier units incorporating direct-view CRT or AMLCD devices. Typical motivation for these upgrades were the alleviation of performance, cost and display device availability concerns. In these systems, the upgraded (projection) displays were one-for-one form / fit replacements for the earlier units. As projection technology has matured, this situation has begun to evolve. The Lockheed-Martin F-35 is the first program in which the cockpit has been specifically designed to take advantage of one of the more unique capabilities of rear projection display technology, namely the ability to replace multiple small screens with a single large conformal viewing surface in the form of a panoramic display. Other programs are expected to follow, since the panoramic formats enable increased mission effectiveness, reduced cost and greater information transfer to the pilot. Some of the advantages and technical challenges associated with panoramic projection displays for avionics applications are described below.

  5. tla1, a DNA insertional transformant of the green alga Chlamydomonas reinhardtii with a truncated light-harvesting chlorophyll antenna size.

    Science.gov (United States)

    Polle, Juergen E W; Kanakagiri, Sarada-Devi; Melis, Anastasios

    2003-05-01

    DNA insertional mutagenesis and screening of the green alga Chlamydomonas reinhardtii was employed to isolate tla1, a stable transformant having a truncated light-harvesting chlorophyll antenna size. Molecular analysis showed a single plasmid insertion into an open reading frame of the nuclear genome corresponding to a novel gene ( Tla1) that encodes a protein of 213 amino acids. Genetic analysis showed co-segregation of plasmid and tla1 phenotype. Biochemical analyses showed the tla1 mutant to be chlorophyll deficient, with a functional chlorophyll antenna size of photosystem I and photosystem II being about 50% and 65% of that of the wild type, respectively. It contained a correspondingly lower amount of light-harvesting proteins than the wild type and had lower steady-state levels of Lhcb mRNA. The tla1 strain required a higher light intensity for the saturation of photosynthesis and showed greater solar conversion efficiencies and a higher photosynthetic productivity than the wild type under mass culture conditions. Results are discussed in terms of the tla1 mutation, its phenotype, and the role played by the Tla1 gene in the regulation of the photosynthetic chlorophyll antenna size in C. reinhardtii.

  6. NRT2.4 and NRT2.5 Are Two Half-Size Transporters from the Chlamydomonas NRT2 Family

    Directory of Open Access Journals (Sweden)

    Jose Javier Higuera

    2016-03-01

    Full Text Available The NRT2 transporters mediate High Affinity Nitrate/NitriteTransport (HAN/NiT, which are essential for nitrogen acquisition from these inorganic forms. The NRT2 proteins are encoded by a multigene family in plants, and contain 12 transmembrane-spanning domains. Chlamydomonas reinhardtii has six NRT2, two of which (NRT2.5 and NRT2.4 are located in Chromosome III, in tandem head to tail. cDNAs for these genes were isolated and their sequence revealed that they correspond to half-size NRT2 transporters each containing six transmembrane domains. NRT2.5 has long N- and C- termini sequences without known homology. NRT2.4 also contains long termini sequences but smaller than NRT2.5. Expression of both studied genes occurred at a very low level, slightly in darkness, and was not modified by the N or C source. Silencing of NRT2.4 by specific artificial miRNA resulted in the inhibition of nitrite transport in the absence of other HANNiT (NRT2.1/NAR2 in the cell genetic background. Nitrite transport activity in the Hansenula polymorpha Δynt::URA3 Leu2 mutant was restored by expressing CrNRT2.4. These results indicate that half-size NRT2 transporters are present in photosynthetic organisms and that NRT2.4 is a HANiT.

  7. Small - Display Cartography

    DEFF Research Database (Denmark)

    Nissen, Flemming; Hvas, Anders; Münster-Swendsen, Jørgen

    This report comprises the work carried out in the work-package of small display cartography. The work-package has aimed at creating a general framework for the small-display cartography. A solid framework facilitates an increased use of spatial data in mobile devices - thus enabling, together...... with the rapidly evolving positioning techniques, a new category of position-dependent, map-based services to be introduced. The report consists of the following parts: Part I: Categorization of handheld devices, Part II: Cartographic design for small-display devices, Part III: Study on the GiMoDig Client ? Portal...... Appendix E consisting of confidential material and Appendix F with examples.   The work has resulted in a small device categorisation, a cartographic design specification for a small-display device for a specified navigation task as well as a comparison between OpenLS and an in house developed protocol...

  8. Flexible displays, rigid designs?

    DEFF Research Database (Denmark)

    Hornbæk, Kasper

    2015-01-01

    Rapid technological progress has enabled a wide range of flexible displays for computing devices, but the user experience--which we're only beginning to understand--will be the key driver for successful designs....

  9. Military display performance parameters

    Science.gov (United States)

    Desjardins, Daniel D.; Meyer, Frederick

    2012-06-01

    The military display market is analyzed in terms of four of its segments: avionics, vetronics, dismounted soldier, and command and control. Requirements are summarized for a number of technology-driving parameters, to include luminance, night vision imaging system compatibility, gray levels, resolution, dimming range, viewing angle, video capability, altitude, temperature, shock and vibration, etc., for direct-view and virtual-view displays in cockpits and crew stations. Technical specifications are discussed for selected programs.

  10. Liquid Crystal Airborne Display

    Science.gov (United States)

    1977-08-01

    81/2X 11- 10 -9 .8 display using a large advertising alphanimeric ( TCI ) has been added to the front of the optical box used in the F-4 aircraft for HUD...properties over a wide range of tempera - tures, including normal room temperature. What are Liquid Crystals? Liquid crystals have been classified in three...natic fanctions and to present data needed for the semi- automatic and manual control of system functions. Existing aircraft using CRT display

  11. Raster graphics display library

    Science.gov (United States)

    Grimsrud, Anders; Stephenson, Michael B.

    1987-01-01

    The Raster Graphics Display Library (RGDL) is a high level subroutine package that give the advanced raster graphics display capabilities needed. The RGDL uses FORTRAN source code routines to build subroutines modular enough to use as stand-alone routines in a black box type of environment. Six examples are presented which will teach the use of RGDL in the fastest, most complete way possible. Routines within the display library that are used to produce raster graphics are presented in alphabetical order, each on a separate page. Each user-callable routine is described by function and calling parameters. All common blocks that are used in the display library are listed and the use of each variable within each common block is discussed. A reference on the include files that are necessary to compile the display library is contained. Each include file and its purpose are listed. The link map for MOVIE.BYU version 6, a general purpose computer graphics display system that uses RGDL software, is also contained.

  12. Phenotypic Characterization of a Female Sterile Mutant in Rice

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    A female sterile mutant, derived from a spontaneous mutation, wasfirst discovered in rice (Oryza sativa L. ssp.indica) restorer line 202R. With normal flowering, the mutant exhibits an extremely Iow seed-setting rate. When the mutant is crossed as a pollen donor, the seeds set normally; whereas when it is used as a pollen receiver,no seeds are obtained even with mixed pollen grains of different varieties sprinkled over the stigmas. The floret of the mutant, consisting of six stamens and one pistil, looks the same as that of the wild type in the malefemale organs, except that less than 10% of the mutant florets have three stigmas on the ovary. Although the mutant has a low seed-setting rate, Its pollen fertility is approximately 87.1%, which is equal to that of the wild type. In addition, more than 90% of the mature embryo sacs of the mutant have complete inner structures. At every stage after pollination, the sperm, embryo, and endosperm are not found in the mutant embryo sac,whereas the disintegration of the egg cell that does not accomplish fertilization is visible. Through observations with a fluorescence microscope, we have found that the pollen grains germinate normally, whereas the pollen tube abnormally elongates in the style-transmitting tissue. The mutant pollen tubes display various defects in the style, such as slower elongation, conversed elongation, distorted elongation, swollen tips, or branched tips. As a result, the growth of the pollen tubes ceases in the style, and, therefore, the pollen tubes cannot reach the embryo sac and the process of double fertilization is blocked. Based on these observations,we conclude that this mutant, designated as fs-202R, is a novel type of female sterile mutation in rice, which causes the arrest of the elongation of the pollen tube.

  13. RNAi knock-down of LHCBM1, 2 and 3 increases photosynthetic H2 production efficiency of the green alga Chlamydomonas reinhardtii.

    Directory of Open Access Journals (Sweden)

    Melanie Oey

    Full Text Available Single cell green algae (microalgae are rapidly emerging as a platform for the production of sustainable fuels. Solar-driven H2 production from H2O theoretically provides the highest-efficiency route to fuel production in microalgae. This is because the H2-producing hydrogenase (HYDA is directly coupled to the photosynthetic electron transport chain, thereby eliminating downstream energetic losses associated with the synthesis of carbohydrate and oils (feedstocks for methane, ethanol and oil-based fuels. Here we report the simultaneous knock-down of three light-harvesting complex proteins (LHCMB1, 2 and 3 in the high H2-producing Chlamydomonas reinhardtii mutant Stm6Glc4 using an RNAi triple knock-down strategy. The resultant Stm6Glc4L01 mutant exhibited a light green phenotype, reduced expression of LHCBM1 (20.6% ±0.27%, LHCBM2 (81.2% ±0.037% and LHCBM3 (41.4% ±0.05% compared to 100% control levels, and improved light to H2 (180% and biomass (165% conversion efficiencies. The improved H2 production efficiency was achieved at increased solar flux densities (450 instead of ∼100 µE m(-2 s(-1 and high cell densities which are best suited for microalgae production as light is ideally the limiting factor. Our data suggests that the overall improved photon-to-H2 conversion efficiency is due to: 1 reduced loss of absorbed energy by non-photochemical quenching (fluorescence and heat losses near the photobioreactor surface; 2 improved light distribution in the reactor; 3 reduced photoinhibition; 4 early onset of HYDA expression and 5 reduction of O2-induced inhibition of HYDA. The Stm6Glc4L01 phenotype therefore provides important insights for the development of high-efficiency photobiological H2 production systems.

  14. Engineering the chloroplast targeted malarial vaccine antigens in Chlamydomonas starch granules.

    Directory of Open Access Journals (Sweden)

    David Dauvillée

    Full Text Available BACKGROUND: Malaria, an Anopheles-borne parasitic disease, remains a major global health problem causing illness and death that disproportionately affects developing countries. Despite the incidence of malaria, which remains one of the most severe infections of human populations, there is no licensed vaccine against this life-threatening disease. In this context, we decided to explore the expression of Plasmodium vaccine antigens fused to the granule bound starch synthase (GBSS, the major protein associated to the starch matrix in all starch-accumulating plants and algae such as Chlamydomonas reinhardtii. METHODS AND FINDINGS: We describe the development of genetically engineered starch granules containing plasmodial vaccine candidate antigens produced in the unicellular green algae Chlamydomonas reinhardtii. We show that the C-terminal domains of proteins from the rodent Plasmodium species, Plasmodium berghei Apical Major Antigen AMA1, or Major Surface Protein MSP1 fused to the algal granule bound starch synthase (GBSS are efficiently expressed and bound to the polysaccharide matrix. Mice were either immunized intraperitoneally with the engineered starch particles and Freund adjuvant, or fed with the engineered particles co-delivered with the mucosal adjuvant, and challenged intraperitoneally with a lethal inoculum of P. Berghei. Both experimental strategies led to a significantly reduced parasitemia with an extension of life span including complete cure for intraperitoneal delivery as assessed by negative blood thin smears. In the case of the starch bound P. falciparum GBSS-MSP1 fusion protein, the immune sera or purified immunoglobulin G of mice immunized with the corresponding starch strongly inhibited in vitro the intra-erythrocytic asexual development of the most human deadly plasmodial species. CONCLUSION: This novel system paves the way for the production of clinically relevant plasmodial antigens as algal starch-based particles

  15. Rapid construction and screening of artificial microRNA systems in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Hu, Jinlu; Deng, Xuan; Shao, Ning; Wang, Gaohong; Huang, Kaiyao

    2014-09-01

    The unicellular green algae Chlamydomonas reinhardtii is a classic model for the study of flagella/cilia and photosynthesis, and it has recently been exploited for producing biopharmaceuticals and biofuel. Due to the low frequency of homologous recombination, reverse genetic manipulation in Chlamydomonas relies mainly on miRNA- and siRNA-based knockdown methods. However, the difficulty in constructing artificial miRNA vectors, laborious screening of knockdown transformants, and undesired epigenetic silencing of exogenous miRNA constructs limit their application. We have established a one-step procedure to construct an artificial miRNA precursor by annealing eight oligonucleotides of approximately 40 nucleotides. In the final construct, the Gaussia princeps luciferase gene (G-Luc) is positioned between the promoter and the artificial miRNA precursor so that knockdown strains may quickly be screened by visualizing luciferase luminescence using a photon-counting camera. Furthermore, the luciferase activity of transformants correlates with the knockdown level of two test target proteins: the chloroplast protein VIPP1 (vesicle inducing protein in plastids 1) and the flagellar protein CDPK3 (calcium-dependent protein kinase 3). Adding an intron from RBCS2 (ribulose bisphosphate carboxylase/oxygenase small subunit 2) to the miRNA construct enhanced both the luciferase activity and the miRNA knockdown efficiency. A second miRNA vector incorporated the promoter of the nitrate reductase gene to allow inducible expression of the artificial miRNA. These vectors will facilitate application of the artificial miRNA and provide tools for studying the mechanism of epigenetics in Chlamydomonas, and may also be adapted for use in other model organisms.

  16. Acclimation of Antarctic Chlamydomonas to the sea-ice environment: a transcriptomic analysis.

    Science.gov (United States)

    Liu, Chenlin; Wang, Xiuliang; Wang, Xingna; Sun, Chengjun

    2016-07-01

    The Antarctic green alga Chlamydomonas sp. ICE-L was isolated from sea ice. As a psychrophilic microalga, it can tolerate the environmental stress in the sea-ice brine, such as freezing temperature and high salinity. We performed a transcriptome analysis to identify freezing stress responding genes and explore the extreme environmental acclimation-related strategies. Here, we show that many genes in ICE-L transcriptome that encoding PUFA synthesis enzymes, molecular chaperon proteins, and cell membrane transport proteins have high similarity to the gens from Antarctic bacteria. These ICE-L genes are supposed to be acquired through horizontal gene transfer from its symbiotic microbes in the sea-ice brine. The presence of these genes in both sea-ice microalgae and bacteria indicated the biological processes they involved in are possibly contributing to ICE-L success in sea ice. In addition, the biological pathways were compared between ICE-L and its closely related sister species, Chlamydomonas reinhardtii and Volvox carteri. In ICE-L transcripome, many sequences homologous to the plant or bacteria proteins in the post-transcriptional, post-translational modification, and signal-transduction KEGG pathways, are absent in the nonpsychrophilic green algae. These complex structural components might imply enhanced stress adaptation capacity. At last, differential gene expression analysis at the transcriptome level of ICE-L indicated that genes that associated with post-translational modification, lipid metabolism, and nitrogen metabolism are responding to the freezing treatment. In conclusion, the transcriptome of Chlamydomonas sp. ICE-L is very useful for exploring the mutualistic interaction between microalgae and bacteria in sea ice; and discovering the specific genes and metabolism pathways responding to the freezing acclimation in psychrophilic microalgae.

  17. The Ultimate Display

    CERN Document Server

    Fluke, C J

    2016-01-01

    Astronomical images and datasets are increasingly high-resolution and multi-dimensional. The vast majority of astronomers perform all of their visualisation and analysis tasks on low-resolution, two-dimensional desktop monitors. If there were no technological barriers to designing the ultimate stereoscopic display for astronomy, what would it look like? What capabilities would we require of our compute hardware to drive it? And are existing technologies even close to providing a true 3D experience that is compatible with the depth resolution of human stereoscopic vision? We consider the CAVE2 (an 80 Megapixel, hybrid 2D and 3D virtual reality environment directly integrated with a 100 Tflop/s GPU-powered supercomputer) and the Oculus Rift (a low- cost, head-mounted display) as examples at opposite financial ends of the immersive display spectrum.

  18. Protocol: methodology for chromatin immunoprecipitation (ChIP in Chlamydomonas reinhardtii

    Directory of Open Access Journals (Sweden)

    Strenkert Daniela

    2011-11-01

    Full Text Available Abstract We report on a detailed chromatin immunoprecipitation (ChIP protocol for the unicellular green alga Chlamydomonas reinhardtii. The protocol is suitable for the analysis of nucleosome occupancy, histone modifications and transcription factor binding sites at the level of mononucleosomes for targeted and genome-wide studies. We describe the optimization of conditions for crosslinking, chromatin fragmentation and antibody titer determination and provide recommendations and an example for the normalization of ChIP results as determined by real-time PCR.

  19. Virtual Auditory Displays

    Science.gov (United States)

    2000-01-01

    timbre , intensity, distance, room modeling, radio communication Virtual Environments Handbook Chapter 4 Virtual Auditory Displays Russell D... musical note “A” as a pure sinusoid, there will be 440 condensations and rarefactions per second. The distance between two adjacent condensations or...and complexity are pitch, loudness, and timbre respectively. This distinction between physical and perceptual measures of sound properties is an

  20. Refreshing Refreshable Braille Displays.

    Science.gov (United States)

    Russomanno, Alexander; O'Modhrain, Sile; Gillespie, R Brent; Rodger, Matthew W M

    2015-01-01

    The increased access to books afforded to blind people via e-publishing has given them long-sought independence for both recreational and educational reading. In most cases, blind readers access materials using speech output. For some content such as highly technical texts, music, and graphics, speech is not an appropriate access modality as it does not promote deep understanding. Therefore blind braille readers often prefer electronic braille displays. But, these are prohibitively expensive. The search is on, therefore, for a low-cost refreshable display that would go beyond current technologies and deliver graphical content as well as text. And many solutions have been proposed, some of which reduce costs by restricting the number of characters that can be displayed, even down to a single braille cell. In this paper, we demonstrate that restricting tactile cues during braille reading leads to poorer performance in a letter recognition task. In particular, we show that lack of sliding contact between the fingertip and the braille reading surface results in more errors and that the number of errors increases as a function of presentation speed. These findings suggest that single cell displays which do not incorporate sliding contact are likely to be less effective for braille reading.

  1. Virtual acoustic displays

    Science.gov (United States)

    Wenzel, Elizabeth M.

    1991-01-01

    A 3D auditory display can potentially enhance information transfer by combining directional and iconic information in a quite naturalistic representation of dynamic objects in the interface. Another aspect of auditory spatial clues is that, in conjunction with other modalities, it can act as a potentiator of information in the display. For example, visual and auditory cues together can reinforce the information content of the display and provide a greater sense of presence or realism in a manner not readily achievable by either modality alone. This phenomenon will be particularly useful in telepresence applications, such as advanced teleconferencing environments, shared electronic workspaces, and monitoring telerobotic activities in remote or hazardous situations. Thus, the combination of direct spatial cues with good principles of iconic design could provide an extremely powerful and information-rich display which is also quite easy to use. An alternative approach, recently developed at ARC, generates externalized, 3D sound cues over headphones in realtime using digital signal processing. Here, the synthesis technique involves the digital generation of stimuli using Head-Related Transfer Functions (HRTF's) measured in the two ear-canals of individual subjects. Other similar approaches include an analog system developed by Loomis, et. al., (1990) and digital systems which make use of transforms derived from normative mannikins and simulations of room acoustics. Such an interface also requires the careful psychophysical evaluation of listener's ability to accurately localize the virtual or synthetic sound sources. From an applied standpoint, measurement of each potential listener's HRTF's may not be possible in practice. For experienced listeners, localization performance was only slightly degraded compared to a subject's inherent ability. Alternatively, even inexperienced listeners may be able to adapt to a particular set of HRTF's as long as they provide adequate

  2. Dephosphorylation Pathway of D-myo-Inositol 1,4,5-trisphosphate in the Unicellular Green Alga Chlamydomonas eugametos

    NARCIS (Netherlands)

    Klerk, Hans; Himbergen, John A.J. van; Musgrave, Alan; Haastert, Peter J.M. van; Ende, Herman van den

    1994-01-01

    In vitro dephosphorylation of D-myo-inositol 1,4,5-trisphosphate [Ins(l,4,5)P-3] by vegetative cells, gametes and zygotes of the green alga Chlamydomonas eugametos was studied using a soluble cell fraction as enzyme source and labelled Ins(1,4,5)P-3 as substrate. This compound was dephosphorylated y

  3. Origin of pronounced differences in 77 K fluorescence of the green alga Chlamydomonas reinhardtii in state 1 and 2

    NARCIS (Netherlands)

    Ünlü, Caner; Polukhina, Iryna; Amerongen, van Herbert

    2016-01-01

    In response to changes in the reduction state of the plastoquinone pool in its thylakoid membrane, the green alga Chlamydomonas reinhardtti is performing state transitions: remodelling of its thylakoid membrane leads to a redistribution of excitations over photosystems I and II (PSI and PSII). Th

  4. Measurement of Lipid Droplet Accumulation Kinetics in Chlamydomonas reinhardtii Using Seoul-Fluor

    Directory of Open Access Journals (Sweden)

    Noo Li Jeon

    2013-10-01

    Full Text Available Alternative energy resources have become an important issue due to the limited stocks of petroleum-based fuel. Microalgae, a source of renewable biodiesel, use solar light to convert CO2 into lipid droplets (LDs. Quantification of LDs in microalgae is required for developing and optimizing algal bioprocess engineering. However, conventional quantification methods are both time and labor-intensive and difficult to apply in high-throughput screening systems. LDs in plant and mammalian cells can be visualized by staining with various fluorescence probes such as the Nile Red, BODIPY, and Seoul-Fluor (SF series. This report describes the optimization of LD staining in Chlamydomonas reinhardtii with SF probes via systematic variations of dye concentration, staining time, temperature, and pH. A protocol for quantitative measurement of accumulation kinetics of LDs in C. reinhardtii was developed using a spectrofluorimeter and the accuracy of LD size measurement was confirmed by transmission electron microscopy (TEM. Our results indicate that our spectrofluorimeter-based measurement approach can monitor kinetics of intracellular LDs (in control and nitrogen-source-starved Chlamydomonas reinhardtii accumulation that has not been possible in the case of conventional imaging-based methods. Our results presented here confirmed that an SF44 can be a powerful tool for in situ monitoring and tracking of intracellular LDs formation.

  5. Proteomic Alterations of Antarctic Ice Microalga Chlamydomonas sp. Under Low-Temperature Stress

    Institute of Scientific and Technical Information of China (English)

    Guang-Feng Kan; Jin-Lai Miao; Cui-Juan Shi; Guang-You Li

    2006-01-01

    Antarctic ice microalga can survive and thrive in cold channels or pores in the Antarctic ice layer. In order to understand the adaptive mechanisms to low temperature, in the present study we compared two-dimensional polyacrylamide gel electrophoresis (2-DE) profiles of normal and low temperature-stressed Antarctic ice microalga Chlamydomonas sp. cells. In addition, new protein spots induced by low temperature were identified with peptide mass fingerprinting based on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and database searching. Well-resolved and reproducible 2-DE patterns of both normal and low temperature-stressed cells were acquired. A total of 626 spots was detected in control cells and 652 spots were detected in the corresponding low temperature-stressed cells. A total of 598 spots was matched between normal and stressed cells. Two newly synthesized proteins (a and b) in low temperature-stressed cells were characterized. Protein spot A (53 kDa, pI 6.0) was similar to isopropylmalate/homocitrate/citramalate synthases, which act in the transport and metabolism of amino acids. Protein spot b (25 kDa, pI 8.0) was related to glutathione S-transferase, which functions as a scavenger of active oxygen, free radicals, and noxious metabolites. The present study is valuable for the application of ice microalgae, establishing an ice microalga Chlamydomonas sp. proteome database, and screening molecular biomarkers for further studies.

  6. Characterization of Hydrocortisone Biometabolites and 18S rRNA Gene in Chlamydomonas reinhardtii Cultures

    Directory of Open Access Journals (Sweden)

    Seyed Bagher Mosavi-Azam

    2008-10-01

    Full Text Available A unicellular microalga, Chlamydomonas reinhardtii, was isolated from rice paddy-field soil and water samples and used in the biotransformation of hydrocortisone (1. This strain has not been previously tested for steroid bioconversion. Fermentation was carried out in BG-11 medium supplemented with 0.05% substrate at 25ºC for 14 days of incubation. The products obtained were chromatographically purified and characterized using spectroscopic methods. 11b,17b-Dihydroxyandrost-4-en-3-one (2, 11b-hydroxyandrost-4-en-3,17-dione (3, 11b,17a,20b,21-tetrahydroxypregn-4-en-3-one (4 and prednisolone (5 were the main products of the bioconversion. The observed bioreaction features were the side chain degradation of the substrate to give compounds 2 and 3 and the 20-ketone reduction and 1,2-dehydrogenation affording compounds 4 and 5, respectively. A time course study showed the accumulation of product 2 from the second day of the fermentation and of compounds 3, 4 and 5 from the third day. All the metabolites reached their maximum concentration in seven days. Microalgal 18S rRNA gene was also amplified by PCR. PCR products were sequenced to confirm their authenticity as 18S rRNA gene of microalgae. The result of PCR blasted with other sequenced microalgae in NCBI showed 100% homology to the 18S small subunit rRNA of two Chlamydomonas reinhardtii spp.

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

    Science.gov (United States)

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

    2015-01-01

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

  8. Biofixation of Carbon dioxide by Chlamydomonas sp. in a Tubular Photobioreactor

    Directory of Open Access Journals (Sweden)

    H Hadiyanto

    2012-04-01

    Full Text Available The biogas production from anaerobic digestion is a potential fuel for power generators application, if biogas can be upgraded to the same standards as fossil natural gas by CO2, H2S, and other non-combustible component removal. Microalgae Chlamydomonas sp. has potency to biofix the carbon dioxide and can be used as an additional food ingredient. The variations of flow rate and carbon dioxide concentration in the process resulting different value of biomass production and carbon dioxide biofixation. Biomass production at 40% carbon dioxide concentration obtained 5.685 gr/dm3 at 10% carbon dioxide concentration obtained 4.892 gr/dm3. The greatest value of carbon dioxide absorption occurs at a 40% concentration amounting to 12.09%. The rate of growth and productivity of microalgae tend to rise in 10% and 20% (%v carbon dioxide concentration, but began started a constant at 30% and 40% (%v carbon dioxide concentration. Biomass production tends to increase in light conditions while a constant in dark conditions. This study used Chlamydomonas sp. as media culture and performed on bubble column and tubular reactor with 6 litres of culture medium at a temperature of 28oC and atmospheric pressure.

  9. Biofixation of Carbon dioxide by Chlamydomonas sp. in a Tubular Photobioreactor

    Directory of Open Access Journals (Sweden)

    H Hadiyanto

    2012-02-01

    Full Text Available The biogas production from anaerobic digestion is a potential fuel for power generators application, if biogas can be upgraded to the same standards as fossil natural gas by CO2, H2S, and other non-combustible component removal. Microalgae Chlamydomonas sp. has potency to biofix the carbon dioxide and can be used as an additional food ingredient. The variations of flow rate and carbon dioxide concentration in the process resulting different value of biomass production and carbon dioxide biofixation. Biomass production at 40% carbon dioxide concentration obtained 5.685 gr/dm3 at 10% carbon dioxide concentration obtained 4.892 gr/dm3. The greatest value of carbon dioxide absorption occurs at a 40% concentration amounting to 12.09%. The rate of growth and productivity of microalgae tend to rise in 10% and 20% (%v carbon dioxide concentration, but began started a constant at 30% and 40% (%v carbon dioxide concentration. Biomass production tends to increase in light conditions while a constant in dark conditions. This study used Chlamydomonas sp. as media culture and performed on bubble column and tubular reactor with 6 litres of culture medium at a temperature of 28oC and atmospheric pressure.

  10. Analyses of tomato fruit brightness mutants uncover both cutin-deficient and cutin-abundant mutants and a new hypomorphic allele of GDSL lipase.

    Science.gov (United States)

    Petit, Johann; Bres, Cécile; Just, Daniel; Garcia, Virginie; Mauxion, Jean-Philippe; Marion, Didier; Bakan, Bénédicte; Joubès, Jérôme; Domergue, Frédéric; Rothan, Christophe

    2014-02-01

    The cuticle is a protective layer synthesized by epidermal cells of the plants and consisting of cutin covered and filled by waxes. In tomato (Solanum lycopersicum) fruit, the thick cuticle embedding epidermal cells has crucial roles in the control of pathogens, water loss, cracking, postharvest shelf-life, and brightness. To identify tomato mutants with modified cuticle composition and architecture and to further decipher the relationships between fruit brightness and cuticle in tomato, we screened an ethyl methanesulfonate mutant collection in the miniature tomato cultivar Micro-Tom for mutants with altered fruit brightness. Our screen resulted in the isolation of 16 glossy and 8 dull mutants displaying changes in the amount and/or composition of wax and cutin, cuticle thickness, and surface aspect of the fruit as characterized by optical and environmental scanning electron microscopy. The main conclusions on the relationships between fruit brightness and cuticle features were as follows: (1) screening for fruit brightness is an effective way to identify tomato cuticle mutants; (2) fruit brightness is independent from wax load variations; (3) glossy mutants show either reduced or increased cutin load; and (4) dull mutants display alterations in epidermal cell number and shape. Cuticle composition analyses further allowed the identification of groups of mutants displaying remarkable cuticle changes, such as mutants with increased dicarboxylic acids in cutin. Using genetic mapping of a strong cutin-deficient mutation, we discovered a novel hypomorphic allele of GDSL lipase carrying a splice junction mutation, thus highlighting the potential of tomato brightness mutants for advancing our understanding of cuticle formation in plants.

  11. Refrigerated display cabinets; Butikskyla

    Energy Technology Data Exchange (ETDEWEB)

    Fahlen, Per

    2000-07-01

    This report summarizes experience from SP research and assignments regarding refrigerated transport and storage of food, mainly in the retail sector. It presents the fundamentals of heat and mass transfer in display cabinets with special focus on indirect systems and secondary refrigerants. Moreover, the report includes a brief account of basic food hygiene and the related regulations. The material has been compiled for educational purposes in the Masters program at Chalmers Technical University.

  12. Phenotypic characterization and virulence of a sae- agr- mutant of Staphylococcus aureus.

    Science.gov (United States)

    Giraudo, A T; Rampone, H; Calzolari, A; Nagel, R

    1996-02-01

    A sae::Tn551 agr::tetM double mutant was constructed and characterized. The production of several exoproteins (e.g., beta-hemolysin, DNase, and proteases) by this mutant was determined and found to be lower than the already diminished production of either isogenic single mutant sae- or agr-. The double mutant also showed, like the agr- mutant, null production of alpha- and delta-hemolysins and diminished levels of lipase. The reduced levels of many exoproteins in the double mutant as compared with their already diminished levels in either single mutant suggest that there is an additive or synergistic interaction between the two mutations involved, sae- and agr-. However, inactivation of both loci, sae and agr, had a different effect on the two exoproteins that are up regulated in the agr- mutant; thus, coagulase dropped to levels close to the null levels of the sae- parental strain, while extracellular protein A displayed the high levels characteristic of the agr- single mutant. The virulence of the sae- agr- double mutant, determined by intraperitoneal injection in mice, was found to be significantly diminished as compared with that of the sae+ agr+ parental strain or the sae- agr+ single mutant.

  13. Book Display as Adult Service

    Directory of Open Access Journals (Sweden)

    Matthew S. Moore

    1997-03-01

    Full Text Available 無Book display as an adult service is defined as choosing and positioning adult books from the collection to increase their circulation. The author contrasts bookstore arrangement for sales versus library arrangement for access. The paper considers the library-as-a-whole as a display, examines the right size for an in-library display, and discusses mass displays, end-caps, on-shelf displays, and the Tiffany approach. The author proposes that an effective display depends on an imaginative, unifying theme, and that book displays are part of the joy of libraries.

  14. Handbook of Visual Display Technology

    CERN Document Server

    Cranton, Wayne; Fihn, Mark

    2012-01-01

    The Handbook of Visual Display Technology is a unique work offering a comprehensive description of the science, technology, economic and human interface factors associated with the displays industry. An invaluable compilation of information, the Handbook will serve as a single reference source with expert contributions from over 150 international display professionals and academic researchers. All classes of display device are covered including LCDs, reflective displays, flexible solutions and emissive devices such as OLEDs and plasma displays, with discussion of established principles, emergent technologies, and particular areas of application. The wide-ranging content also encompasses the fundamental science of light and vision, image manipulation, core materials and processing techniques, display driving and metrology.

  15. A millifluidic study of cell-to-cell heterogeneity in growth-rate and cell-division capability in populations of isogenic cells of Chlamydomonas reinhardtii.

    Directory of Open Access Journals (Sweden)

    Shima P Damodaran

    Full Text Available To address possible cell-to-cell heterogeneity in growth dynamics of isogenic cell populations of Chlamydomonas reinhardtii, we developed a millifluidic drop-based device that not only allows the analysis of populations grown from single cells over periods of a week, but is also able to sort and collect drops of interest, containing viable and healthy cells, which can be used for further experimentation. In this study, we used isogenic algal cells that were first synchronized in mixotrophic growth conditions. We show that these synchronized cells, when placed in droplets and kept in mixotrophic growth conditions, exhibit mostly homogeneous growth statistics, but with two distinct subpopulations: a major population with a short doubling-time (fast-growers and a significant subpopulation of slowly dividing cells (slow-growers. These observations suggest that algal cells from an isogenic population may be present in either of two states, a state of restricted division and a state of active division. When isogenic cells were allowed to propagate for about 1000 generations on solid agar plates, they displayed an increased heterogeneity in their growth dynamics. Although we could still identify the original populations of slow- and fast-growers, drops inoculated with a single progenitor cell now displayed a wider diversity of doubling-times. Moreover, populations dividing with the same growth-rate often reached different cell numbers in stationary phase, suggesting that the progenitor cells differed in the number of cell divisions they could undertake. We discuss possible explanations for these cell-to-cell heterogeneities in growth dynamics, such as mutations, differential aging or stochastic variations in metabolites and macromolecules yielding molecular switches, in the light of single-cell heterogeneities that have been reported among isogenic populations of other eu- and prokaryotes.

  16. A millifluidic study of cell-to-cell heterogeneity in growth-rate and cell-division capability in populations of isogenic cells of Chlamydomonas reinhardtii.

    Science.gov (United States)

    Damodaran, Shima P; Eberhard, Stephan; Boitard, Laurent; Rodriguez, Jairo Garnica; Wang, Yuxing; Bremond, Nicolas; Baudry, Jean; Bibette, Jérôme; Wollman, Francis-André

    2015-01-01

    To address possible cell-to-cell heterogeneity in growth dynamics of isogenic cell populations of Chlamydomonas reinhardtii, we developed a millifluidic drop-based device that not only allows the analysis of populations grown from single cells over periods of a week, but is also able to sort and collect drops of interest, containing viable and healthy cells, which can be used for further experimentation. In this study, we used isogenic algal cells that were first synchronized in mixotrophic growth conditions. We show that these synchronized cells, when placed in droplets and kept in mixotrophic growth conditions, exhibit mostly homogeneous growth statistics, but with two distinct subpopulations: a major population with a short doubling-time (fast-growers) and a significant subpopulation of slowly dividing cells (slow-growers). These observations suggest that algal cells from an isogenic population may be present in either of two states, a state of restricted division and a state of active division. When isogenic cells were allowed to propagate for about 1000 generations on solid agar plates, they displayed an increased heterogeneity in their growth dynamics. Although we could still identify the original populations of slow- and fast-growers, drops inoculated with a single progenitor cell now displayed a wider diversity of doubling-times. Moreover, populations dividing with the same growth-rate often reached different cell numbers in stationary phase, suggesting that the progenitor cells differed in the number of cell divisions they could undertake. We discuss possible explanations for these cell-to-cell heterogeneities in growth dynamics, such as mutations, differential aging or stochastic variations in metabolites and macromolecules yielding molecular switches, in the light of single-cell heterogeneities that have been reported among isogenic populations of other eu- and prokaryotes.

  17. Indy mutants: live long and prosper

    Directory of Open Access Journals (Sweden)

    Stewart eFrankel

    2012-02-01

    Full Text Available Indy encodes the fly homologue of a mammalian transporter of di and tricarboxylatecomponents of the Krebs cycle. Reduced expression of fly Indy or two of the C. elegansIndy homologs leads to an increase in life span. Fly and worm tissues that play key roles inintermediary metabolism are also the places where Indy genes are expressed. One of themouse homologs of Indy (mIndy is mainly expressed in the liver. It has been hypothesizedthat decreased INDY activity creates a state similar to caloric restriction (CR. Thishypothesis is supported by the physiological similarities between Indy mutant flies on highcalorie food and control flies on CR, such as increased physical activity and decreases inweight, egg production, triglyceride levels, starvation resistance, and insulin signaling. Inaddition, Indy mutant flies undergo changes in mitochondrial biogenesis also observed inCR animals. Recent findings with mIndy knockout mice support and extend the findingsfrom flies. mIndy-/- mice display an increase in hepatic mitochondrial biogenesis, lipidoxidation and decreased hepatic lipogenesis. When mIndy-/- mice are fed high calorie foodthey are protected from adiposity and insulin resistance. These findings point to INDY as apotential drug target for the treatment of metabolic syndrome, type 2 diabetes and obesity.

  18. Sulphate, more than a nutrient, protects the microalga Chlamydomonas moewusii from cadmium toxicity

    Energy Technology Data Exchange (ETDEWEB)

    Mera, Roi; Torres, Enrique, E-mail: torres@udc.es; Abalde, Julio

    2014-03-01

    Highlights: • Sulphate effect on cadmium toxicity in the microalga Chlamydomonas moewusii Gerloff. • Cadmium increases the sulphur requirements in Chlamydomonas moewusii. • Kinetic coefficients for sulphate utilization and cadmium effect on them. • Sulphate and cadmium influence on the biosynthesis of low-molecular mass thiols. • Cadmium toxicity reduction by sulphate due to higher biosynthesis of thiols. - Abstract: Sulphur is an essential macroelement that plays important roles in living organisms. The thiol rich sulphur compounds, such as cysteine, γ-Glu–Cys, glutathione and phytochelatins participate in the tolerance mechanisms against cadmium toxicity. Plants, algae, yeasts and most prokaryotes cover their demand for reduced sulphur by reduction of inorganic sulphate. The aim of this study was to investigate, using a bifactorial experimental design, the effect of different sulphate concentrations in the nutrient solution on cadmium toxicity in the freshwater microalga Chlamydomonas moewusii. Cell growth, kinetic parameters of sulphate utilization and intracellular concentrations of low-molecular mass thiol compounds were determined. A mathematical model to describe the growth of this microalga based on the effects of sulphate and cadmium was obtained. An ANOVA revealed an interaction between them, 16% of the effect sizes was explained by this interaction. A higher amount of sulphate in the culture medium allowed a higher cadmium tolerance due to an increase in the thiol compound biosynthesis. The amount of low-molecular mass thiol compounds, mainly phytochelatins, synthesized by this microalga was significantly dependent on the sulphate and cadmium concentrations; the higher phytochelatin content was obtained in cultures with 4 mg Cd/L and 1 mM sulphate. The maximum EC{sub 50} value (based on nominal cadmium concentration) reached for this microalga was 4.46 ± 0.42 mg Cd/L when the sulphate concentration added to the culture medium was also 1 m

  19. Latest development of display technologies

    Science.gov (United States)

    Gao, Hong-Yue; Yao, Qiu-Xiang; Liu, Pan; Zheng, Zhi-Qiang; Liu, Ji-Cheng; Zheng, Hua-Dong; Zeng, Chao; Yu, Ying-Jie; Sun, Tao; Zeng, Zhen-Xiang

    2016-09-01

    In this review we will focus on recent progress in the field of two-dimensional (2D) and three-dimensional (3D) display technologies. We present the current display materials and their applications, including organic light-emitting diodes (OLEDs), flexible OLEDs quantum dot light emitting diodes (QLEDs), active-matrix organic light emitting diodes (AMOLEDs), electronic paper (E-paper), curved displays, stereoscopic 3D displays, volumetric 3D displays, light field 3D displays, and holographic 3D displays. Conventional 2D display devices, such as liquid crystal devices (LCDs) often result in ambiguity in high-dimensional data images because of lacking true depth information. This review thus provides a detailed description of 3D display technologies.

  20. Light-intensity-dependent expression of Lhc gene family encoding light-harvesting chlorophyll-a/b proteins of photosystem II in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Teramoto, Haruhiko; Nakamori, Akira; Minagawa, Jun; Ono, Taka-aki

    2002-09-01

    Excessive light conditions repressed the levels of mRNAs accumulation of multiple Lhc genes encoding light-harvesting chlorophyll-a/b (LHC) proteins of photosystem (PS)II in the unicellular green alga, Chlamydomonas reinhardtii. The light intensity required for the repression tended to decrease with lowering temperature or CO(2) concentration. The responses of six LhcII genes encoding the major LHC (LHCII) proteins and two genes (Lhcb4 and Lhcb5) encoding the minor LHC proteins of PSII (CP29 and CP26) were similar. The results indicate that the expression of these Lhc genes is coordinately repressed when the energy input through the antenna systems exceeds the requirement for CO(2) assimilation. The Lhc mRNA level repressed under high-light conditions was partially recovered by adding the electron transport inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea, suggesting that redox signaling via photosynthetic electron carriers is involved in the gene regulation. However, the mRNA level was still considerably lower under high-light than under low-light conditions even in the presence of 3-(3,4-dichlorophenyl)-1,1-dimethylurea. Repression of the Lhc genes by high light was prominent even in the mutants deficient in the reaction center(s) of PSII or both PSI and PSII. The results indicate that two alternative processes are involved in the repression of Lhc genes under high-light conditions, one of which is independent of the photosynthetic reaction centers and electron transport events.

  1. Inhomogeneous distribution of Chlamydomonas in a cylindrical container with a bubble plume.

    Science.gov (United States)

    Nonaka, Yuki; Kikuchi, Kenji; Numayama-Tsuruta, Keiko; Kage, Azusa; Ueno, Hironori; Ishikawa, Takuji

    2016-01-19

    Swimming microalgae show various taxes, such as phototaxis and gravitaxis, which sometimes result in the formation of a cell-rich layer or a patch in a suspension. Despite intensive studies on the effects of shear flow and turbulence on the inhomogeneous distribution of microalgae, the effect of a bubble plume has remained unclear. In this study, we used Chlamydomonas as model microalgae, and investigated the spatial distribution of cells in a cylindrical container with a bubble plume. The results illustrate that cells become inhomogeneously distributed in the suspension due to their motility and photo-responses. A vortical ring distribution was observed below the free surface when the bubble flow rate was sufficiently small. We performed a scaling analysis on the length scale of the vortical ring, which captured the main features of the experimental results. These findings are important in understanding transport phenomena in a microalgae suspension with a bubble plume.

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

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

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

  3. 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.

  4. Efficient phototrophic production of a high-value sesquiterpenoid from the eukaryotic microalga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Lauersen, Kyle J; Baier, Thomas; Wichmann, Julian; Wördenweber, Robin; Mussgnug, Jan H; Hübner, Wolfgang; Huser, Thomas; Kruse, Olaf

    2016-11-01

    The heterologous expression of terpene synthases in microbial hosts has opened numerous possibilities for bioproduction of desirable metabolites. Photosynthetic microbial hosts present a sustainable alternative to traditional fermentative systems, using freely available (sun)light and carbon dioxide as inputs for bio-production. Here, we report the expression of a patchoulol synthase from Pogostemon cablin Benth in the model green microalga Chlamydomonas reinhardtii. The sesquiterpenoid patchoulol was produced from the alga and was used as a marker of sesquiterpenoid production capacity. A novel strategy for gene loading was employed and patchoulol was produced up to 922±242µgg(-1) CDW in six days. We additionally investigated the effect of carbon source on sesquiterpenoid productivity from C. reinhardtii in scale-up batch cultivations. It was determined that up to 1.03mgL(-1) sesquiterpenoid products could be produced in completely photoautotrophic conditions and that the alga exhibited altered sesquiterpenoid production metabolism related to carbon source.

  5. Quantification of phytochelatins in Chlamydomonas reinhardtii using ferrocene-based derivatization.

    Science.gov (United States)

    Bräutigam, Anja; Bomke, Susanne; Pfeifer, Thorben; Karst, Uwe; Krauss, Gerd-Joachim; Wesenberg, Dirk

    2010-08-01

    A method for the identification and quantification of canonic and isoforms of phytochelatins (PCs) from Chlamydomonas reinhardtii was developed. After disulfide reduction with tris(2-carboxyethyl)phosphine (TCEP) PCs were derivatized with ferrocenecarboxylic acid (2-maleimidoyl)ethylamide (FMEA) in order to avoid oxidation of the free thiol functions during analysis. Liquid chromatography (LC) coupled to electrospray mass spectrometry (ESI-MS) and inductively coupled plasma-mass spectrometry (ICP-MS) was used for rapid and quantitative analysis of the precolumn derivatized PCs. PC(2-4), CysGSH, CysPC(2-4), CysPC(2)desGly, CysPC(2)Glu and CysPC(2)Ala were determined in the algal samples depending on the exposure of the cells to cadmium ions.

  6. Cell types and their status in Chlamydomonas-like algae (Chlorophyceae on agar medium culture

    Directory of Open Access Journals (Sweden)

    M.М. Pavlovska

    2014-04-01

    Full Text Available The classification of cell types under agar culture was proposed. Six cell morphotypes were allocated. The statuses were identified depending on the reduction of monade attributes of cells. The variants of transition from one cell morphotype to another under dissolving mucilage were shown. The monade, cocciod, palmeloid and gloeocysta morphotypes approximately equally represented in all clades. The asterococcus and mucogleocysta morphotypes presented only in Reinhardtinia аnd Oogamochlamydinia clades. Any morphotype isn’t typical for all clades of Chlamydomonas-like algae at once. The most of morphotypes numbers (5 from 6 are presented in Reinhardtinia clade. This demonstrates the diversity of the Reinhardtinia clade species. There are only one morphotype presented in Polytominia and Monadinia clades. There are four morphotypes presented in Oogamochlamydinia clade, three – in Moewusinia, two morphotypes – in Chloromonadinia.

  7. Stable expression of a bifunctional diterpene synthase in the chloroplast of Chlamydomonas reinhardtii

    DEFF Research Database (Denmark)

    Zedler, Julie A Z; Gangl, Doris; Hamberger, Björn Robert;

    2015-01-01

    Chlamydomonas reinhardtii has been shown to hold significant promise as a production platform for recombinant proteins, but transformation of the nuclear genome is still a non-trivial process due to random gene insertion and frequent silencing. Insertion of transgenes into the chloroplasts...... is an alternative strategy, and we report here the stable expression of a large (91 kDa) protein in the chloroplast using a recently developed low-cost transformation protocol. Moreover, selection of transformants is based on restoration of prototrophy using an endogenous gene (psbH) as the marker, thereby allowing...... the generation of transgenic lines without the use of antibiotic-resistance genes. Here, we have expressed a bifunctional diterpene synthase in C. reinhardtii chloroplasts. Homoplasmic transformants were obtained with the expressed enzyme accounting for 3.7 % of total soluble protein. The enzyme was purified...

  8. DNA-free two-gene knockout in Chlamydomonas reinhardtii via CRISPR-Cas9 ribonucleoproteins.

    Science.gov (United States)

    Baek, Kwangryul; Kim, Duk Hyoung; Jeong, Jooyeon; Sim, Sang Jun; Melis, Anastasios; Kim, Jin-Soo; Jin, EonSeon; Bae, Sangsu

    2016-07-28

    Microalgae are versatile organisms capable of converting CO2, H2O, and sunlight into fuel and chemicals for domestic and industrial consumption. Thus, genetic modifications of microalgae for enhancing photosynthetic productivity, and biomass and bio-products generation are crucial for both academic and industrial applications. However, targeted mutagenesis in microalgae with CRISPR-Cas9 is limited. Here we report, a one-step transformation of Chlamydomonas reinhardtii by the DNA-free CRISPR-Cas9 method rather than plasmids that encode Cas9 and guide RNAs. Outcome was the sequential CpFTSY and ZEP two-gene knockout and the generation of a strain constitutively producing zeaxanthin and showing improved photosynthetic productivity.

  9. Zinc Deficiency Impacts CO2 Assimilation and Disrupts Copper Homeostasis in Chlamydomonas reinhardtii*

    Science.gov (United States)

    Malasarn, Davin; Kropat, Janette; Hsieh, Scott I.; Finazzi, Giovanni; Casero, David; Loo, Joseph A.; Pellegrini, Matteo; Wollman, Francis-André; Merchant, Sabeeha S.

    2013-01-01

    Zinc is an essential nutrient because of its role in catalysis and in protein stabilization, but excess zinc is deleterious. We distinguished four nutritional zinc states in the alga Chlamydomonas reinhardtii: toxic, replete, deficient, and limited. Growth is inhibited in zinc-limited and zinc-toxic cells relative to zinc-replete cells, whereas zinc deficiency is visually asymptomatic but distinguished by the accumulation of transcripts encoding ZIP family transporters. To identify targets of zinc deficiency and mechanisms of zinc acclimation, we used RNA-seq to probe zinc nutrition-responsive changes in gene expression. We identified genes encoding zinc-handling components, including ZIP family transporters and candidate chaperones. Additionally, we noted an impact on two other regulatory pathways, the carbon-concentrating mechanism (CCM) and the nutritional copper regulon. Targets of transcription factor Ccm1 and various CAH genes are up-regulated in zinc deficiency, probably due to reduced carbonic anhydrase activity, validated by quantitative proteomics and immunoblot analysis of Cah1, Cah3, and Cah4. Chlamydomonas is therefore not able to grow photoautotrophically in zinc-limiting conditions, but supplementation with 1% CO2 restores growth to wild-type rates, suggesting that the inability to maintain CCM is a major consequence of zinc limitation. The Crr1 regulon responds to copper limitation and is turned on in zinc deficiency, and Crr1 is required for growth in zinc-limiting conditions. Zinc-deficient cells are functionally copper-deficient, although they hyperaccumulate copper up to 50-fold over normal levels. We suggest that zinc-deficient cells sequester copper in a biounavailable form, perhaps to prevent mismetallation of critical zinc sites. PMID:23439652

  10. A revised mineral nutrient supplement increases biomass and growth rate in Chlamydomonas reinhardtii

    Science.gov (United States)

    Kropat, Janette; Hong-Hermesdorf, Anne; Casero, David; Ent, Petr; Castruita, Madeli; Pellegrini, Matteo; Merchant, Sabeeha S.; Malasarn, Davin

    2011-01-01

    Summary Interest in exploiting algae as a biofuel source and the role of inorganic nutrient deficiency in inducing triacylglyceride (TAG) accumulation in cells necessitates a strategy to efficiently formulate species-specific culture media that can easily be manipulated. Using the reference organism Chlamydomonas reinhardtii, we tested the hypothesis that modeling trace element supplements after the cellular ionome would result in optimized cell growth. We determined the trace metal content of several commonly used Chlamydomonas strains in various culture conditions and developed a revised trace element solution to parallel these measurements. Comparison of cells growing in the revised supplement versus a traditional trace element solution revealed faster growth rates and higher maximum cell densities with the revised recipe. RNA-seq analysis of cultures growing in the traditional versus revised medium suggest that the variation in transcriptomes was smaller than that found between different wild-type strains grown in traditional Hutner’s supplement. Visual observation did not reveal defects in cell motility or mating efficiency in the new supplement. Ni2+-inducible expression from the CYC6 promoter remained a useful tool, albeit with an increased requirement for Ni2+ because of the introduction of an EDTA buffer system in the revised medium. Other advantages include more facile preparation of trace element stock solutions, a reduction in total chemical use, a more consistent batch-to-batch formulation, and long-term stability (tested up to 5 years). Under the new growth regime, we analyzed cells growing under different macro- and micronutrient-deficiencies. TAG accumulation in N deficiency is comparable in the new medium. Fe and Zn deficiency also induced TAG accumulation, as suggested by Nile Red staining. This approach can be used to efficiently optimize culture conditions for other algal species to improve growth and to assay cell physiology. PMID:21309872

  11. CO2 acquisition in Chlamydomonas acidophila is influenced mainly by CO2, not phosphorus, availability.

    Science.gov (United States)

    Spijkerman, Elly; Stojkovic, Slobodanka; Beardall, John

    2014-09-01

    The extremophilic green microalga Chlamydomonas acidophila grows in very acidic waters (pH 2.3-3.4), where CO2 is the sole inorganic carbon source. Previous work has revealed that the species can accumulate inorganic carbon (Ci) and exhibits high affinity CO2 utilization under low-CO2 (air-equilibrium) conditions, similar to organisms with an active CO2 concentrating mechanism (CCM), whereas both processes are down-regulated under high CO2 (4.5 % CO2) conditions. Responses of this species to phosphorus (Pi)-limited conditions suggested a contrasting regulation of the CCM characteristics. Therefore, we measured external carbonic anhydrase (CAext) activities and protein expression (CAH1), the internal pH, Ci accumulation, and CO2-utilization in cells adapted to high or low CO2 under Pi-replete and Pi-limited conditions. Results reveal that C. acidophila expressed CAext activity and expressed a protein cross-reacting with CAH1 (the CAext from Chlamydomonas reinhardtii). Although the function of this CA remains unclear, CAext activity and high affinity CO2 utilization were the highest under low CO2 conditions. C. acidophila accumulated Ci and expressed the CAH1 protein under all conditions tested, and C. reinhardtii also contained substantial amounts of CAH1 protein under Pi-limitation. In conclusion, Ci utilization is optimized in C. acidophila under ecologically relevant conditions, which may enable optimal survival in its extreme Ci- and Pi-limited habitat. The exact physiological and biochemical acclimation remains to be further studied.

  12. Survival and proliferation characteristics of the microalga Chlamydomonas sp. ICE-L after hypergravitational stress pretreatment

    Science.gov (United States)

    Gao, Zhengquan; Li, Demao; Meng, Chunxiao; Xu, Dong; Zhang, Xiaowen; Ye, Naihao

    2013-09-01

    Seeking extraterrestrial life, transferring between planets, even migrating to other planets attracts more and more attention of public and scientists. However, to make it clear for the ability to survive the forces studies is prerequisite to enable the speculations by natural means. Gravity is a critical force involved in all the life on Earth and, possibly, others planets. Organisms have been grown in microgravity habitats and in centrifuges to characterize the biological response to a range of gravitational forces and radiation levels in space and on Earth. However, little is known about the profiles of eukaryotic life under conditions of hyperacceleration attributable to extreme gravities. In this study, a eukaryotic extremophile, the Antarctic green microalga Chlamydomonas sp. ICE-L, showed amazing proliferation capacity during and after hypergravitational stress for 30 min to 48 h at 110,200, 423,400, and 670,800g. These extreme gravities also had profound effects on viability, reproduction rate, photosynthesis efficiency, and gene transcriptional expression of this microalga. Most notably, all three supergravities efficiently stimulated algal cell division, but the greater the centrifugal force and the longer the duration of treatment, the lower the viable rate and breeding potential of samples in the following incubation. These results illustrated Chlamydomonas sp. ICE-L is a useful eukaryotic model system candidate for space research. Further studies could provide new insight into the physical limits of life and its evolution and enhance the possibility for interstellar space travel and the quest for extraterrestrial life according to panspermia theory. Also, it indicated that life come from the outer space is not always prokaryotes but may be eukaryotes.

  13. A Chlamydomonas-derived Human Papillomavirus 16 E7 vaccine induces specific tumor protection.

    Directory of Open Access Journals (Sweden)

    Olivia C Demurtas

    Full Text Available BACKGROUND: The E7 protein of the Human Papillomavirus (HPV type 16, being involved in malignant cellular transformation, represents a key antigen for developing therapeutic vaccines against HPV-related lesions and cancers. Recombinant production of this vaccine antigen in an active form and in compliance with good manufacturing practices (GMP plays a crucial role for developing effective vaccines. E7-based therapeutic vaccines produced in plants have been shown to be active in tumor regression and protection in pre-clinical models. However, some drawbacks of in whole-plant vaccine production encouraged us to explore the production of the E7-based therapeutic vaccine in Chlamydomonas reinhardtii, an organism easy to grow and transform and fully amenable to GMP guidelines. METHODOLOGY/PRINCIPAL FINDINGS: An expression cassette encoding E7GGG, a mutated, attenuated form of the E7 oncoprotein, alone or as a fusion with affinity tags (His6 or FLAG, under the control of the C. reinhardtii chloroplast psbD 5' UTR and the psbA 3' UTR, was introduced into the C. reinhardtii chloroplast genome by homologous recombination. The protein was mostly soluble and reached 0.12% of total soluble proteins. Affinity purification was optimized and performed for both tagged forms. Induction of specific anti-E7 IgGs and E7-specific T-cell proliferation were detected in C57BL/6 mice vaccinated with total Chlamydomonas extract and with affinity-purified protein. High levels of tumor protection were achieved after challenge with a tumor cell line expressing the E7 protein. CONCLUSIONS: The C. reinhardtii chloroplast is a suitable expression system for the production of the E7GGG protein, in a soluble, immunogenic form. The production in contained and sterile conditions highlights the potential of microalgae as alternative platforms for the production of vaccines for human uses.

  14. Light-harvesting complex II (LHCII) and its supramolecular organization in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Drop, Bartlomiej; Webber-Birungi, Mariam; Yadav, Sathish K N; Filipowicz-Szymanska, Alicja; Fusetti, Fabrizia; Boekema, Egbert J; Croce, Roberta

    2014-01-01

    LHCII is the most abundant membrane protein on earth. It participates in the first steps of photosynthesis by harvesting sunlight and transferring excitation energy to the core complex. Here we have analyzed the LHCII complex of the green alga Chlamydomonas reinhardtii and its association with the core of Photosystem II (PSII) to form multiprotein complexes. Several PSII supercomplexes with different antenna sizes have been purified, the largest of which contains three LHCII trimers (named S, M and N) per monomeric core. A projection map at a 13Å resolution was obtained allowing the reconstruction of the 3D structure of the supercomplex. The position and orientation of the S trimer are the same as in plants; trimer M is rotated by 45° and the additional trimer (named here as LHCII-N), which is taking the position occupied in plants by CP24, is directly associated with the core. The analysis of supercomplexes with different antenna sizes suggests that LhcbM1, LhcbM2/7 and LhcbM3 are the major components of the trimers in the PSII supercomplex, while LhcbM5 is part of the "extra" LHCII pool not directly associated with the supercomplex. It is also shown that Chlamydomonas LHCII has a slightly lower Chlorophyll a/b ratio than the complex from plants and a blue shifted absorption spectrum. Finally the data indicate that there are at least six LHCII trimers per dimeric core in the thylakoid membranes, meaning that the antenna size of PSII of C. reinhardtii is larger than that of plants.

  15. Impaired exercise tolerance and skeletal muscle myopathy in sulfonylurea receptor-2 mutant mice

    Science.gov (United States)

    Stoller, Douglas; Pytel, Peter; Katz, Sophie; Earley, Judy U.; Collins, Keith; Metcalfe, Jamie; Lang, Roberto M.

    2009-01-01

    By sensing intracellular energy levels, ATP-sensitive potassium (KATP) channels help regulate vascular tone, glucose metabolism, and cardioprotection. SUR2 mutant mice lack full-length KATP channels in striated and smooth muscle and display a complex phenotype of hypertension and coronary vasospasm. SUR2 mutant mice also display baseline cardioprotection and can withstand acute sympathetic stress better than normal mice. We now studied response to a form of chronic stress, namely that induced by 4 wk of daily exercise on SUR2 mutant mice. Control mice increased exercise capacity by 400% over the training period, while SUR2 mutant mice showed little increase in exercise capacity. Unexercised SUR2 mutant showed necrotic and regenerating fibers in multiple muscle skeletal muscles, including quadriceps, tibialis anterior, and diaphragm muscles. Unlike exercised control animals, SUR2 mutant mice did not lose weight, presumably due to less overall exertion. Unexercised SUR2 mutant mice showed a trend of mildly reduced cardiac function, measured by fractional shortening, (46 ± 4% vs. 57 ± 7% for SUR2 mutant and control, respectively), and this decrease was not exacerbated by chronic exercise exposure. Despite an improved response to acute sympathetic stress and baseline cardioprotection, exercise intolerance results from lack of SUR2 KATP channels in mice. PMID:19675276

  16. Isolation of Aneuploid-Generating Mutants of ASPERGILLUS NIDULANS, One of Which Is Defective in Interphase of the Cell Cycle

    OpenAIRE

    Upshall, A; Mortimore, I. D.

    1984-01-01

    A method is described for isolating mutants potentially defective in loci involved in mitotic chromosome segregation. Conditional lethal, heat-sensitive (42°) mutants were assayed at a subrestrictive temperature of 37° for an inflated production of colonies displaying phenotypes and behavior patterns of whole chromosome aneuploids. Of 14 mutants, three showed specificity for one disomic phenotype, whereas 11 generated colonies mosaic for different aneuploid phenotypes. This latter group is de...

  17. Optimal screening of surface-displayed polypeptide libraries.

    Science.gov (United States)

    Boder, E T; Wittrup, K D

    1998-01-01

    Cell surface display of polypeptide libraries combined with flow cytometric cell sorting presents remarkable potential for enhancement of protein-ligand recognition properties. To maximize the utility of this approach, screening and purification conditions must be optimized to take full advantage of the quantitative feature of this technique. In particular, discrimination of improved library mutants from an excess of wild-type polypeptides is dependent upon an effective screening methodology. Fluorescence discrimination profiles for improved library mutants were derived from a mathematical model of expected cell fluorescence intensities for polypeptide libraries screened with fluorescent ligand. Profiles for surface-displayed libraries under equilibrium or kinetic screening conditions demonstrate distinct discrimination optima from which optimal equilibrium and kinetic screening parameters were derived. In addition, a statistical model of low cytometrically analyzed cell populations indicates the importance of low-stringency sorting followed by amplification through regrowth and resorting at increased stringency. This analysis further yields quantitative recommendations for cell-sorting stringency.

  18. Responsibility of regulatory gene expression and repressed protein synthesis for triacylglycerol accumulation on sulfur-starvation in Chlamydomonas reinhardtii

    OpenAIRE

    2014-01-01

    Triacylglycerol (TG) synthesis is induced for energy and carbon storage in algal cells under nitrogen(N)-starved conditions, and helps prevent reactive oxygen species (ROS) production through fatty acid synthesis that consumes excessive reducing power. Here, the regulatory mechanism for the TG content in sulfur(S)-starved cells of Chlamydomonas reinhardtii was examined, in comparison to that in N- or phosphorus(P)-starved cells. S- and N- starved cells exhibited markedly increased TG contents...

  19. Colorimetry for CRT displays.

    Science.gov (United States)

    Golz, Jürgen; MacLeod, Donald I A

    2003-05-01

    We analyze the sources of error in specifying color in CRT displays. These include errors inherent in the use of the color matching functions of the CIE 1931 standard observer when only colorimetric, not radiometric, calibrations are available. We provide transformation coefficients that prove to correct the deficiencies of this observer very well. We consider four different candidate sets of cone sensitivities. Some of these differ substantially; variation among candidate cone sensitivities exceeds the variation among phosphors. Finally, the effects of the recognized forms of observer variation on the visual responses (cone excitations or cone contrasts) generated by CRT stimuli are investigated and quantitatively specified. Cone pigment polymorphism gives rise to variation of a few per cent in relative excitation by the different phosphors--a variation larger than the errors ensuing from the adoption of the CIE standard observer, though smaller than the differences between some candidate cone sensitivities. Macular pigmentation has a larger influence, affecting mainly responses to the blue phosphor. The estimated combined effect of all sources of observer variation is comparable in magnitude with the largest differences between competing cone sensitivity estimates but is not enough to disrupt very seriously the relation between the L and M cone weights and the isoluminance settings of individual observers. It is also comparable with typical instrumental colorimetric errors, but we discuss these only briefly.

  20. LHCb Event display

    CERN Document Server

    Trisovic, Ana

    2014-01-01

    The LHCb Event Display was made for educational purposes at the European Organization for Nuclear Research, CERN in Geneva, Switzerland. The project was implemented as a stand-alone application using C++ and ROOT, a framework developed by CERN for data analysis. This paper outlines the development and architecture of the application in detail, as well as the motivation for the development and the goals of the exercise. The application focuses on the visualization of events recorded by the LHCb detector, where an event represents a set of charged particle tracks in one proton-proton collision. Every particle track is coloured by its type and can be selected to see its essential information such as mass and momentum. The application allows students to save this information and calculate the invariant mass for any pair of particles. Furthermore, the students can use additional calculating tools in the application and build up a histogram of these invariant masses. The goal for the students is to find a $D^0$ par...

  1. Data Display in Qualitative Research

    Directory of Open Access Journals (Sweden)

    Susana Verdinelli PsyD

    2013-02-01

    Full Text Available Visual displays help in the presentation of inferences and conclusions and represent ways of organizing, summarizing, simplifying, or transforming data. Data displays such as matrices and networks are often utilized to enhance data analysis and are more commonly seen in quantitative than in qualitative studies. This study reviewed the data displays used by three prestigious qualitative research journals within a period of three years. The findings include the types of displays used in these qualitative journals, the frequency of use, and the purposes for using visual displays as opposed to presenting data in text.

  2. The ferredoxin-thioredoxin system of a green alga, Chlamydomonas reinhardtii: identification and characterization of thioredoxins and ferredoxin-thioredoxin reductase components

    Science.gov (United States)

    Huppe, H. C.; de Lamotte-Guery, F.; Buchanan, B. B.

    1990-01-01

    The components of the ferredoxin-thioredoxin (FT) system of Chlamydomonas reinhardtii have been purified and characterized. The system resembled that of higher plants in consisting of a ferredoxin-thioredoxin reductase (FTR) and two types of thioredoxin, a single f and two m species, m1 and m2. The Chlamydomonas m and f thioredoxins were antigenically similar to their higher-plant counterparts, but not to one another. The m thioredoxins were recognized by antibodies to both higher plant m and bacterial thioredoxins, whereas the thioredoxin f was not. Chlamydomonas thioredoxin f reacted, although weakly, with the antibody to spinach thioredoxin f. The algal thioredoxin f differed from thioredoxins studied previously in behaving as a basic protein on ion-exchange columns. Purification revealed that the algal thioredoxins had molecular masses (Mrs) typical of thioredoxins from other sources, m1 and m2 being 10700 and f 11500. Chlamydomonas FTR had two dissimilar subunits, a feature common to all FTRs studied thus far. One, the 13-kDa ("similar") subunit, resembled its counterpart from other sources in both size and antigenicity. The other, 10-kDa ("variable") subunit was not recognized by antibodies to any FTR tested. When combined with spinach, (Spinacia oleracea L.) thylakoid membranes, the components of the FT system functioned in the light activation of the standard target enzymes from chloroplasts, corn (Zea mays L.) NADP-malate dehydrogenase (EC 1.1.1.82) and spinach fructose 1,6-bisphosphatase (EC 3.1.3.11) as well as the chloroplast-type fructose 1,6-bisphosphatase from Chlamydomonas. Activity was greatest if ferredoxin and other components of the FT system were from Chlamydomonas. The capacity of the Chlamydomonas FT system to activate autologous FBPase indicates that light regulates the photosynthetic carbon metabolism of green algae as in other oxygenic photosynthetic organisms.

  3. Whole-Genome Resequencing Reveals Extensive Natural Variation in the Model Green Alga Chlamydomonas reinhardtii[OPEN

    Science.gov (United States)

    Hazzouri, Khaled M.; Rosas, Ulises; Bahmani, Tayebeh; Nelson, David R.; Abdrabu, Rasha; Harris, Elizabeth H.; Salehi-Ashtiani, Kourosh; Purugganan, Michael D.

    2015-01-01

    We performed whole-genome resequencing of 12 field isolates and eight commonly studied laboratory strains of the model organism Chlamydomonas reinhardtii to characterize genomic diversity and provide a resource for studies of natural variation. Our data support previous observations that Chlamydomonas is among the most diverse eukaryotic species. Nucleotide diversity is ∼3% and is geographically structured in North America with some evidence of admixture among sampling locales. Examination of predicted loss-of-function mutations in field isolates indicates conservation of genes associated with core cellular functions, while genes in large gene families and poorly characterized genes show a greater incidence of major effect mutations. De novo assembly of unmapped reads recovered genes in the field isolates that are absent from the CC-503 assembly. The laboratory reference strains show a genomic pattern of polymorphism consistent with their origin as the recombinant progeny of a diploid zygospore. Large duplications or amplifications are a prominent feature of laboratory strains and appear to have originated under laboratory culture. Extensive natural variation offers a new source of genetic diversity for studies of Chlamydomonas, including naturally occurring alleles that may prove useful in studies of gene function and the dissection of quantitative genetic traits. PMID:26392080

  4. An improved ARS2-derived nuclear reporter enhances the efficiency and ease of genetic engineering in Chlamydomonas.

    Science.gov (United States)

    Specht, Elizabeth A; Nour-Eldin, Hussam Hassan; Hoang, Kevin T D; Mayfield, Stephen P

    2015-03-01

    The model alga Chlamydomonas reinhardtii has been used to pioneer genetic engineering techniques for high-value protein and biofuel production from algae. To date, most studies of transgenic Chlamydomonas have utilized the chloroplast genome due to its ease of engineering, with a sizeable suite of reporters and well-characterized expression constructs. The advanced manipulation of algal nuclear genomes has been hampered by limited strong expression cassettes, and a lack of high-throughput reporters. We have improved upon an endogenous reporter gene - the ARS2 gene encoding an arylsulfatase enzyme - that was first cloned and characterized decades ago but has not been used extensively. The new construct, derived from ARS2 cDNA, expresses significantly higher levels of reporter protein and transforms more efficiently, allowing qualitative and quantitative screening using a rapid, inexpensive 96-well assay. The improved arylsulfatase expression cassette was used to screen a new transgene promoter from the ARG7 gene, and found that the ARG7 promoter can express the ARS2 reporter as strongly as the HSP70-RBCS2 chimeric promoter that currently ranks as the best available promoter, thus adding to the list of useful nuclear promoters. This enhanced arylsulfatase reporter construct improves the efficiency and ease of genetic engineering within the Chlamydomonas nuclear genome, with potential application to other algal strains.

  5. Unique interactive projection display screen

    Energy Technology Data Exchange (ETDEWEB)

    Veligdan, J.T.

    1997-11-01

    Projection systems continue to be the best method to produce large (1 meter and larger) displays. However, in order to produce a large display, considerable volume is typically required. The Polyplanar Optic Display (POD) is a novel type of projection display screen, which for the first time, makes it possible to produce a large projection system that is self-contained and only inches thick. In addition, this display screen is matte black in appearance allowing it to be used in high ambient light conditions. This screen is also interactive and can be remotely controlled via an infrared optical pointer resulting in mouse-like control of the display. Furthermore, this display need not be flat since it can be made curved to wrap around a viewer as well as being flexible.

  6. Isolation and characterization of rice lesion mimic mutants from a T-DNA tagged population

    Institute of Scientific and Technical Information of China (English)

    LI Shutian; PEI Zhongyou; LUO Lijuan; TIAN Yingchuan; HE Chaozu

    2005-01-01

    A rice ( Oryza sativa L. ssp. japonica cv. Nipponbare) T-DNA tagged population consisting of about 7000 individual lines was generated and screened for rice lesion mimic mutants in the T1 generation. Ten lines were found to develop spontaneous lesions in the absence of pathogen infection and displayed distinct lesion phenotypes. These mutants were tentatively designated as lm1 -lm10 (for lesion mimic), respectively. Lesion formation of lm mutants was developmentally regulated, and all the mutants showed stunted growth and reduced fertility. Genetic analysis demonstrated that all the mutations were recessive, and five partially fertile mutants (lm4-lm8) were derived from different loci. Mimic lesions occurring on the leaves of lm mutants resulted from cell death as revealed by trypan blue staining. Six of them ( lm3 -lm8 ) exhibited enhanced resistance to five bacterial blight isolates, indicating their wide-spectrum resistance to this pathogen. These results imply that some lesion mimic mutations of rice might be involved in disease resistance signaling pathways,and that isolation of these mutated genes may be useful for elucidating molecular mechanisms of plant disease resistance. Among the mutants, only one mutant, lm6, was preliminarily shown to cosegregate with the inserted T-DNA in its T1 generation, making it feasible to isolate the gene responsible for the phenotype of this mutant.

  7. Augmenting digital displays with computation

    Science.gov (United States)

    Liu, Jing

    As we inevitably step deeper and deeper into a world connected via the Internet, more and more information will be exchanged digitally. Displays are the interface between digital information and each individual. Naturally, one fundamental goal of displays is to reproduce information as realistically as possible since humans still care a lot about what happens in the real world. Human eyes are the receiving end of such information exchange; therefore it is impossible to study displays without studying the human visual system. In fact, the design of displays is rather closely coupled with what human eyes are capable of perceiving. For example, we are less interested in building displays that emit light in the invisible spectrum. This dissertation explores how we can augment displays with computation, which takes both display hardware and the human visual system into consideration. Four novel projects on display technologies are included in this dissertation: First, we propose a software-based approach to driving multiview autostereoscopic displays. Our display algorithm can dynamically assign views to hardware display zones based on multiple observers' current head positions, substantially reducing crosstalk and stereo inversion. Second, we present a dense projector array that creates a seamless 3D viewing experience for multiple viewers. We smoothly interpolate the set of viewer heights and distances on a per-vertex basis across the arrays field of view, reducing image distortion, crosstalk, and artifacts from tracking errors. Third, we propose a method for high dynamic range display calibration that takes into account the variation of the chrominance error over luminance. We propose a data structure for enabling efficient representation and querying of the calibration function, which also allows user-guided balancing between memory consumption and the amount of computation. Fourth, we present user studies that demonstrate that the ˜ 60 Hz critical flicker fusion

  8. Wild Accessions and Mutant Resources

    DEFF Research Database (Denmark)

    Kawaguchi, Masayoshi; Sandal, Niels Nørgaard

    2014-01-01

    Lotus japonicus, Lotus burttii, and Lotus filicaulis are species of Lotus genus that are utilized for molecular genetic analysis such as the construction of a linkage map and QTL analysis. Among them, a number of mutants have been isolated from two wild accessions: L. japonicus Gifu B-129...

  9. Flow cytometry pulse width data enables rapid and sensitive estimation of biomass dry weight in the microalgae Chlamydomonas reinhardtii and Chlorella vulgaris.

    Directory of Open Access Journals (Sweden)

    Maurizio Chioccioli

    Full Text Available Dry weight biomass is an important parameter in algaculture. Direct measurement requires weighing milligram quantities of dried biomass, which is problematic for small volume systems containing few cells, such as laboratory studies and high throughput assays in microwell plates. In these cases indirect methods must be used, inducing measurement artefacts which vary in severity with the cell type and conditions employed. Here, we utilise flow cytometry pulse width data for the estimation of cell density and biomass, using Chlorella vulgaris and Chlamydomonas reinhardtii as model algae and compare it to optical density methods. Measurement of cell concentration by flow cytometry was shown to be more sensitive than optical density at 750 nm (OD750 for monitoring culture growth. However, neither cell concentration nor optical density correlates well to biomass when growth conditions vary. Compared to the growth of C. vulgaris in TAP (tris-acetate-phosphate medium, cells grown in TAP + glucose displayed a slowed cell division rate and a 2-fold increased dry biomass accumulation compared to growth without glucose. This was accompanied by increased cellular volume. Laser scattering characteristics during flow cytometry were used to estimate cell diameters and it was shown that an empirical but nonlinear relationship could be shown between flow cytometric pulse width and dry weight biomass per cell. This relationship could be linearised by the use of hypertonic conditions (1 M NaCl to dehydrate the cells, as shown by density gradient centrifugation. Flow cytometry for biomass estimation is easy to perform, sensitive and offers more comprehensive information than optical density measurements. In addition, periodic flow cytometry measurements can be used to calibrate OD750 measurements for both convenience and accuracy. This approach is particularly useful for small samples and where cellular characteristics, especially cell size, are expected to vary

  10. Nanoformulated cell-penetrating survivin mutant and its dual actions

    Directory of Open Access Journals (Sweden)

    Sriramoju B

    2014-07-01

    Full Text Available Bhasker Sriramoju, Rupinder K Kanwar, Jagat R Kanwar Nanomedicine Laboratory of Immunology and Molecular Biomedical Research (NLIMBR, School of Medicine, Faculty of Health, Deakin University, Geelong, Australia Abstract: In this study, we investigated the differential actions of a dominant-negative survivin mutant (SurR9-C84A against cancerous SK-N-SH neuroblastoma cell lines and differentiated SK-N-SH neurons. In both the cases, the mutant protein displayed dual actions, where its effects were cytotoxic toward cancerous cells and proliferative toward the differentiated neurons. This can be explained by the fact that tumorous (undifferentiated SK-N-SH cells have a high endogenous survivin pool and upon treatment with mutant SuR9-C84A causes forceful survivin expression. These events significantly lowered the microtubule dynamics and stability, eventually leading to apoptosis. In the case of differentiated SK-N-SH neurons that express negligible levels of wild-type survivin, the mutant indistinguishably behaved in a wild-type fashion. It also favored cell-cycle progression, forming the chromosome-passenger complex, and stabilized the microtubule-organizing center. Therefore, mutant SurR9-C84A represents a novel therapeutic with its dual actions (cytotoxic toward tumor cells and protective and proliferative toward neuronal cells, and hence finds potential applications against a variety of neurological disorders. In this study, we also developed a novel poly(lactic-co-glycolic acid nanoparticulate formulation to surmount the hurdles associated with the delivery of SurR9-C84A, thus enhancing its effective therapeutic outcome. Keywords: survivin mutant, neurological disorders, protein therapeutics, inhibitor of apoptosis protein family, poly(lactic-co-glycolic acid

  11. Apoc2 loss-of-function zebrafish mutant as a genetic model of hyperlipidemia

    Directory of Open Access Journals (Sweden)

    Chao Liu

    2015-08-01

    Full Text Available Apolipoprotein C-II (APOC2 is an obligatory activator of lipoprotein lipase. Human patients with APOC2 deficiency display severe hypertriglyceridemia while consuming a normal diet, often manifesting xanthomas, lipemia retinalis and pancreatitis. Hypertriglyceridemia is also an important risk factor for development of cardiovascular disease. Animal models to study hypertriglyceridemia are limited, with no Apoc2-knockout mouse reported. To develop a genetic model of hypertriglyceridemia, we generated an apoc2 mutant zebrafish characterized by the loss of Apoc2 function. apoc2 mutants show decreased plasma lipase activity and display chylomicronemia and severe hypertriglyceridemia, which closely resemble the phenotype observed in human patients with APOC2 deficiency. The hypertriglyceridemia in apoc2 mutants is rescued by injection of plasma from wild-type zebrafish or by injection of a human APOC2 mimetic peptide. Consistent with a previous report of a transient apoc2 knockdown, apoc2 mutant larvae have a minor delay in yolk consumption and angiogenesis. Furthermore, apoc2 mutants fed a normal diet accumulate lipid and lipid-laden macrophages in the vasculature, which resemble early events in the development of human atherosclerotic lesions. In addition, apoc2 mutant embryos show ectopic overgrowth of pancreas. Taken together, our data suggest that the apoc2 mutant zebrafish is a robust and versatile animal model to study hypertriglyceridemia and the mechanisms involved in the pathogenesis of associated human diseases.

  12. X-1 on display

    Science.gov (United States)

    1949-01-01

    A Bell Aircraft Corporation X-1 series aircraft on display at an Open House at NACA Muroc Flight Test Unit or High-Speed Flight Research Station hangar on South Base of Edwards Air Force Base, California. (The precise date of the photo is uncertain, but it is probably before 1948.) The instrumentation that was carried aboard the aircraft to gather data is on display. The aircraft data was recorded on oscillograph film that was read, calibrated, and converted into meaningful parameters for the engineers to evaluate from each research flight. In the background of the photo are several early U.S. jets. These include several Lockheed P-80 Shooting Stars, which were used as chase planes on X-1 flights; two Bell P-59 Airacomets, the first U.S. jet pursuit aircraft (fighter in later parlance); and a prototype Republic XP-84 Thunderjet. There were five versions of the Bell X-1 rocket-powered research aircraft that flew at the NACA High-Speed Flight Research Station, Edwards, California. The bullet-shaped X-1 aircraft were built by Bell Aircraft Corporation, Buffalo, N.Y. for the U.S. Army Air Forces (after 1947, U.S. Air Force) and the National Advisory Committee for Aeronautics (NACA). The X-1 Program was originally designated the XS-1 for eXperimental Sonic. The X-1's mission was to investigate the transonic speed range (speeds from just below to just above the speed of sound) and, if possible, to break the 'sound barrier.' Three different X-1s were built and designated: X-1-1, X-1-2 (later modified to become the X-1E), and X-1-3. The basic X-1 aircraft were flown by a large number of different pilots from 1946 to 1951. The X-1 Program not only proved that humans could go beyond the speed of sound, it reinforced the understanding that technological barriers could be overcome. The X-1s pioneered many structural and aerodynamic advances including extremely thin, yet extremely strong wing sections; supersonic fuselage configurations; control system requirements; powerplant

  13. Laser illuminated flat panel display

    Energy Technology Data Exchange (ETDEWEB)

    Veligdan, J.T.

    1995-12-31

    A 10 inch laser illuminated flat panel Planar Optic Display (POD) screen has been constructed and tested. This POD screen technology is an entirely new concept in display technology. Although the initial display is flat and made of glass, this technology lends itself to applications where a plastic display might be wrapped around the viewer. The display screen is comprised of hundreds of planar optical waveguides where each glass waveguide represents a vertical line of resolution. A black cladding layer, having a lower index of refraction, is placed between each waveguide layer. Since the cladding makes the screen surface black, the contrast is high. The prototype display is 9 inches wide by 5 inches high and approximately I inch thick. A 3 milliwatt HeNe laser is used as the illumination source and a vector scanning technique is employed.

  14. Influence of agglomeration of cerium oxide nanoparticles and speciation of cerium(III) on short term effects to the green algae Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Röhder, Lena A. [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Department of Environmental Toxicology, Dübendorf 8600 (Switzerland); ETH-Zurich, Institute of Biogeochemistry and Pollutant Dynamics, Zürich 8092 (Switzerland); Brandt, Tanja [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Department of Environmental Toxicology, Dübendorf 8600 (Switzerland); Sigg, Laura [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Department of Environmental Toxicology, Dübendorf 8600 (Switzerland); ETH-Zurich, Institute of Biogeochemistry and Pollutant Dynamics, Zürich 8092 (Switzerland); Behra, Renata, E-mail: Renata.behra@eawag.ch [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Department of Environmental Toxicology, Dübendorf 8600 (Switzerland)

    2014-07-01

    Highlights: • Phosphate-dispersed CeO₂ NP did not affect photosynthetic yield in C. reinhardtii. • Agglomerated CeO₂ NP slightly decreased photosynthetic yield. • Cerium(III) was shown to affect photosynthetic yield and intracellular ROS level. • Slight effects of CeO₂ NP were caused by dissolved Ce³⁺ ions present in suspensions. • Wild type and cell wall free mutant of C. reinhardtii showed the same sensitivity. - Abstract: Cerium oxide nanoparticles (CeO₂ NP) are increasingly used in industrial applications and may be released to the aquatic environment. The fate of CeO₂ NP and effects on algae are largely unknown. In this study, the short term effects of CeO₂ NP in two different agglomeration states on the green algae Chlamydomonas reinhardtii were examined. The role of dissolved cerium(III) on toxicity, its speciation and the dissolution of CeO₂ NP were considered. The role of cell wall of C. reinhardtii as a barrier and its influence on the sensitivity to CeO₂ NP and cerium(III) was evaluated by testing both, the wild type and the cell wall free mutant of C. reinhardtii. Characterization showed that CeO₂ NP had a surface charge of ~0 mV at physiological pH and agglomerated in exposure media. Phosphate stabilized CeO₂ NP at pH 7.5 over 24 h. This effect was exploited to test CeO₂ NP dispersed in phosphate with a mean size of 140 nm and agglomerated in absence of phosphate with a mean size of 2000 nm. The level of dissolved cerium(III) in CeO₂ NP suspensions was very low and between 0.1 and 27 nM in all tested media. Exposure of C. reinhardtii to Ce(NO₃)₃ decreased the photosynthetic yield in a concentration dependent manner with EC₅₀ of 7.5 ± 0.84 μM for wild type and EC₅₀ of 6.3 ± 0.53 μM for the cell wall free mutant. The intracellular level of reactive oxygen species (ROS) increased upon exposure to Ce(NO₃)₃ with effective concentrations similar to those inhibiting photosynthesis. The agglomerated Ce

  15. Miniature information displays: primary applications

    Science.gov (United States)

    Alvelda, Phillip; Lewis, Nancy D.

    1998-04-01

    Positioned to replace current liquid crystal display technology in many applications, miniature information displays have evolved to provide several truly portable platforms for the world's growing personal computing and communication needs. The technology and functionality of handheld computer and communicator systems has finally surpassed many of the standards that were originally established for desktop systems. In these new consumer electronics, performance, display size, packaging, power consumption, and cost have always been limiting factors for fabricating genuinely portable devices. The rapidly growing miniature information display manufacturing industry is making it possible to bring a wide range of highly anticipated new products to new markets.

  16. Maintenance Procedure Display: Head Mounted Display (HMD) Evaluations

    Science.gov (United States)

    Whitmore, Milrian; Litaker, Harry L., Jr.; Solem, Jody A.; Holden, Kritina L.; Hoffman, Ronald R.

    2007-01-01

    A viewgraph presentation describing maintenance procedures for head mounted displays is shown. The topics include: 1) Study Goals; 2) Near Eye Displays (HMDs); 3) Design; 4) Phase I-Evaluation Methods; 5) Phase 1 Results; 6) Improved HMD Mounting; 7) Phase 2 -Evaluation Methods; 8) Phase 2 Preliminary Results; and 9) Next Steps.

  17. Comparison between medium-chain acyl-CoA dehydrogenase mutant proteins overexpressed in bacterial and mammalian cells

    DEFF Research Database (Denmark)

    Jensen, T G; Bross, P; Andresen, B S;

    1995-01-01

    Medium-chain acyl-CoA dehydrogenase (MCAD) deficiency is a potentially lethal inherited defect in the beta-oxidation of fatty acids. By comparing the behaviour of five missense MCAD mutant proteins expressed in COS cells and in Escherichia coli, we can define some of these as "pure folding mutants......." Upon expression in E. coli, these mutant proteins produce activity levels in the range of the wild-type enzyme only if the chaperonins GroESL are co-overproduced. When overexpressed in COS cells, the pure folding mutants display enzyme activities comparable to the wild-type enzyme. The results suggest...

  18. Residual plastids of bleached mutants of Euglena gracilis and their effects on the expression of nucleus-encoded genes

    Institute of Scientific and Technical Information of China (English)

    WANG Jiangxin; SHI Zhixin; XU Xudong

    2004-01-01

    Bleached mutants of Euglena gracilis were obtained by treatment with ofloxacin (Ofl)and streptomycin (Sm) respectively. As shown by electron microscopy, the residual plastids contain prothylakoids in an Ofl mutant, and the highly developed and tightly stacked membranous structure found in cells of two Sm mutants. Nine genes of the plastid genome were examined with PCR, showing that ribosomal protein genes and most other plastid genes were lost in all but one Sm mutant. Using differential display and RT-PCR, it was shown that chloroplast degeneration could cause changes in transcription of certain nucleus-encoded genes during heterotrophic growth in darkness.

  19. Flexible Bistable Cholesteric Reflective Displays

    Science.gov (United States)

    Yang, Deng-Ke

    2006-03-01

    Cholesteric liquid crystals (ChLCs) exhibit two stable states at zero field condition-the reflecting planar state and the nonreflecting focal conic state. ChLCs are an excellent candidate for inexpensive and rugged electronic books and papers. This paper will review the display cell structure,materials and drive schemes for flexible bistable cholesteric (Ch) reflective displays.

  20. Nitrogen-Sparing Mechanisms in Chlamydomonas Affect the Transcriptome, the Proteome, and Photosynthetic Metabolism.

    Science.gov (United States)

    Schmollinger, Stefan; Mühlhaus, Timo; Boyle, Nanette R; Blaby, Ian K; Casero, David; Mettler, Tabea; Moseley, Jeffrey L; Kropat, Janette; Sommer, Frederik; Strenkert, Daniela; Hemme, Dorothea; Pellegrini, Matteo; Grossman, Arthur R; Stitt, Mark; Schroda, Michael; Merchant, Sabeeha S

    2014-04-01

    Nitrogen (N) is a key nutrient that limits global primary productivity; hence, N-use efficiency is of compelling interest in agriculture and aquaculture. We used Chlamydomonas reinhardtii as a reference organism for a multicomponent analysis of the N starvation response. In the presence of acetate, respiratory metabolism is prioritized over photosynthesis; consequently, the N-sparing response targets proteins, pigments, and RNAs involved in photosynthesis and chloroplast function over those involved in respiration. Transcripts and proteins of the Calvin-Benson cycle are reduced in N-deficient cells, resulting in the accumulation of cycle metabolic intermediates. Both cytosolic and chloroplast ribosomes are reduced, but via different mechanisms, reflected by rapid changes in abundance of RNAs encoding chloroplast ribosomal proteins but not cytosolic ones. RNAs encoding transporters and enzymes for metabolizing alternative N sources increase in abundance, as is appropriate for the soil environmental niche of C. reinhardtii. Comparison of the N-replete versus N-deplete proteome indicated that abundant proteins with a high N content are reduced in N-starved cells, while the proteins that are increased have lower than average N contents. This sparing mechanism contributes to a lower cellular N/C ratio and suggests an approach for engineering increased N-use efficiency.

  1. Nitrogen-Sparing Mechanisms in Chlamydomonas Affect the Transcriptome, the Proteome, and Photosynthetic Metabolism[W

    Science.gov (United States)

    Schmollinger, Stefan; Mühlhaus, Timo; Boyle, Nanette R.; Blaby, Ian K.; Casero, David; Mettler, Tabea; Moseley, Jeffrey L.; Kropat, Janette; Sommer, Frederik; Strenkert, Daniela; Hemme, Dorothea; Pellegrini, Matteo; Grossman, Arthur R.; Stitt, Mark; Schroda, Michael; Merchant, Sabeeha S.

    2014-01-01

    Nitrogen (N) is a key nutrient that limits global primary productivity; hence, N-use efficiency is of compelling interest in agriculture and aquaculture. We used Chlamydomonas reinhardtii as a reference organism for a multicomponent analysis of the N starvation response. In the presence of acetate, respiratory metabolism is prioritized over photosynthesis; consequently, the N-sparing response targets proteins, pigments, and RNAs involved in photosynthesis and chloroplast function over those involved in respiration. Transcripts and proteins of the Calvin-Benson cycle are reduced in N-deficient cells, resulting in the accumulation of cycle metabolic intermediates. Both cytosolic and chloroplast ribosomes are reduced, but via different mechanisms, reflected by rapid changes in abundance of RNAs encoding chloroplast ribosomal proteins but not cytosolic ones. RNAs encoding transporters and enzymes for metabolizing alternative N sources increase in abundance, as is appropriate for the soil environmental niche of C. reinhardtii. Comparison of the N-replete versus N-deplete proteome indicated that abundant proteins with a high N content are reduced in N-starved cells, while the proteins that are increased have lower than average N contents. This sparing mechanism contributes to a lower cellular N/C ratio and suggests an approach for engineering increased N-use efficiency. PMID:24748044

  2. The Proteome of Copper, Iron, Zinc, and Manganese Micronutrient Deficiency in Chlamydomonas reinhardtii*

    Science.gov (United States)

    Hsieh, Scott I.; Castruita, Madeli; Malasarn, Davin; Urzica, Eugen; Erde, Jonathan; Page, M. Dudley; Yamasaki, Hiroaki; Casero, David; Pellegrini, Matteo; Merchant, Sabeeha S.; Loo, Joseph A.

    2013-01-01

    Trace metals such as copper, iron, zinc, and manganese play important roles in several biochemical processes, including respiration and photosynthesis. Using a label-free, quantitative proteomics strategy (MSE), we examined the effect of deficiencies in these micronutrients on the soluble proteome of Chlamydomonas reinhardtii. We quantified >103 proteins with abundances within a dynamic range of 3 to 4 orders of magnitude and demonstrated statistically significant changes in ∼200 proteins in each metal-deficient growth condition relative to nutrient-replete media. Through analysis of Pearson's coefficient, we also examined the correlation between protein abundance and transcript abundance (as determined via RNA-Seq analysis) and found moderate correlations under all nutritional states. Interestingly, in a subset of transcripts known to significantly change in abundance in metal-replete and metal-deficient conditions, the correlation to protein abundance is much stronger. Examples of new discoveries highlighted in this work include the accumulation of O2 labile, anaerobiosis-related enzymes (Hyd1, Pfr1, and Hcp2) in copper-deficient cells; co-variation of Cgl78/Ycf54 and coprogen oxidase; the loss of various stromal and lumenal photosynthesis-related proteins, including plastocyanin, in iron-limited cells; a large accumulation (from undetectable amounts to over 1,000 zmol/cell) of two COG0523 domain-containing proteins in zinc-deficient cells; and the preservation of photosynthesis proteins in manganese-deficient cells despite known losses in photosynthetic function in this condition. PMID:23065468

  3. Transcriptome Analysis of Manganese-deficient Chlamydomonas reinhardtii Provides Insight on the Chlorophyll Biosynthesis Pathway

    Energy Technology Data Exchange (ETDEWEB)

    Lockhart, Ainsley; Zvenigorodsky, Natasha; Pedraza, Mary Ann; Lindquist, Erika

    2011-08-11

    The biosynthesis of chlorophyll and other tetrapyrroles is a vital but poorly understood process. Recent genomic advances with the unicellular green algae Chlamydomonas reinhardtii have created opportunity to more closely examine the mechanisms of the chlorophyll biosynthesis pathway via transcriptome analysis. Manganese is a nutrient of interest for complex reactions because of its multiple stable oxidation states and role in molecular oxygen coordination. C. reinhardtii was cultured in Manganese-deplete Tris-acetate-phosphate (TAP) media for 24 hours and used to create cDNA libraries for sequencing using Illumina TruSeq technology. Transcriptome analysis provided intriguing insight on possible regulatory mechanisms in the pathway. Evidence supports similarities of GTR (Glutamyl-tRNA synthase) to its Chlorella vulgaris homolog in terms of Mn requirements. Data was also suggestive of Mn-related compensatory up-regulation for pathway proteins CHLH1 (Manganese Chelatase), GUN4 (Magnesium chelatase activating protein), and POR1 (Light-dependent protochlorophyllide reductase). Intriguingly, data suggests possible reciprocal expression of oxygen dependent CPX1 (coproporphyrinogen III oxidase) and oxygen independent CPX2. Further analysis using RT-PCR could provide compelling evidence for several novel regulatory mechanisms in the chlorophyll biosynthesis pathway.

  4. Prospects of using unicellular algae protein in biological life-support systems. [Chlorella, Chlamydomonas, Spirulina, Euglena

    Energy Technology Data Exchange (ETDEWEB)

    Antonyan, A.A.; Abakumova, I.A.; Meleshko, G.I.; Vlasova, T.F.

    The concentration, amino acid composition and biological value of proteins of unicellular algae belonging to various taxonomic groups (Chlorella, Chlamydomonas, Spirulina, Euglena) were investigated. With respect to their characteristics, these algae hold promise as components of biological life-support systems (BLSS). Indices characterizing the protein and biomass quality and biological value were calculated. Such indices as A/E (where A is an essential amino acid and E is the sum total of amino acids), anti-E/T (where anti-E is nitrogen of essential amino acids and T is its sum total), amino acid number, factor of digestibility in vitro were high enough and close to the respective parameters of the reference protein. Animal experiments showed high biological value of the algal biomass and the lack of its toxic or other adverse effects. It is suggested that the differences in the protein composition associated with various algal forms and cultivation conditions can be used to produce balanced diets by varying the portion of each form of the photoautotropic component of BLSS.

  5. Two equilibration pools of chlorophylls in the Photosystem I core antenna of Chlamydomonas reinhardtii.

    Science.gov (United States)

    Gibasiewicz, Krzysztof; Ramesh, V M; Lin, Su; Redding, Kevin; Woodbury, Neal W; Webber, Andrew N

    2007-04-01

    Femtosecond transient absorption spectroscopy was applied for a comparative study of excitation decay in several different Photosystem I (PSI) core preparations from the green alga Chlamydomonas reinhardtii. For PSI cores with a fully interconnected network of chlorophylls, the excitation energy was equilibrated over a pool of chlorophylls absorbing at approximately 683 nm, independent of excitation wavelength [Gibasiewicz et al. J Phys Chem B 105:11498-11506, 2001; J Phys Chem B 106:6322-6330, 2002]. In preparations with impaired connectivity between chlorophylls, we have found that the spectrum of chlorophylls connected to the reaction center (i.e., with approximately 20 ps decay time) over which the excitation is equilibrated becomes excitation-wavelength-dependent. Excitation at 670 nm is finally equilibrated over chlorophylls absorbing at approximately 675 nm, whereas excitation at 695 nm or 700 nm is equilibrated over chlorophylls absorbing at approximately 683 nm. This indicates that in the vicinity of the reaction center there are two spectrally different and spatially separated pools of chlorophylls that are equally capable of effective excitation energy transfer to the reaction center. We propose that they are related to the two groups of central PSI core chlorophylls lying on the opposite sides of reaction center.

  6. Controlling expression of genes in the unicellular alga Chlamydomonas reinhardtii with a vitamin-repressible riboswitch.

    Science.gov (United States)

    Ramundo, Silvia; Rochaix, Jean-David

    2015-01-01

    Chloroplast genomes of land plants and algae contain generally between 100 and 150 genes. These genes are involved in plastid gene expression and photosynthesis and in various other tasks. The function of some chloroplast genes is still unknown and some of them appear to be essential for growth and survival. Repressible and reversible expression systems are highly desirable for functional and biochemical characterization of these genes. We have developed a genetic tool that allows one to regulate the expression of any coding sequence in the chloroplast genome of the unicellular alga Chlamydomonas reinhardtii. Our system is based on vitamin-regulated expression of the nucleus-encoded chloroplast Nac2 protein, which is specifically required for the expression of any plastid gene fused to the psbD 5'UTR. With this approach, expression of the Nac2 gene in the nucleus and, in turn, that of the chosen chloroplast gene artificially driven by the psbD 5'UTR, is controlled by the MetE promoter and Thi4 riboswitch, which can be inactivated in a reversible way by supplying vitamin B12 and thiamine to the growth medium, respectively. This system opens interesting possibilities for studying the assembly and turnover of chloroplast multiprotein complexes such as the photosystems, the ribosome, and the RNA polymerase. It also provides a way to overcome the toxicity often associated with the expression of proteins of biotechnological interest in the chloroplast.

  7. Process development for hydrogen production with Chlamydomonas reinhardtii based on growth and product formation kinetics.

    Science.gov (United States)

    Lehr, Florian; Morweiser, Michael; Rosello Sastre, Rosa; Kruse, Olaf; Posten, Clemens

    2012-11-30

    Certain strains of microalgae are long known to produce hydrogen under anaerobic conditions. In Chlamydomonas reinhardtii the oxygen-sensitive hydrogenase enzyme recombines electrons from the chloroplast electron transport chain with protons to form molecular hydrogen directly inside the chloroplast. A sustained hydrogen production can be obtained under low sulfur conditions in C. reinhardtii, reducing the net oxygen evolution by reducing the photosystem II activity and thereby overcoming the inhibition of the hydrogenases. The development of specially adapted hydrogen production strains led to higher yields and optimized biological process preconditions. So far sustainable hydrogen production required a complete exchange of the growth medium to establish sulfur-deprived conditions after biomass growth. In this work we demonstrate the transition from the biomass growth phase to the hydrogen production phase in a single batch culture only by exact dosage of sulfur. This eliminates the elaborate and energy intensive solid-liquid separation step and establishes a process strategy to proceed further versus large scale production. This strategy has been applied to determine light dependent biomass growth and hydrogen production kinetics to assess the potential of H₂ production with C. reinhardtii as a basis for scale up and further process optimization.

  8. Antagonistic and synergistic effects of light irradiation on the effects of copper on Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Cheloni, Giulia; Cosio, Claudia; Slaveykova, Vera I., E-mail: vera.slaveykova@unige.ch

    2014-10-15

    Highlights: • Light intensity and spectral composition affect Cu uptake and effects to C. reinhardtii. • High light (HL) reduced Cu effect on growth inhibition, oxidative stress and damage. • HL in combination with Cu up-regulated genes involved in the antioxidant responses. • HL with increased UVB radiation exacerbated Cu uptake and Cu-induced toxic effects. - Abstract: The present study showed the important role of light intensity and spectral composition on Cu uptake and effects on green alga Chlamydomonas reinhardtii. High-intenisty light (HL) increased cellular Cu concentrations, but mitigated the Cu-induced decrease in chlorophyll fluorescence, oxidative stress and lipid peroxidation at high Cu concentrations, indicating that Cu and HL interact in an antagonistic manner. HL up-regulated the transcription of genes involved in the antioxidant response in C. reinhardtii and thus reduced the oxidative stress upon exposure to Cu and HL. Combined exposure to Cu and UVBR resulted in an increase of cellular Cu contents and caused severe oxidative damage to the cells. The observed effects were higher than the sum of the effects corresponding to exposure to UVBR or Cu alone suggesting a synergistic interaction.

  9. Relief of arsenate toxicity by Cd-stimulated phytochelatin synthesis in the green alga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Kobayashi, Isao; Fujiwara, Shoko; Saegusa, Hirotaka; Inouhe, Masahiro; Matsumoto, Hiroko; Tsuzuki, Mikio

    2006-01-01

    In most photosynthetic organisms, inorganic arsenic taken up into the cells inhibits photosynthesis and cellular growth. In a green alga, Chlamydomonas reinhardtii, 0.5 mM arsenate inhibited photosynthesis almost completely within 30 min. However, in cells acclimated with a sublethal concentration (0.05 to 0.1 mM) of Cd, the inhibition of photosynthesis at 30 min after the addition of arsenate was relieved by more than 50%. The concentrations of arsenic incorporated into the cells were not significantly different between the Cd-acclimated and the non-acclimated cells. The Cd-acclimated cells accumulated Cd and synthesized phytochelatin (PC) peptides, which are known to play an important role in detoxification of heavy metals in plants. By the addition of an inhibitor of glutathione (an intermediate in the PC biosynthetic pathway) biosynthesis, buthionine sulfoximine, cells lost not only Cd tolerance but also arsenate tolerance. These results suggest that glutathione and/or PCs synthesized in Cd-acclimated cells are involved in mechanisms of arsenate tolerance.

  10. Integrated quantitative analysis of nitrogen stress response in Chlamydomonas reinhardtii using metabolite and protein profiling.

    Science.gov (United States)

    Wase, Nishikant; Black, Paul N; Stanley, Bruce A; DiRusso, Concetta C

    2014-03-01

    Nitrogen starvation induces a global stress response in microalgae that results in the accumulation of lipids as a potential source of biofuel. Using GC-MS-based metabolite and iTRAQ-labeled protein profiling, we examined and correlated the metabolic and proteomic response of Chlamydomonas reinhardtii under nitrogen stress. Key amino acids and metabolites involved in nitrogen sparing pathways, methyl group transfer reactions, and energy production were decreased in abundance, whereas certain fatty acids, citric acid, methionine, citramalic acid, triethanolamine, nicotianamine, trehalose, and sorbitol were increased in abundance. Proteins involved in nitrogen assimilation, amino acid metabolism, oxidative phosphorylation, glycolysis, TCA cycle, starch, and lipid metabolism were elevated compared with nonstressed cultures. In contrast, the enzymes of the glyoxylate cycle, one carbon metabolism, pentose phosphate pathway, the Calvin cycle, photosynthetic and light harvesting complex, and ribosomes were reduced. A noteworthy observation was that citrate accumulated during nitrogen stress coordinate with alterations in the enzymes that produce or utilize this metabolite, demonstrating the value of comparing protein and metabolite profiles to understand complex patterns of metabolic flow. Thus, the current study provides unique insight into the global metabolic adjustments leading to lipid storage during N starvation for application toward advanced biofuel production technologies.

  11. Chlamydomonas as a model for biofuels and bio-products production.

    Science.gov (United States)

    Scranton, Melissa A; Ostrand, Joseph T; Fields, Francis J; Mayfield, Stephen P

    2015-05-01

    Developing renewable energy sources is critical to maintaining the economic growth of the planet while protecting the environment. First generation biofuels focused on food crops like corn and sugarcane for ethanol production, and soybean and palm for biodiesel production. Second generation biofuels based on cellulosic ethanol produced from terrestrial plants, has received extensive funding and recently pilot facilities have been commissioned, but to date output of fuels from these sources has fallen well short of what is needed. Recent research and pilot demonstrations have highlighted the potential of algae as one of the most promising sources of sustainable liquid transportation fuels. Algae have also been established as unique biofactories for industrial, therapeutic, and nutraceutical co-products. Chlamydomonas reinhardtii's long established role in the field of basic research in green algae has paved the way for understanding algal metabolism and developing genetic engineering protocols. These tools are now being utilized in C. reinhardtii and in other algal species for the development of strains to maximize biofuels and bio-products yields from the lab to the field.

  12. Advances in the biotechnology of hydrogen production with the microalga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Torzillo, Giuseppe; Scoma, Alberto; Faraloni, Cecilia; Giannelli, Luca

    2015-01-01

    Biological hydrogen production is being evaluated for use as a fuel, since it is a promising substitute for carbonaceous fuels owing to its high conversion efficiency and high specific energy content. The basic advantages of biological hydrogen production over other "green" energy sources are that it does not compete for agricultural land use, and it does not pollute, as water is the only by-product of the combustion. These characteristics make hydrogen a suitable fuel for the future. Among several biotechnological approaches, photobiological hydrogen production carried out by green microalgae has been intensively investigated in recent years. A select group of photosynthetic organisms has evolved the ability to harness light energy to drive hydrogen gas production from water. Of these, the microalga Chlamydomonas reinhardtii is considered one of the most promising eukaryotic H2 producers. In this model microorganism, light energy, H2O and H2 are linked by two excellent catalysts, the photosystem 2 (PSII) and the [FeFe]-hydrogenase, in a pathway usually referred to as direct biophotolysis. This review summarizes the main advances made over the past decade as an outcome of the discovery of the sulfur-deprivation process. Both the scientific and technical barriers that need to be overcome before H2 photoproduction can be scaled up to an industrial level are examined. Actual and theoretical limits of the efficiency of the process are also discussed. Particular emphasis is placed on algal biohydrogen production outdoors, and guidelines for an optimal photobioreactor design are suggested.

  13. Recombination and Heterologous Expression of Allophycocyanin Gene in the Chloroplast of Chlamydomonas reinhardtii

    Institute of Scientific and Technical Information of China (English)

    Zhong-Liang SU; Kai-Xian QIAN; Cong-Ping TAN; Chun-Xiao MENG; Song QIN

    2005-01-01

    Heterogeneous expression of multiple genes in the nucleus of transgenic plants requires the introduction of an individual gene and the subsequent backcross to reconstitute multi-subunit proteins or metabolic pathways. In order to accomplish the expression of multiple genes in a single transformation event, we inserted both large and small subunits of allophycocyanin gene (apcA and apcB) into Chlamydomonas reinhardtii chloroplast expression vector, resulting in papc-S. The constructed vector was then introduced into the chloroplast of C. reinhardtii by micro-particle bombardment. Polymerase chain reaction and Southern blot analysis revealed that the two genes had integrated into the chloroplast genome. Western blot and enzyme-linked immunosorbent assay showed that the two genes from the prokaryotic cyanobacteria could be correctly expressed in the chloroplasts of C. reinhardtii. The expressed foreign protein in transformants accounted for about 2%-3% of total soluble proteins. These findings pave the way to the reconstitution of multi-subunit proteins or metabolic pathways in transgenic C. reinhardtii chloroplasts in a single transformation event.

  14. Linear systems analysis of the ciliary steering behavior associated with negative-phototaxis in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Josef, Keith; Saranak, Jureepan; Foster, Kenneth W

    2006-12-01

    In response to light stimulation Chlamydomonas reinhardtii changes the beating frequency, beating pattern, and beating synchrony of the trans and cis cilia to steer the freely-swimming cell relative to light sources. To understand the cell steering behavior the impulse responses of the beating frequency and stroke velocity of each cilium have been obtained with high temporal resolution on cells held with a micropipette. Interestingly the response of each cilium is quite different. The trans cilium responds with less delay than the cis cilium for both beating frequency and stroke velocity. For light stimulation at 2 Hz, the critical cell-rotation frequency, both responses of the trans and cis cilia are about 180 degrees out of phase. The trans-cilium beating frequency response peaks at a stimulus frequency of 5-6 Hz, higher than the cis at 1-2 Hz. The stroke velocities of the trans and cis cilia have the same stimulus-frequency response (2 Hz), but the trans cilium has a shorter delay than the cis. The times to maximum response are much shorter than the time for a rotation of the cell. The use of two different approaches that enable the trans cilium to respond ahead of the cis for both the beating frequency and stroke velocity responses suggests the importance of both responses to phototaxis. Internal cell processing responsible for the time course of the responses is proposed.

  15. Enhancement of lipid production and fatty acid profiling in Chlamydomonas reinhardtii, CC1010 for biodiesel production.

    Science.gov (United States)

    Karpagam, R; Preeti, R; Ashokkumar, B; Varalakshmi, P

    2015-11-01

    Lipid from microalgae is one of the putative oil resources to facilitate the biodiesel production during this era of energy dissipation and environmental pollution. In this study, the key parameters such as biomass productivity, lipid productivity and lipid content were evaluated at the early stationary phase of Chlamydomonas reinhardtii, CC1010 cultivated in nutrient starved (nitrogen, phosphorous), glucose (0.05%, 0.1%, 0.15% and 0.2%) and vitamin B12 supplementation (0.001%, 0.002% and 0.003%) in Tris-Acetate-Phosphate (TAP) medium. The lipid content in nitrogen starved media was 61% which is 2.34 folds higher than nutrient sufficient TAP medium. Glucose supplementation has lead to proportional increase in biomass productivity with the increasing concentration of glucose whereas vitamin B12 supplementations had not shown any influence in lipid and biomass production. Further, fatty acid methyl ester (FAME) profiling of C. reinhardtii, CC 1010 has revealed more than 80% of total SFA (saturated fatty acid) and MUFA (mono unsaturated fatty acid) content. Quality checking parameters of biodiesel like cetane number, saponification value, iodine number and degree of unsaturation were analyzed and the biodiesel fuel properties were found to be appropriate as per the international standards, EN 14214 and ASTM D6751. Conclusively, among all the treatments, nitrogen starvation with 0.1% glucose supplementation had yielded high lipid content in C. reinhardtii, CC 1010.

  16. Ciprofloxacin toxicity and its co-metabolic removal by a freshwater microalga Chlamydomonas mexicana.

    Science.gov (United States)

    Xiong, Jiu-Qiang; Kurade, Mayur B; Kim, Jung Rae; Roh, Hyun-Seog; Jeon, Byong-Hun

    2017-02-05

    This study evaluated the toxicity and cellular stresses of ciprofloxacin (CIP) and its co-metabolic removal in a freshwater microalga Chlamydomonas mexicana. The toxicological effects of CIP on C. mexicana were assessed by studying the growth and biochemical characteristics of the microalga including total chlorophyll, carotenoid content, malondialdehyde (MDA) and superoxide dismutase (SOD) activity. The calculated effective concentration (EC50) of CIP on C. mexicana was 65±4mgL(-1) at 96h. The growth of C. mexicana was significantly inhibited at increased concentrations of CIP, showing 36±1, 75±3. and 88±3% inhibition at 40, 60 and 100mgL(-1) CIP, respectively, compared to the control after 11days of cultivation. The total chlorophyll, carotenoid, MDA and SOD activity were significantly increased as a result of relatively high concentrations of CIP stress. C. mexicana showed 13±1% removal of CIP (2mgL(-1)) after 11days of cultivation; however, the addition of an electron donor (sodium acetate, 4gL(-1)) highly enhanced the removal of CIP (2mgL(-1)) by>3-fold after 11days. Kinetic studies showed that removal of CIP followed a first-order model (R(2) 0.94-0.97) with the apparent rate constants (k) ranging from 0.0121 to 0.079 d(-1).

  17. Influence of sulphate on the reduction of cadmium toxicity in the microalga Chlamydomonas moewusii.

    Science.gov (United States)

    Mera, Roi; Torres, Enrique; Abalde, Julio

    2016-06-01

    Cadmium is considered as one of the most hazardous metals for living organism and ecosystems. Environmental factors play an important role since they alter the toxicity of metals by varying the bioavailability of these elements for the organisms. The aim of the present study was to investigate, using the freshwater microalga Chlamydomonas moewusii, the existence of an interaction between cadmium and sulphate as a factor that varied the toxicity of this metal. Different cell parameters such as cell growth, content of chlorophylls and biosynthesis of phytochelatins (PCs) were determined. A two-way ANOVA showed that the interaction had a significant effect size of 21% (pmicroalga and around of a 6% on the content of chlorophylls/cell. The effect of this inhibition was that when the concentration of sulphate increased, a lower toxic effect of cadmium on the growth and on the content of chlorophylls was observed. In addition, the increase of sulphate concentration allowed the biosynthesis of a higher amount of PCs and/or PCs with higher chain length. This higher biosynthesis was responsible for the reduction of the toxic effect of cadmium and explained the interaction.

  18. Toxicity of atrazine and its bioaccumulation and biodegradation in a green microalga, Chlamydomonas mexicana.

    Science.gov (United States)

    Kabra, Akhil N; Ji, Min-Kyu; Choi, Jaewon; Kim, Jung Rae; Govindwar, Sanjay P; Jeon, Byong-Hun

    2014-11-01

    This study evaluated the toxicity of herbicide atrazine, along with its bioaccumulation and biodegradation in the green microalga Chlamydomonas mexicana. At low concentration (10 μg L(-1)), atrazine had no profound effect on the microalga, while higher concentrations (25, 50, and 100 μg L(-1)) imposed toxicity, leading to inhibition of cell growth and chlorophyll a accumulation by 22 %, 33 %, and 36 %, and 13 %, 24 %, and 27 %, respectively. Atrazine 96-h EC50 for C. mexicana was estimated to be 33 μg L(-1). Microalga showed a capability to accumulate atrazine in the cell and to biodegrade the cell-accumulated atrazine resulting in 14-36 % atrazine degradation at 10-100 μg L(-1). Increasing atrazine concentration decreased the total fatty acids (from 102 to 75 mg g(-1)) and increased the unsaturated fatty acid content in the microalga. Carbohydrate content increased gradually with the increase in atrazine concentration up to 15 %. This study shows that C. mexicana has the capability to degrade atrazine and can be employed for the remediation of atrazine-contaminated streams.

  19. Towards elucidation of the toxic mechanism of copper on the model green alga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Jiang, Yongguang; Zhu, Yanli; Hu, Zhangli; Lei, Anping; Wang, Jiangxin

    2016-09-01

    Toxic effects of copper on aquatic organisms in polluted water bodies have garnered particular attention in recent years. Microalgae play an important role in aquatic ecosystems, and they are sensitive to heavy metal pollution. Thus, it is important to clarify the mechanism of copper toxicity first for ecotoxicology studies. In this study, the physiological, biochemical and gene expression characteristics of a model green microalga, Chlamydomonas reinhardtii, with 0, 50, 150 and 250 μM copper treatments were investigated. The response of C. reinhardtii to copper stress was significantly shown at a dose dependent manner. Inhibition of cell growth and variation of total chlorophyll content were observed with copper treatments. The maximum photochemical efficiency of PSII, actual photochemical efficiency of PSII and photochemical quenching value decreased in the 250 μM copper treatment with minimum values equal to 28, 24 and 60 % of the control values respectively. The content of lipid peroxidation biomarker malondialdehyde with copper treatments increased with a maximum value sevenfold higher than the control value. Inhibition of cell growth and photosynthesis was ascribed to peroxidation of membrane lipids. The glutathione content and activities of antioxidant enzymes, glutathione S-transferase, glutathione peroxidase, superoxide dismutase and peroxidase were induced by copper. Interestingly, the expression of antioxidant genes and the photosynthetic gene decreased in most copper treatments. In conclusion, oxidative stress caused by production of excess reactive oxidative species might be the major mechanism of copper toxicity on C. reinhardtii.

  20. Phytotoxicity of 15 common pharmaceuticals on the germination of Lactuca sativa and photosynthesis of Chlamydomonas reinhardtii.

    Science.gov (United States)

    Pino, Ma Rosa; Muñiz, Selene; Val, Jonatan; Navarro, Enrique

    2016-11-01

    Pharmaceuticals reach terrestrial environments through the application of treated wastewaters and biosolids to agricultural soils. We have investigated the toxicity of 15 common pharmaceuticals, classified as nonsteroidal anti-inflammatory drugs (NSAIDs), blood lipid-lowering agents, β-blockers and antibiotics, in two photosynthetic organisms. Twelve pharmaceuticals caused inhibitory effects on the radicle and hypocotyl elongation of Lactuca sativa seeds. The EC50 values obtained were in the range of 170-5656 mg L(-1) in the case of the radicle and 188-4558 mg L(-1) for the hypocotyl. Propranolol was the most toxic drug for both root and hypocotyl elongation, followed by the NSAIDs, then gemfibrozil and tetracycline. Other effects, such as root necrosis, inhibition of root growth and curly hairs, were detected. However, even at the highest concentrations tested (3000 mg L(-1)), seed germination was not affected. NSAIDs decreased the photosynthetic yield of Chlamydomonas reinhardtii, but only salicylic acid showed EC50 values below 1000 mg L(-1). The first effects detected at low concentrations, together with the concentrations found in environmental samples, indicate that the use of biosolids and wastewaters containing pharmaceuticals should be regulated and their compositions assessed in order to prevent medium- and long-term impacts on agricultural soils and crops.

  1. Light/electricity conversion by defined cocultures of Chlamydomonas and Geobacter.

    Science.gov (United States)

    Nishio, Koichi; Hashimoto, Kazuhito; Watanabe, Kazuya

    2013-04-01

    Biological energy-conversion systems are attractive in terms of their self-organizing and self-sustaining properties and are expected to be applied towards environmentally friendly bioenergy processes. Recent studies have demonstrated that sustainable light/electricity-conversion systems, termed microbial solar cells (MSCs), can be constructed using naturally occurring microbial communities. To better understand the energy-conversion mechanisms in microbial communities, the present study attempted to construct model MSCs comprised of defined cocultures of a green alga, Chlamydomonas reinhardtii, and an iron-reducing bacterium, Geobacter sulfurreducens, and examined their metabolism and interactions in MSCs. When MSC bioreactors were inoculated with these microbes and irradiated on a 12-h light/dark cycle, periodic current was generated in the dark with energy-conversion efficiencies of 0.1%. Metabolite analyses revealed that G. sulfurreducens generated current by oxidizing formate that was produced by C. reinhardtii in the dark. These results demonstrate that the light/electricity conversion occurs via syntrophic interactions between phototrophs and electricity-generating bacteria. Based on the results and data in literatures, it is estimated that the excretion of organics by the phototroph was the bottleneck step in the syntrophic light/electricity conversion. We also discuss differences between natural-community and defined-coculture MSCs.

  2. An omics based assessment of cadmium toxicity in the green alga Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Jamers, An; Blust, Ronny; De Coen, Wim [Laboratory for Ecophysiology, Biochemistry and Toxicology, Department of Biology, University of Antwerp, Groenenborgerlaan 171, 2020 Antwerp (Belgium); Griffin, Julian L. [Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 2QA (United Kingdom); Jones, Oliver A.H., E-mail: oliver.jones@rmit.edu.au [School of Applied Sciences, RMIT University, GPO Box 2476, Melbourne, VIC 3001 (Australia)

    2013-01-15

    The effects of cadmium were assessed in the freshwater alga Chlamydomonas reinhardtii. Algae were exposed to concentrations of 0, 8.1 or 114.8 {mu}M of cadmium and growth rates, gene transcription and metabolite profiles were examined after 48 and 72 h of exposure. In algae exposed to 8.1 {mu}M Cd, several genes were differentially transcribed after 48 h but no adverse growth related effects were detected. A transient effect on both gene transcription patterns and metabolite profiles could be discerned after 48 h of exposure but the majority of these changes disappeared after 72 h. In contrast, all effects were more pronounced at the 114.8 {mu}M cadmium exposure. Here growth was clearly reduced and transcription of a large number of genes involved in oxidative stress defense mechanisms was differentially increased. Metabolites involved in the glutathione synthesis pathway (an important antioxidant defense) were also affected but the effects of cadmium were found to be more pronounced at the transcript level than in the metabolome, suggesting that the former exhibits greater sensitivity toward cadmium exposure.

  3. A simple and non-invasive method for nuclear transformation of intact-walled Chlamydomonas reinhardtii.

    Directory of Open Access Journals (Sweden)

    Sora Kim

    Full Text Available Genetic engineering in microalgae is gaining attraction but nuclear transformation methods available so far are either inefficient or require special equipment. In this study, we employ positively charged nanoparticles, 3-aminopropyl-functionalized magnesium phyllosilicate (aminoclay, approximate unit cell composition of [H2N(CH23]8Si8Mg6O12(OH4, for nuclear transformation into eukaryotic microalgae. TEM and EDX analysis of the process of transformation reveals that aminoclay coats negatively-charged DNA biomolecules and forms a self-assembled hybrid nanostructure. Subsequently, when this nanostructure is mixed with microalgal cells and plated onto selective agar plates with high friction force, cell wall is disrupted facilitating delivery of plasmid DNA into the cell and ultimately to the nucleus. This method is not only simple, inexpensive, and non-toxic to cells but also provides efficient transformation (5.03×10(2 transformants/µg DNA, second only to electroporation which needs advanced instrumentation. We present optimized parameters for efficient transformation including pre-treatment, friction force, concentration of foreign DNA/aminoclay, and plasticity of agar plates. It is also confirmed the successful integration and stable expression of foreign gene in Chlamydomonas reinhardtii through molecular methods.

  4. Characterization of a Native Algae Species Chlamydomonas debaryana: Strain Selection, Bioremediation Ability, and Lipid Characterization

    Directory of Open Access Journals (Sweden)

    Bo Zhang

    2014-08-01

    Full Text Available Native microalgal species may offer a favorable combination of both wastewater treatment and biofuel production. In this research, a green microalgae, Chlamydomonas debaryana, was isolated from a local lagoon, screened for its lipid content using flow cytometry, and further identified with microscopic observations and DNA sequence analysis. When using swine wastewater as a medium, the biomass yields were between 0.6 and 1.62 g/L, giving a median value of 1.11 g/L. By increasing mass transfer rates and providing sufficient light intensity, the microalgal growth was intrinsically enhanced. The growth of C. debaryana reduced most nutritional contents of the wastewater except iron. When combining the microalgal growth and nutrient removal, C. debaryana was able to utilize 1.3 to 1.6×103 mg COD (chemical oxygen demand/g biomass, 55 to 90 ppm ammonia/g biomass, and 48 to 89 ppm phosphorous/g biomass, The lipid content of C. debaryana was 19.9 ± 4.3% of cell dry weight. The transesterified microalgal oil mostly consisted of 14 kinds of fatty acids, ranging from C5 to C22, which can be refined into renewable jet fuel or used as sources of omega-3 and omega-6 fatty acids.

  5. Trophic transfer of gold nanoparticles from Euglena gracilis or Chlamydomonas reinhardtii to Daphnia magna.

    Science.gov (United States)

    Lee, Woo-Mi; Yoon, Sung-Ji; Shin, Yu-Jin; An, Youn-Joo

    2015-06-01

    Understanding the trophic transfer of nanoparticles (NPs) is important because NPs are small enough to easily penetrate into organisms. In this study, we evaluated the trophic transfer of gold NPs (AuNPs) within the aquatic food chain. We observed AuNPs transfer from 2 species of primary producers (Chlamydomonas reinhardtii or Euglena gracilis) to the primary consumer (Daphnia magna). Also, bioaccumulation of AuNPs in E. gracilis was higher than that in C. reinhardtii. The reasons for the difference in Au accumulation may be the physical structure of these organisms, and the surface area that is available for interaction with NPs. C. reinhardtii has a cell wall that may act as a barrier to the penetration of NPs. The size of E. gracilis is larger than that of C. reinhardtii. This study demonstrates the trophic transfer of AuNPs from a general producer to a consumer in an aquatic environment.

  6. A small multifunctional pentatricopeptide repeat protein in the chloroplast of Chlamydomonas reinhardtii.

    Science.gov (United States)

    Jalal, Abdullah; Schwarz, Christian; Schmitz-Linneweber, Christian; Vallon, Olivier; Nickelsen, Jörg; Bohne, Alexandra-Viola

    2015-03-01

    Organellar biogenesis is mainly regulated by nucleus-encoded factors, which act on various steps of gene expression including RNA editing, processing, splicing, stabilization, and translation initiation. Among these regulatory factors, pentatricopeptide repeat (PPR) proteins form the largest family of RNA binding proteins, with hundreds of members in flowering plants. In striking contrast, the genome of the unicellular green alga Chlamydomonas reinhardtii encodes only 14 such proteins. In this study, we analyzed PPR7, the smallest and most highly expressed PPR protein in C. reinhardtii. Green fluorescent protein-based localization and gel-filtration analysis revealed that PPR7 forms a part of a high-molecular-weight ribonucleoprotein complex in the chloroplast stroma. RIP-chip analysis of PPR7-bound RNAs demonstrated that the protein associates with a diverse set of chloroplast transcripts in vivo, i.e. rrnS, psbH, rpoC2, rbcL, atpA, cemA-atpH, tscA, and atpI-psaJ. Furthermore, the investigation of PPR7 RNAi strains revealed that depletion of PPR7 results in a light-sensitive phenotype, accompanied by altered levels of its target RNAs that are compatible with the defects in their maturation or stabilization. PPR7 is thus an unusual type of small multifunctional PPR protein, which interacts, probably in conjunction with other RNA binding proteins, with numerous target RNAs to promote a variety of post-transcriptional events.

  7. Identification of regulatory network hubs that control lipid metabolism in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Gargouri, Mahmoud; Park, Jeong-Jin; Holguin, F Omar; Kim, Min-Jeong; Wang, Hongxia; Deshpande, Rahul R; Shachar-Hill, Yair; Hicks, Leslie M; Gang, David R

    2015-08-01

    Microalgae-based biofuels are promising sources of alternative energy, but improvements throughout the production process are required to establish them as economically feasible. One of the most influential improvements would be a significant increase in lipid yields, which could be achieved by altering the regulation of lipid biosynthesis and accumulation. Chlamydomonas reinhardtii accumulates oil (triacylglycerols, TAG) in response to nitrogen (N) deprivation. Although a few important regulatory genes have been identified that are involved in controlling this process, a global understanding of the larger regulatory network has not been developed. In order to uncover this network in this species, a combined omics (transcriptomic, proteomic and metabolomic) analysis was applied to cells grown in a time course experiment after a shift from N-replete to N-depleted conditions. Changes in transcript and protein levels of 414 predicted transcription factors (TFs) and transcriptional regulators (TRs) were monitored relative to other genes. The TF and TR genes were thus classified by two separate measures: up-regulated versus down-regulated and early response versus late response relative to two phases of polar lipid synthesis (before and after TAG biosynthesis initiation). Lipidomic and primary metabolite profiling generated compound accumulation levels that were integrated with the transcript dataset and TF profiling to produce a transcriptional regulatory network. Evaluation of this proposed regulatory network led to the identification of several regulatory hubs that control many aspects of cellular metabolism, from N assimilation and metabolism, to central metabolism, photosynthesis and lipid metabolism.

  8. The response of Chlamydomonas reinhardtii to nitrogen deprivation: a systems biology analysis.

    Science.gov (United States)

    Park, Jeong-Jin; Wang, Hongxia; Gargouri, Mahmoud; Deshpande, Rahul R; Skepper, Jeremy N; Holguin, F Omar; Juergens, Matthew T; Shachar-Hill, Yair; Hicks, Leslie M; Gang, David R

    2015-02-01

    Drastic alteration in macronutrients causes large changes in gene expression in the photosynthetic unicellular alga Chlamydomonas reinhardtii. Preliminary data suggested that cells follow a biphasic response to this change hinging on the initiation of lipid accumulation, and we hypothesized that drastic repatterning of metabolism also followed this biphasic modality. To test this hypothesis, transcriptomic, proteomic, and metabolite changes that occur under nitrogen (N) deprivation were analyzed. Eight sampling times were selected covering the progressive slowing of growth and induction of oil synthesis between 4 and 6 h after N deprivation. Results of the combined, systems-level investigation indicated that C. reinhardtii cells sense and respond on a large scale within 30 min to a switch to N-deprived conditions turning on a largely gluconeogenic metabolic state, which then transitions to a glycolytic stage between 4 and 6 h after N depletion. This nitrogen-sensing system is transduced to carbon- and nitrogen-responsive pathways, leading to down-regulation of carbon assimilation and chlorophyll biosynthesis, and an increase in nitrogen metabolism and lipid biosynthesis. For example, the expression of nearly all the enzymes for assimilating nitrogen from ammonium, nitrate, nitrite, urea, formamide/acetamide, purines, pyrimidines, polyamines, amino acids and proteins increased significantly. Although arginine biosynthesis enzymes were also rapidly up-regulated, arginine pool size changes and isotopic labeling results indicated no increased flux through this pathway.

  9. Effect of aluminum on cellular division and photosynthetic electron transport in Euglena gracilis and Chlamydomonas acidophila.

    Science.gov (United States)

    Perreault, François; Dewez, David; Fortin, Claude; Juneau, Philippe; Diallo, Amirou; Popovic, Radovan

    2010-04-01

    The present study investigated aluminum's effect on cellular division and the photosynthetic processes in Euglena gracilis and Chlamydomonas acidophila at pH 3.0, at which Al is present mostly as Al(3+), AlSO(4) (+), and Al(SO(4))(2) (-). These algal species were exposed to 100, 188, and 740 microM Al, and after 24 h cell-bound Al was significantly different from control only for the highest concentration tested. However, very different effects of Al on algal cellular division, biomass per cell, and photosynthetic activity were found. Aluminum stimulated cell division but decreased at some level biomass per cell in C. acidophila. Primary photochemistry of photosynthesis, as Photosystem II quantum yield, and energy dissipation via nonphotochemical activity were slightly affected. However, for E. gracilis, under the same conditions, Al did not show a stimulating effect on cellular division or photosynthetic activity. Primary photochemical activity was diminished, and energy dissipation via nonphotochemical pathways was strongly increased. Therefore, when Al is highly available in aquatic ecosystems, these effects may indicate very different response mechanisms that are dependent on algal species.

  10. Alleviation of copper-induced oxidative damage in Chlamydomonas reinhardtii by carbon monoxide.

    Science.gov (United States)

    Zheng, Qi; Meng, Qian; Wei, Yuan Yuan; Yang, Zhi Min

    2011-08-01

    Carbon monoxide (CO) is an endogenous gaseous molecule in plants and animals. Recent studies have shown that it is one of the most essential cellular components regulating many aspects of plant growth and development. However, whether CO regulates the green algae adaptive response to heavy metal toxicity is unknown. The present study investigated the role of CO in regulating Cu-induced oxidative stress in eukaryotic algae Chlamydomonas reinhardtii. Cells pretreated with 5 μM CO for 30 min and followed by exposure to 5 μM Cu(II) for 4 days showed attenuated toxicity. The CO-improved growth of algae was correlated with reduced lipid peroxidation and increased chlorophyll accumulation. The beneficial effect of CO was confirmed by histochemical staining with reactive oxygen species. Further, treatment with 5 μM CO increased the activity of catalase with Cu. However, a reduced superoxide dismutase activity was observed in the CO + Cu-treated algae compared to the control (activity of Cu treatment alone). Under the same condition, the activity of ascorbate peroxidase was not significantly changed. These results suggest that CO can play an important role in regulating the response of algae to Cu stress.

  11. Three-dimensional display technologies.

    Science.gov (United States)

    Geng, Jason

    2013-01-01

    The physical world around us is three-dimensional (3D), yet traditional display devices can show only two-dimensional (2D) flat images that lack depth (i.e., the third dimension) information. This fundamental restriction greatly limits our ability to perceive and to understand the complexity of real-world objects. Nearly 50% of the capability of the human brain is devoted to processing visual information [Human Anatomy & Physiology (Pearson, 2012)]. Flat images and 2D displays do not harness the brain's power effectively. With rapid advances in the electronics, optics, laser, and photonics fields, true 3D display technologies are making their way into the marketplace. 3D movies, 3D TV, 3D mobile devices, and 3D games have increasingly demanded true 3D display with no eyeglasses (autostereoscopic). Therefore, it would be very beneficial to readers of this journal to have a systematic review of state-of-the-art 3D display technologies.

  12. Flat panel display - Impurity doping technology for flat panel displays

    Energy Technology Data Exchange (ETDEWEB)

    Suzuki, Toshiharu [Advanced Technology Planning, Sumitomo Eaton Nova Corporation, SBS Tower 9F, 10-1, Yoga 4-chome, Setagaya-ku, 158-0097 Tokyo (Japan)]. E-mail: suzuki_tsh@senova.co.jp

    2005-08-01

    Features of the flat panel displays (FPDs) such as liquid crystal display (LCD) and organic light emitting diode (OLED) display, etc. using low temperature poly-Si (LTPS) thin film transistors (TFTs) are briefly reviewed comparing with other FPDs. The requirements for fabricating TFTs used for high performance FPDs and system on glass (SoG) are addressed. This paper focuses on the impurity doping technology, which is one of the key technologies together with crystallization by laser annealing, formation of high quality gate insulator and gate-insulator/poly-Si interface. The issues to be solved in impurity doping technology for state of the art and future TFTs are clarified.

  13. Chemotyping of yeast mutants using robotics.

    Science.gov (United States)

    Rieger, K J; El-Alama, M; Stein, G; Bradshaw, C; Slonimski, P P; Maundrell, K

    1999-07-01

    By now, the EUROFAN programme for the functional analysis of genes from the yeast genome has attained its cruising speed. Indeed, several hundreds of yeast mutants with no phenotype as tested by growth on standard media and no significant sequence similarity to proteins of known function are available through the efforts of various laboratories. Based on the methodology initiated during the pilot project on yeast chromosome III (Yeast 13, 1547-1562, 1997) we adapted it to High Throughput Screening (HTS), using robotics. The first 100 different gene deletions from EUROSCARF, constructed in an FY1679 strain background, were run against a collection of about 300 inhibitors. Many of these inhibitors have not been reported until now to interfere in vivo with growth of Saccharomyces cerevisiae. In the present paper we provide a list of novel growth conditions and a compilation of 49 yeast deletants (from chromosomes II, IV, VII, X, XIV, XV) corresponding to 58% of the analysed genes, with at least one clear and stringent phenotype. The majority of these deletants are sensitive to one or two compounds (monotropic phenotype) while a distinct subclass of deletants displays a hyper-pleiotropic phenotype with sensitivities to a dozen or more compounds. Therefore, chemotyping of unknown genes with a large spectrum of drugs opens new vistas for a more in-depth functional analysis and a more precise definition of molecular targets.

  14. Phenotypic comparison of samdc and spe mutants reveals complex relationships of polyamine metabolism in Ustilago maydis.

    Science.gov (United States)

    Valdés-Santiago, Laura; Cervantes-Chávez, José Antonio; Winkler, Robert; León-Ramírez, Claudia G; Ruiz-Herrera, José

    2012-03-01

    Synthesis of spermidine involves the action of two enzymes, spermidine synthase (Spe) and S-adenosylmethionine decarboxylase (Samdc). Previously we cloned and disrupted the gene encoding Spe as a first approach to unravel the biological function of spermidine in Ustilago maydis. With this background, the present study was designed to provide a better understanding of the role played by Samdc in the regulation of the synthesis of this polyamine. With this aim we proceeded to isolate and delete the gene encoding Samdc from U. maydis, and made a comparative analysis of the phenotypes of samdc and spe mutants. Both spe and samdc mutants behaved as spermidine auxotrophs, and were more sensitive than the wild-type strain to different stress conditions. However, the two mutants displayed significant differences: in contrast to spe mutants, samdc mutants were more sensitive to LiCl stress, high spermidine concentrations counteracted their dimorphic deficiency, and they were completely avirulent. It is suggested that these differences are possibly related to differences in exogenous spermidine uptake or the differential location of the respective enzymes in the cell. Alternatively, since samdc mutants accumulate higher levels of S-adenosylmethionine (SAM), whereas spe mutants accumulate decarboxylated SAM, the known opposite roles of these metabolites in the processes of methylation and differentiation offer an additional attractive hypothesis to explain the phenotypic differences of the two mutants, and provide insights into the additional roles of polyamine metabolism in the physiology of the cell.

  15. Bacterial mutants for enhanced succinate production

    NARCIS (Netherlands)

    Baart, G.J.E.; Beauprez, J.J.R.; Foulquie, M.M.R.; Heijnen, J.J.; Maertens, J.

    2010-01-01

    The present invention relates to a method for obtaining enhanced metabolite production in micro-organisms, and to mutants and/or transformants obtained with said method. More particularly, it relates to bacterial mutants and/or transformants for enhanced succinate production, especially mutants and/

  16. Problem-Solving Test: Tryptophan Operon Mutants

    Science.gov (United States)

    Szeberenyi, Jozsef

    2010-01-01

    This paper presents a problem-solving test that deals with the regulation of the "trp" operon of "Escherichia coli." Two mutants of this operon are described: in mutant A, the operator region of the operon carries a point mutation so that it is unable to carry out its function; mutant B expresses a "trp" repressor protein unable to bind…

  17. Tone compatibility between HDR displays

    Science.gov (United States)

    Bist, Cambodge; Cozot, Rémi; Madec, Gérard; Ducloux, Xavier

    2016-09-01

    High Dynamic Range (HDR) is the latest trend in television technology and we expect an in ux of HDR capable consumer TVs in the market. Initial HDR consumer displays will operate on a peak brightness of about 500-1000 nits while in the coming years display peak brightness is expected to go beyond 1000 nits. However, professionally graded HDR content can range from 1000 to 4000 nits. As with Standard Dynamic Range (SDR) content, we can expect HDR content to be available in variety of lighting styles such as low key, medium key and high key video. This raises concerns over tone-compatibility between HDR displays especially when adapting to various lighting styles. It is expected that dynamic range adaptation between HDR displays uses similar techniques as found with tone mapping and tone expansion operators. In this paper, we survey simple tone mapping methods of 4000 nits color-graded HDR content for 1000 nits HDR displays. We also investigate tone expansion strategies when HDR content graded in 1000 nits is displayed on 4000 nits HDR monitors. We conclude that the best tone reproduction technique between HDR displays strongly depends on the lighting style of the content.

  18. 衣藻有性生殖的分子机制%Molecular Mechanism of Chlamydomonas Mating

    Institute of Scientific and Technical Information of China (English)

    李修岭; 李夜光

    2009-01-01

    衣藻作为分子生物学研究的模式材料,被广泛用于植物光合作用、鞭毛组装与功能、细胞周期与节律、细胞信号传导与光感受、细胞识别等重要生物学过程的研究,而且衣藻有性生殖的分子机制与人类某些疾病的发生机制存在联系.该文对国内外近年来有关莱茵衣藻在有性生殖过程中凝集素的动态分布,包括鞭毛粘连、补充、传递、脱粘连、凝集素合成的正调节,以及与性凝集素行为有关的基因研究进展进行综述,以阐明衣藻有性生殖的分子机制,为人类的疾病研究提供参考.%As a model organism for studying photosynthesis of plant,flagella assembly and function,cell cycle and circadian rhythms,signal transduction,light perception and cell recognition,Chlamydomonas has been investigated widely.This review reported the advances in molecular mechanism of Chlamydomonas reinhardtii mating.We focused on adhension,tipping,disadhension and regulation of synthesis of sex agglutinins as well as some genes related with adhension of agglutinins.The molecular mechanism of Chlamydomonas mating might provide reference to the study of some human diseases.

  19. Polyclonal antibodies against the TLA1 protein also recognize with high specificity the D2 reaction center protein of PSII in the green alga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Mitra, Mautusi; Dewez, David; García-Cerdán, Jose Gines; Melis, Anastasios

    2012-04-01

    The Chlamydomonas reinhardtii DNA-insertional transformant truncated light-harvesting antenna 1 (tla1) mutant, helped identify the novel TLA1 gene (GenBank Accession # AF534570-71) as an important genetic determinant in the chlorophyll antenna size of photosynthesis. Down-regulation in the amount of the TLA1 23 kDa protein in the cell resulted in smaller chlorophyll antenna size for both photosystems (in Tetali et al. Planta 225:813-829, 2007). Specific polyclonal antibodies, raised against the recombinant TLA1 protein, showed a cross-reaction with the predicted 23 kDa TLA1 protein in C. reinhardtii protein extracts, but also showed a strong cross-reaction with a protein band migrating to 28.5 kDa. Questions of polymorphism, or posttranslational modification of the TLA1 protein were raised as a result of the unexpected 28.5 kDa cross-reaction. Work in this paper aimed to elucidate the nature of the unexpected 28.5 kDa cross-reaction, as this was deemed to be important in terms of the functional role of the TLA1 protein in the regulation of the chlorophyll antenna size of photosynthesis. Immuno-precipitation of the 28.5 kDa protein, followed by LC-mass spectrometry, showed amino acid sequences ascribed to the psbD/D2 reaction center protein of PSII. The common antigenic determinant between TLA1 and D2 was shown to be a stretch of nine conserved amino acids V-F-L(V)LP-GNAL in the C-terminus of the two proteins, constituting a high antigenicity "GNAL" domain. Antibodies raised against the TLA1 protein containing this domain recognized both the TLA1 and the D2 protein. Conversely, antibodies raised against the TLA1 protein minus the GNAL domain specifically recognized the 23 kDa TLA1 protein and failed to recognize the 28.5 kDa D2 protein. D2 antibodies raised against an oligopeptide containing this domain also cross-reacted with the TLA1 protein. It is concluded that the 28.5 kDa cross-reaction of C. reinhardtii protein extracts with antiTLA1 antibodies is due to

  20. Quantitative screening of yeast surface-displayed polypeptide libraries by magnetic bead capture.

    Science.gov (United States)

    Yeung, Yik A; Wittrup, K Dane

    2002-01-01

    Magnetic bead capture is demonstrated here to be a feasible alternative for quantitative screening of favorable mutants from a cell-displayed polypeptide library. Flow cytometric sorting with fluorescent probes has been employed previously for high throughput screening for either novel binders or improved mutants. However, many laboratories do not have ready access to this technology as a result of the limited availability and high cost of cytometers, restricting the use of cell-displayed libraries. Using streptavidin-coated magnetic beads and biotinylated ligands, an alternative approach to cell-based library screening for improved mutants was developed. Magnetic bead capture probability of labeled cells is shown to be closely correlated with the surface ligand density. A single-pass enrichment ratio of 9400 +/- 1800-fold, at the expense of 85 +/- 6% binder losses, is achieved from screening a library that contains one antibody-displaying cell (binder) in 1.1 x 10(5) nondisplaying cells. Additionally, kinetic screening for an initial high affinity to low affinity (7.7-fold lower) mutant ratio of 1:95,000, the magnetic bead capture method attains a single-pass enrichment ratio of 600 +/- 200-fold with a 75 +/- 24% probability of loss for the higher affinity mutant. The observed high loss probabilities can be straightforwardly compensated for by library oversampling, given the inherently parallel nature of the screen. Overall, these results demonstrate that magnetic beads are capable of quantitatively screening for novel binders and improved mutants. The described methods are directly analogous to procedures in common use for phage display and should lower the barriers to entry for use of cell surface display libraries.

  1. Ultraminiature, Micropower Multipurpose Display Project

    Data.gov (United States)

    National Aeronautics and Space Administration — High information content electronic displays remain the most difficult element of the human-machine interface to effectively miniaturize. Mobile applications need a...

  2. Color speckle in laser displays

    Science.gov (United States)

    Kuroda, Kazuo

    2015-07-01

    At the beginning of this century, lighting technology has been shifted from discharge lamps, fluorescent lamps and electric bulbs to solid-state lighting. Current solid-state lighting is based on the light emitting diodes (LED) technology, but the laser lighting technology is developing rapidly, such as, laser cinema projectors, laser TVs, laser head-up displays, laser head mounted displays, and laser headlamps for motor vehicles. One of the main issues of laser displays is the reduction of speckle noise1). For the monochromatic laser light, speckle is random interference pattern on the image plane (retina for human observer). For laser displays, RGB (red-green-blue) lasers form speckle patterns independently, which results in random distribution of chromaticity, called color speckle2).

  3. ENERGY STAR Certified Displays - Deprecated

    Data.gov (United States)

    U.S. Environmental Protection Agency — This dataset is up-to-date but newer better data can be retrieved at: https://data.energystar.gov/dataset/ENERGY-STAR-Certified-Displays/xsyb-v8gs Certified models...

  4. Ten inch Planar Optic Display

    Energy Technology Data Exchange (ETDEWEB)

    Beiser, L. [Beiser (Leo) Inc., Flushing, NY (United States); Veligdan, J. [Brookhaven National Lab., Upton, NY (United States)

    1996-04-01

    A Planar Optic Display (POD) is being built and tested for suitability as a high brightness replacement for the cathode ray tube, (CRT). The POD display technology utilizes a laminated optical waveguide structure which allows a projection type of display to be constructed in a thin (I to 2 inch) housing. Inherent in the optical waveguide is a black cladding matrix which gives the display a black appearance leading to very high contrast. A Digital Micromirror Device, (DMD) from Texas Instruments is used to create video images in conjunction with a 100 milliwatt green solid state laser. An anamorphic optical system is used to inject light into the POD to form a stigmatic image. In addition to the design of the POD screen, we discuss: image formation, image projection, and optical design constraints.

  5. Identification of a Long Rice Spikelet Mutant

    Institute of Scientific and Technical Information of China (English)

    WU Xian-jun; WANG Bin; HAN Zan-ping; XIE Zhao-hui; MOU Chun-hong; WANG Xu-dong

    2004-01-01

    A spontaneously occurring rice (Oryza sativa L. ) mutant, characterized by homeotic conversion in glumes and stamens, was found in the progeny of a cross. The mutant showed long glumes and glumaceous lodicules and morphological transformation of stamens into pistils. Mutant florets consisted of 1 to 3 completely developed pistils, some pistilloid stamens with filaments, but tipped by bulged tissue and 0 to 3 stigmas. It seens that the mutant phenotype of the homeotic conversions in glumes and stamens is similar to that of the B loss-of-function mutants in Arabidopsis and Antirrhinum. The mutant is controlled by a single recessive gene as a segregation ratio of 3:1 (wild type to mutant plants) was observed in the F2 generation.

  6. The path to triacylglyceride obesity in the sta6 strain of Chlamydomonas reinhardtii.

    Science.gov (United States)

    Goodenough, Ursula; Blaby, Ian; Casero, David; Gallaher, Sean D; Goodson, Carrie; Johnson, Shannon; Lee, Jae-Hyeok; Merchant, Sabeeha S; Pellegrini, Matteo; Roth, Robyn; Rusch, Jannette; Singh, Manmilan; Umen, James G; Weiss, Taylor L; Wulan, Tuya

    2014-05-01

    When the sta6 (starch-null) strain of the green microalga Chlamydomonas reinhardtii is nitrogen starved in acetate and then "boosted" after 2 days with additional acetate, the cells become "obese" after 8 days, with triacylglyceride (TAG)-filled lipid bodies filling their cytoplasm and chloroplasts. To assess the transcriptional correlates of this response, the sta6 strain and the starch-forming cw15 strain were subjected to RNA-Seq analysis during the 2 days prior and 2 days after the boost, and the data were compared with published reports using other strains and growth conditions. During the 2 h after the boost, ∼425 genes are upregulated ≥2-fold and ∼875 genes are downregulated ≥2-fold in each strain. Expression of a small subset of "sensitive" genes, encoding enzymes involved in the glyoxylate and Calvin-Benson cycles, gluconeogenesis, and the pentose phosphate pathway, is responsive to culture conditions and genetic background as well as to boosting. Four genes-encoding a diacylglycerol acyltransferase (DGTT2), a glycerol-3-P dehydrogenase (GPD3), and two candidate lipases (Cre03.g155250 and Cre17.g735600)-are selectively upregulated in the sta6 strain. Although the bulk rate of acetate depletion from the medium is not boost enhanced, three candidate acetate permease-encoding genes in the GPR1/FUN34/YaaH superfamily are boost upregulated, and 13 of the "sensitive" genes are strongly responsive to the cell's acetate status. A cohort of 64 autophagy-related genes is downregulated by the boost. Our results indicate that the boost serves both to avert an autophagy program and to prolong the operation of key pathways that shuttle carbon from acetate into storage lipid, the combined outcome being enhanced TAG accumulation, notably in the sta6 strain.

  7. The Path to Triacylglyceride Obesity in the sta6 Strain of Chlamydomonas reinhardtii

    Science.gov (United States)

    Blaby, Ian; Casero, David; Gallaher, Sean D.; Goodson, Carrie; Johnson, Shannon; Lee, Jae-Hyeok; Merchant, Sabeeha S.; Pellegrini, Matteo; Roth, Robyn; Rusch, Jannette; Singh, Manmilan; Umen, James G.; Weiss, Taylor L.; Wulan, Tuya

    2014-01-01

    When the sta6 (starch-null) strain of the green microalga Chlamydomonas reinhardtii is nitrogen starved in acetate and then “boosted” after 2 days with additional acetate, the cells become “obese” after 8 days, with triacylglyceride (TAG)-filled lipid bodies filling their cytoplasm and chloroplasts. To assess the transcriptional correlates of this response, the sta6 strain and the starch-forming cw15 strain were subjected to RNA-Seq analysis during the 2 days prior and 2 days after the boost, and the data were compared with published reports using other strains and growth conditions. During the 2 h after the boost, ∼425 genes are upregulated ≥2-fold and ∼875 genes are downregulated ≥2-fold in each strain. Expression of a small subset of “sensitive” genes, encoding enzymes involved in the glyoxylate and Calvin-Benson cycles, gluconeogenesis, and the pentose phosphate pathway, is responsive to culture conditions and genetic background as well as to boosting. Four genes—encoding a diacylglycerol acyltransferase (DGTT2), a glycerol-3-P dehydrogenase (GPD3), and two candidate lipases (Cre03.g155250 and Cre17.g735600)—are selectively upregulated in the sta6 strain. Although the bulk rate of acetate depletion from the medium is not boost enhanced, three candidate acetate permease-encoding genes in the GPR1/FUN34/YaaH superfamily are boost upregulated, and 13 of the “sensitive” genes are strongly responsive to the cell's acetate status. A cohort of 64 autophagy-related genes is downregulated by the boost. Our results indicate that the boost serves both to avert an autophagy program and to prolong the operation of key pathways that shuttle carbon from acetate into storage lipid, the combined outcome being enhanced TAG accumulation, notably in the sta6 strain. PMID:24585881

  8. Experimental Definition and Validation of Protein Coding Transcripts in Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Kourosh Salehi-Ashtiani; Jason A. Papin

    2012-01-13

    Algal fuel sources promise unsurpassed yields in a carbon neutral manner that minimizes resource competition between agriculture and fuel crops. Many challenges must be addressed before algal biofuels can be accepted as a component of the fossil fuel replacement strategy. One significant challenge is that the cost of algal fuel production must become competitive with existing fuel alternatives. Algal biofuel production presents the opportunity to fine-tune microbial metabolic machinery for an optimal blend of biomass constituents and desired fuel molecules. Genome-scale model-driven algal metabolic design promises to facilitate both goals by directing the utilization of metabolites in the complex, interconnected metabolic networks to optimize production of the compounds of interest. Using Chlamydomonas reinhardtii as a model, we developed a systems-level methodology bridging metabolic network reconstruction with annotation and experimental verification of enzyme encoding open reading frames. We reconstructed a genome-scale metabolic network for this alga and devised a novel light-modeling approach that enables quantitative growth prediction for a given light source, resolving wavelength and photon flux. We experimentally verified transcripts accounted for in the network and physiologically validated model function through simulation and generation of new experimental growth data, providing high confidence in network contents and predictive applications. The network offers insight into algal metabolism and potential for genetic engineering and efficient light source design, a pioneering resource for studying light-driven metabolism and quantitative systems biology. Our approach to generate a predictive metabolic model integrated with cloned open reading frames, provides a cost-effective platform to generate metabolic engineering resources. While the generated resources are specific to algal systems, the approach that we have developed is not specific to algae and

  9. Hydrogen production by Chlamydomonas reinhardtii: an elaborate interplay of electron sources and sinks

    Energy Technology Data Exchange (ETDEWEB)

    Hemschemeier, A; Happe, T. [Ruhr Univ Bochum, AG Photobiotechnol, Lehrstuhl Biochem Pflanzen, Fak Biol, D-44780 Bochum (Germany); Fouchard, S [Univ Nantes, CNRS, Lab GEPEA, UMR 6144, F-44602 St Nazaire (France); Cournac, L; Peltier, G. [CEA Cadarache, DSV, IBEB, SBVME, Lab Bioenerget et Biotechnol Bacteries et Microalgues, CEA-CNRS-Univ Aix Marseille, UMR 6191, F-13108 St Paul Les Durance (France)

    2008-07-01

    The unicellular green alga Chlamydomonas reinhardtii possesses a [FeFe]-hydrogenase HydA1 (EC 1.12.7.2), which is coupled to the photosynthetic electron transport chain. Large amounts of H{sub 2} are produced in a light-dependent reaction for several days when C. reinhardtii cells are deprived of sulfur. Under these conditions, the cells drastically change their physiology from aerobic photosynthetic growth to an anaerobic resting state. The understanding of the underlying physiological processes is not only important for getting further insights into the adaptability of photosynthesis, but will help to optimize the biotechnological application of algae as H{sub 2} producers. Two of the still most disputed questions regarding H{sub 2} generation by C. reinhardtii concern the electron source for H{sub 2} evolution and the competition of the hydrogenase with alternative electron sinks. We analyzed the H{sub 2} metabolism of S-depleted C. reinhardtii cultures utilizing a special mass spectrometer setup and investigated the influence of photosystem II (PSII)- or ribulose-bisphosphate-carboxylase/oxygenase (Rubisco)-deficiency. We show that electrons for H{sub 2}-production are provided both by PSII activity and by a non-photochemical plastoquinone reduction pathway, which is dependent on previous PSII activity. In a Rubisco-deficient strain, which produces H{sub 2} also in the presence of sulfur, H{sub 2} generation seems to be the only significant electron sink for PSII activity and rescues this strain at least partially from a light-sensitive phenotype.The latter indicates that the down-regulation of assimilatory pathways in S-deprived C. reinhardtii cells is one of the important prerequisites for a sustained H{sub 2} evolution. (authors)

  10. Cellular oxido-reductive proteins of Chlamydomonas reinhardtii control the biosynthesis of silver nanoparticles

    Directory of Open Access Journals (Sweden)

    Barwal Indu

    2011-12-01

    Full Text Available Abstract Background Elucidation of molecular mechanism of silver nanoparticles (SNPs biosynthesis is important to control its size, shape and monodispersity. The evaluation of molecular mechanism of biosynthesis of SNPs is of prime importance for the commercialization and methodology development for controlling the shape and size (uniform distribution of SNPs. The unicellular algae Chlamydomonas reinhardtii was exploited as a model system to elucidate the role of cellular proteins in SNPs biosynthesis. Results The C. reinhardtii cell free extract (in vitro and in vivo cells mediated synthesis of silver nanoparticles reveals SNPs of size range 5 ± 1 to 15 ± 2 nm and 5 ± 1 to 35 ± 5 nm respectively. In vivo biosynthesized SNPs were localized in the peripheral cytoplasm and at one side of flagella root, the site of pathway of ATP transport and its synthesis related enzymes. This provides an evidence for the involvement of oxidoreductive proteins in biosynthesis and stabilization of SNPs. Alteration in size distribution and decrease of synthesis rate of SNPs in protein-depleted fractions confirmed the involvement of cellular proteins in SNPs biosynthesis. Spectroscopic and SDS-PAGE analysis indicate the association of various proteins on C. reinhardtii mediated in vivo and in vitro biosynthesized SNPs. We have identified various cellular proteins associated with biosynthesized (in vivo and in vitro SNPs by using MALDI-MS-MS, like ATP synthase, superoxide dismutase, carbonic anhydrase, ferredoxin-NADP+ reductase, histone etc. However, these proteins were not associated on the incubation of pre-synthesized silver nanoparticles in vitro. Conclusion Present study provides the indication of involvement of molecular machinery and various cellular proteins in the biosynthesis of silver nanoparticles. In this report, the study is mainly focused towards understanding the role of diverse cellular protein in the synthesis and capping of silver

  11. Inorganic polyphosphate occurs in the cell wall of Chlamydomonas reinhardtii and accumulates during cytokinesis

    Directory of Open Access Journals (Sweden)

    Freimoser Florian M

    2007-09-01

    Full Text Available Abstract Background Inorganic polyphosphate (poly P, linear chains of phosphate residues linked by energy rich phosphoanhydride bonds, is found in every cell and organelle and is abundant in algae. Depending on its localization and concentration, poly P is involved in various biological functions. It serves, for example, as a phosphate store and buffer against alkali, is involved in energy metabolism and regulates the activity of enzymes. Bacteria defective in poly P synthesis are impaired in biofilm development, motility and pathogenicity. PolyP has also been found in fungal cell walls and bacterial envelopes, but has so far not been measured directly or stained specifically in the cell wall of any plant or alga. Results Here, we demonstrate the presence of poly P in the cell wall of Chlamydomonas reinhardtii by staining with specific poly P binding proteins. The specificity of the poly P signal was verified by various competition experiments, by staining with different poly P binding proteins and by correlation with biochemical quantification. Microscopical investigation at different time-points during growth revealed fluctuations of the poly P signal synchronous with the cell cycle: The poly P staining peaked during late cytokinesis and was independent of the high intracellular poly P content, which fluctuated only slightly during the cell cycle. Conclusion The presented staining method provides a specific and sensitive tool for the study of poly P in the extracellular matrices of algae and could be used to describe the dynamic behaviour of cell wall poly P during the cell cycle. We assume that cell wall poly P and intracellular poly P are regulated by distinct mechanisms and it is suggested that cell wall bound poly P might have important protective functions against toxic compounds or pathogens during cytokinesis, when cells are more vulnerable.

  12. L,L-diaminopimelate aminotransferase from Chlamydomonas reinhardtii: a target for algaecide development.

    Directory of Open Access Journals (Sweden)

    Renwick C J Dobson

    Full Text Available In some bacterial species and photosynthetic cohorts, including algae, the enzyme L,L-diaminopimelate aminotransferase (DapL (E.C. 2.6.1.83 is involved in the anabolism of the essential amino acid L-lysine. DapL catalyzes the conversion of tetrahydrodipicolinate (THDPA to L,L-diaminopimelate (L,L-DAP, in one step bypassing the DapD, DapC and DapE enzymatic reactions present in the acyl DAP pathways. Here we present an in vivo and in vitro characterization of the DapL ortholog from the alga Chlamydomonas reinhardtii (Cr-DapL. The in vivo analysis illustrated that the enzyme is able to functionally complement the E. coli dap auxotrophs and was essential for plant development in Arabidopsis. In vitro, the enzyme was able to inter-convert THDPA and L,L-DAP, showing strong substrate specificity. Cr-DapL was dimeric in both solution and when crystallized. The structure of Cr-DapL was solved in its apo form, showing an overall architecture of a α/β protein with each monomer in the dimer adopting a pyridoxal phosphate-dependent transferase-like fold in a V-shaped conformation. The active site comprises residues from both monomers in the dimer and shows some rearrangement when compared to the apo-DapL structure from Arabidopsis. Since animals do not possess the enzymatic machinery necessary for the de novo synthesis of the amino acid L-lysine, enzymes involved in this pathway are attractive targets for the development of antibiotics, herbicides and algaecides.

  13. Preventive effect of the microalga Chlamydomonas debaryana on the acute phase of experimental colitis in rats.

    Science.gov (United States)

    Avila-Román, Javier; Talero, Elena; Alcaide, Antonio; Reyes, Carolina de Los; Zubía, Eva; García-Mauriño, Sofía; Motilva, Virginia

    2014-10-14

    Inflammatory bowel diseases (IBD) are characterised by chronic uncontrolled inflammation of intestinal mucosa. Diet and nutritional factors have emerged as possible interventions for IBD. Microalgae are rich sources of n-3 PUFA and derived oxylipins. Oxylipins are lipid mediators involved in the resolution of many inflammatory disorders. The aim of the present study was to investigate the effects of the oxylipin-containing biomass of the microalga Chlamydomonas debaryana and its major oxylipin constituent, (9Z,11E,13S,15Z)-13-hydroxyoctadeca-9,11,15-trienoic acid ((13S)-HOTE), on acute 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced colitis in rats. Lyophilised microalgal biomass and (13S)-HOTE were administered by oral route 48, 24 and 1 h before the induction of colitis and 24 h later, and the rats were killed after 48 h. The treatment with the lyophilised microalga and (13S)-HOTE improved body-weight loss and colon shortening, as well as attenuated the extent of colonic damage and increased mucus production. Cellular neutrophil infiltration, with the subsequent increase in myeloperoxidase levels induced by TNBS, were also reduced after the administration of the lyophilised microalga or (13S)-HOTE. The anti-inflammatory effects of these treatments were confirmed by the inhibition of colonic TNF-α production. Moreover, lyophilised microalga or (13S)-HOTE down-regulated cyclo-oxygenase-2 and inducible nitric oxide synthase expression. The present study was the first to show the prophylactic effects of a lyophilised biomass sample of the microalga C. debaryana and the oxylipin (13S)-HOTE on TNBS-induced acute colitis in rats. Our findings suggest that the microalga C. debaryana or derived oxylipins could be used as nutraceuticals in the treatment of the active phase of IBD.

  14. Sulphate, more than a nutrient, protects the microalga Chlamydomonas moewusii from cadmium toxicity.

    Science.gov (United States)

    Mera, Roi; Torres, Enrique; Abalde, Julio

    2014-03-01

    Sulphur is an essential macroelement that plays important roles in living organisms. The thiol rich sulphur compounds, such as cysteine, γ-Glu-Cys, glutathione and phytochelatins participate in the tolerance mechanisms against cadmium toxicity. Plants, algae, yeasts and most prokaryotes cover their demand for reduced sulphur by reduction of inorganic sulphate. The aim of this study was to investigate, using a bifactorial experimental design, the effect of different sulphate concentrations in the nutrient solution on cadmium toxicity in the freshwater microalga Chlamydomonas moewusii. Cell growth, kinetic parameters of sulphate utilization and intracellular concentrations of low-molecular mass thiol compounds were determined. A mathematical model to describe the growth of this microalga based on the effects of sulphate and cadmium was obtained. An ANOVA revealed an interaction between them, 16% of the effect sizes was explained by this interaction. A higher amount of sulphate in the culture medium allowed a higher cadmium tolerance due to an increase in the thiol compound biosynthesis. The amount of low-molecular mass thiol compounds, mainly phytochelatins, synthesized by this microalga was significantly dependent on the sulphate and cadmium concentrations; the higher phytochelatin content was obtained in cultures with 4 mg Cd/L and 1mM sulphate. The maximum EC50 value (based on nominal cadmium concentration) reached for this microalga was 4.46 ± 0.42 mg Cd/L when the sulphate concentration added to the culture medium was also 1mM. An increase in the sulphate concentration, in deficient environments, could alleviate the toxic effect of this metal; however, a relative excess is also negative. The results obtained showed a substrate inhibition for this nutrient. An uncompetitive model for sulphate was chosen to establish the mathematical model that links both factors.

  15. High yield secretion of recombinant proteins from the microalga Chlamydomonas reinhardtii.

    Science.gov (United States)

    Ramos-Martinez, E M; Fimognari, L; Sakuragi, Y

    2017-02-16

    Microalga-based biomanufacturing of recombinant proteins is attracting growing attention due to its advantages in safety, metabolic diversity, scalability, and sustainability. Secretion of recombinant proteins can accelerate the use of microalgal platforms by allowing post-translational modifications and easy recovery of products from the culture media. However, currently, the yields of secreted recombinant proteins are low, which hampers the commercial application of this strategy. This study aimed at expanding the genetic tools for enhancing secretion of recombinant proteins in Chlamydomonas reinhardtii, a widely used green microalga as a model organism and a potential industrial biotechnology platform. We demonstrated that the putative signal sequence from C. reinhardtii gametolysin can assist the secretion of the yellow fluorescent protein Venus into the culture media. In order to increase the secretion yields, Venus was C-terminally fused with synthetic glycomodules comprised of tandem serine (Ser) and proline (Pro) repeats of 10 and 20 units [hereafter (SP)n, wherein n=10 or 20]. The yields of the (SP)n-fused Venus were higher than Venus without the glycomodule by up to 12 folds, with the maximum yield of 15 mg L(-1) . Moreover, the presence of the glycomodules confererred an enhanced proteolytic protein stability. The Venus-(SP)n proteins were shown to be glycosylated, and a treatment of the cells with Brefeldin A led to a suggestion that glycosylation of the (SP)n glycomodules starts in the endoplasmic reticulum (ER). Taken together, the results demonstrate the utility of the gametolysin signal sequence and (SP)n glycomodule to promote a more efficient biomanufacturing of microalgae-based recombinant proteins. This article is protected by copyright. All rights reserved.

  16. Characterization of lead induced metal-phytochelatin complexes in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Scheidegger, Christian; Sigg, Laura; Behra, Renata

    2011-11-01

    Accumulation of Pb and induction of phytochelatin synthesis were observed in Chlamydomonas reinhardtii upon Pb(II) exposure. Our aim was to examine whether Pb(II) is bound by phytochelatins (PCs) in C. reinhardtii and to examine formed complexes for their stoichiometry and composition. Metal-phytochelatin (Me-PC) complexes induced by Pb were isolated by size-exclusion chromatography in 13 collected fractions, which were analyzed for their PC and metal content by high-performance liquid chromatography and inductively coupled plasma mass spectrometry. A recovery of more than 90% of Pb from standard Pb-PC₂ complexes within the total volume of the size-exclusion column indicated the adequacy of the method for Pb-PC(n) complex separation and characterization. Phytochelatins were detected mainly in a molecular weight ranging from 1,000 to 5,300 daltons (Da), indicating the formation of complexes with various stoichiometries. Approximately 72% of total PC₂ eluted in the range from 1,000 to 1,600 Da, and 80% of total PC₃ eluted in the molecular weight range from 1,600 to 2,300 Da. The distribution of Cu, Zn, and Pb showed that more than 70% of these metals were associated with the high-molecular-weight fractions. Copper, zinc, and lead were also observed in PC-containing fractions, suggesting the formation of various Me-PC complexes. The results of the present study indicate that the role of PCs in Pb detoxification is minor, because only 13% of total Pb was associated with PCs.

  17. Saturating Light Induces Sustained Accumulation of Oil in Plastidal Lipid Droplets in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Goold, Hugh Douglas; Cuiné, Stéphan; Légeret, Bertrand; Liang, Yuanxue; Brugière, Sabine; Auroy, Pascaline; Javot, Hélène; Tardif, Marianne; Jones, Brian; Beisson, Fred; Peltier, Gilles; Li-Beisson, Yonghua

    2016-08-01

    Enriching algal biomass in energy density is an important goal in algal biotechnology. Nitrogen (N) starvation is considered the most potent trigger of oil accumulation in microalgae and has been thoroughly investigated. However, N starvation causes the slow down and eventually the arrest of biomass growth. In this study, we show that exposing a Chlamydomonas reinhardtii culture to saturating light (SL) under a nonlimiting CO2 concentration in turbidostatic photobioreactors induces a sustained accumulation of lipid droplets (LDs) without compromising growth, which results in much higher oil productivity than N starvation. We also show that the polar membrane lipid fraction of SL-induced LDs is rich in plastidial lipids (approximately 70%), in contrast to N starvation-induced LDs, which contain approximately 60% lipids of endoplasmic reticulum origin. Proteomic analysis of LDs isolated from SL-exposed cells identified more than 200 proteins, including known proteins of lipid metabolism, as well as 74 proteins uniquely present in SL-induced LDs. LDs induced by SL and N depletion thus differ in protein and lipid contents. Taken together, lipidomic and proteomic data thus show that a large part of the sustained oil accumulation occurring under SL is likely due to the formation of plastidial LDs. We discuss our data in relation to the different metabolic routes used by microalgae to accumulate oil reserves depending on cultivation conditions. Finally, we propose a model in which oil accumulation is governed by an imbalance between photosynthesis and growth, which can be achieved by impairing growth or by boosting photosynthetic carbon fixation, with the latter resulting in higher oil productivity.

  18. Tying Down Loose Ends in the Chlamydomonas Genome: Functional Significance of Abundant Upstream Open Reading Frames

    Directory of Open Access Journals (Sweden)

    Frederick R. Cross

    2016-02-01

    Full Text Available The Chlamydomonas genome has been sequenced, assembled, and annotated to produce a rich resource for genetics and molecular biology in this well-studied model organism. The annotated genome is very rich in open reading frames upstream of the annotated coding sequence (‘uORFs’: almost three quarters of the assigned transcripts have at least one uORF, and frequently more than one. This is problematic with respect to the standard ‘scanning’ model for eukaryotic translation initiation. These uORFs can be grouped into three classes: class 1, initiating in-frame with the coding sequence (CDS (thus providing a potential in-frame N-terminal extension; class 2, initiating in the 5′ untranslated sequences (5UT and terminating out-of-frame in the CDS; and class 3, initiating and terminating within the 5UT. Multiple bioinformatics criteria (including analysis of Kozak consensus sequence agreement and BLASTP comparisons to the closely related Volvox genome, and statistical comparison to cds and to random sequence controls indicate that of ∼4000 class 1 uORFs, approximately half are likely in vivo translation initiation sites. The proposed resulting N-terminal extensions in many cases will sharply alter the predicted biochemical properties of the encoded proteins. These results suggest significant modifications in ∼2000 of the ∼20,000 transcript models with respect to translation initiation and encoded peptides. In contrast, class 2 uORFs may be subject to purifying selection, and the existent ones (surviving selection are likely inefficiently translated. Class 3 uORFs are found in more than half of transcripts, frequently multiple times per transcript; however, they are remarkably similar to random sequence expectations with respect to size, number, and composition, and therefore may in most cases be selectively neutral.

  19. Use of microalgae Chlamydomonas reinhardtii for production of double-stranded RNA against shrimp virus

    Directory of Open Access Journals (Sweden)

    Parinyachat Somchai

    2016-05-01

    Full Text Available RNA interference has been proposed to be a promising tool for combating shrimp viruses. Antiviral double-stranded (dsRNA has been mostly produced in Escherichia coli-expression system because of its high efficiency and inexpensive operations. However, overusing the bacteria may raise concerns regarding public health and environmental contamination, and seeking for a new dsRNA production platform would be alternative for future molecular farming. In this study, we exploited the green microalgae Chlamydomonas reinhardtii to produce dsRNA targeting the lethal shrimp yellow head virus (YHV. The expression plasmid pSL18 for C. reinhardtii was constructed to contain YHV-specific hairpin RNA expression cassette, and the successful assembly of pSL18-YHV was confirmed by PCR and enzymatic digestions. Glass bead method was employed for transformation of C. reinhardtii nuclear genome with pSL18-YHV. Microalgal expression of dsRNA-YHV, approximately 45 ng from 100-mL culture, was detected by qRT-PCR. Oral feeding experiment on postlarval shrimp revealed that the formulated feed with C. reinhardtii expressing dsRNA-YHV, at the ratio of 1 × 108 transformants per gram feed, improved 22% survival rate after YHV challenge. The present study suggests that C. reinhardtii can be bioengineered to produce viral-specific dsRNA for shrimp viral disease control, and the developed qRT-PCR could detect microalgal dsRNA with detection limit of subpicogram.

  20. Structure, circadian regulation and bioinformatic analysis of the unique sigma factor gene in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Carter, Matthew L; Smith, Annette C; Kobayashi, Hirokazu; Purton, Saul; Herrin, David L

    2004-01-01

    In higher plants, the transcription of plastid genes is mediated by at least two types of RNA polymerase (RNAP); a plastid-encoded bacterial RNAP in which promoter specificity is conferred by nuclear-encoded sigma factors, and a nuclear-encoded phage-like RNAP. Green algae, however, appear to possess only the bacterial enzyme. Since transcription of much, if not most, of the chloroplast genome in Chlamydomonas reinhardtii is regulated by the circadian clock and the nucleus, we sought to identify sigma factor genes that might be responsible for this regulation. We describe a nuclear gene (RPOD) that is predicted to encode an 80 kDa protein that, in addition to a predicted chloroplast transit peptide at the N-terminus, has the conserved motifs (2.1- 4.2) diagnostic of bacterial sigma-70 factors. We also identified two motifs not previously recognized for sigma factors, adjacent PEST sequences and a leucine zipper, both suggested to be involved in protein-protein interactions. PEST sequences were also found in approximately 40% of sigma factors examined, indicating they may be of general significance. Southern blot hybridization and BLAST searches of the genome and EST databases suggest that RPODmay be the only sigma factor gene in C. reinhardtii. The levels of RPODmRNA increased 2- 3-fold in the mid-to-late dark period of light-dark cycling cells, just prior to, or coincident with, the peak in chloroplast transcription. Also, the dark-period peak in RPOD mRNA persisted in cells shifted to continuous light or continuous dark for at least one cycle, indicating that RPODis under circadian clock control. These results suggest that regulation of RPODexpression contributes to the circadian clock's control of chloroplast transcription.

  1. Estrogens Suppress a Behavioral Phenotype in Zebrafish Mutants of the Autism Risk Gene, CNTNAP2.

    Science.gov (United States)

    Hoffman, Ellen J; Turner, Katherine J; Fernandez, Joseph M; Cifuentes, Daniel; Ghosh, Marcus; Ijaz, Sundas; Jain, Roshan A; Kubo, Fumi; Bill, Brent R; Baier, Herwig; Granato, Michael; Barresi, Michael J F; Wilson, Stephen W; Rihel, Jason; State, Matthew W; Giraldez, Antonio J

    2016-02-17

    Autism spectrum disorders (ASDs) are a group of devastating neurodevelopmental syndromes that affect up to 1 in 68 children. Despite advances in the identification of ASD risk genes, the mechanisms underlying ASDs remain unknown. Homozygous loss-of-function mutations in Contactin Associated Protein-like 2 (CNTNAP2) are strongly linked to ASDs. Here we investigate the function of Cntnap2 and undertake pharmacological screens to identify phenotypic suppressors. We find that zebrafish cntnap2 mutants display GABAergic deficits, particularly in the forebrain, and sensitivity to drug-induced seizures. High-throughput behavioral profiling identifies nighttime hyperactivity in cntnap2 mutants, while pharmacological testing reveals dysregulation of GABAergic and glutamatergic systems. Finally, we find that estrogen receptor agonists elicit a behavioral fingerprint anti-correlative to that of cntnap2 mutants and show that the phytoestrogen biochanin A specifically reverses the mutant behavioral phenotype. These results identify estrogenic compounds as phenotypic suppressors and illuminate novel pharmacological pathways with relevance to autism.

  2. Bioinformatic Analysis of the Nitrate Reductase Gene in Antartic Ice Algae Chlamydomonas sp. ICE-L%南极衣藻Chlamydomonas sp.ICE-L硝酸还原酶基因的生物信息学分析

    Institute of Scientific and Technical Information of China (English)

    林敏卓; 刘晨临; 黄晓航; 杨平平

    2012-01-01

    Nitrate reductase (NR) plays an important role in the abiotic stress adaptation in plants by regulating nitrogen metabolism. A nitrate reductase (NR) gene of Antarctic ice algae, Chlamydomonas sp. ICE-L, was identified from the cDNA library and sequenced. The encoded protein sequence of NR gene was investigated by bioinformatic analysis. Through sequence alignment the active sites of ICE-L NR protein sequence which may related to stress acclimation was identified. In addition, the tertiary structure of ICE- L NR protein sequence was predicted. The full-length of Chlamydomonas ICE-L NR gene contained an open reading frame of 2,589 bp encoding a nitrate reductase of 863 amino acids. Phylogenetic analysis showed that the gene was homologous to known green algae NRs with identity of 63%, 61%, 60% and 54% to Volvox carteri, Chlamydomonas reinhardtii, Dunaliella tertiolecta and Chlorella vulgaris respectively. The functional prediction analysis revealed that NR gene sequence has 3 different functional domains which was similar to higher plant. This bioinformatic analysis about NR gene of ICE- L will help us further understand and deeply expand the recearch on the acclimatizing mechanism of Antarctic ice alga Chlamydomonas in the extreme environment from the angle of NR gene.%硝酸还原酶(NR)除调节植物的氮代谢外,在植物的各种非生物胁迫的适应过程中也发挥着重要的作用.从南极冰藻Chlamydomonas sp.ICE-L的cDNA文库中筛选到了硝酸还原酶的全长基因,对其进行测序并对其编码的蛋白序列进行了生物信息学分析,构建了NR的系统进化树,通过多序列比对探讨了可能与该酶逆境适应性相关的活性位点,并对该蛋白进行了三级结构预测分析.结果显示,NR基因的编码区长2 589 bp,编码863个氨基酸.在以氨基酸序列构建的系统进化树中,南极衣藻的NR序列和其他绿藻类的聚在一起,与团藻、莱茵衣藻、杜氏盐藻和小球藻

  3. Mapping of barley alpha-amylases and outer subsite mutants reveals dynamic high-affinity subsites and barriers in the long substrate binding cleft

    DEFF Research Database (Denmark)

    Kandra, L.; Abou Hachem, Maher; Gyemant, G.;

    2006-01-01

    as binding barriers. Barley a-amylase I mutants Y105A and T212Y at subsite -6 and +4 resulted in release or anchoring of bound substrate, thus modifying the affinities of other high-affinity subsites (-2 and +2) and barriers. The double mutant Y105A-T212Y displayed a hybrid subsite affinity profile...

  4. Phosphors for flat panel emissive displays

    Energy Technology Data Exchange (ETDEWEB)

    Anderson, M.T.; Walko, R.J.; Phillips, M.L.F.

    1995-07-01

    An overview of emissive display technologies is presented. Display types briefly described include: cathode ray tubes (CRTs), field emission displays (FEDs), electroluminescent displays (ELDs), and plasma display panels (PDPs). The critical role of phosphors in further development of the latter three flat panel emissive display technologies is outlined. The need for stable, efficient red, green, and blue phosphors for RGB fall color displays is emphasized.

  5. Comparative genomics in Chlamydomonas and Plasmodium identifies an ancient nuclear envelope protein family essential for sexual reproduction in protists, fungi, plants, and vertebrates.

    Science.gov (United States)

    Ning, Jue; Otto, Thomas D; Pfander, Claudia; Schwach, Frank; Brochet, Mathieu; Bushell, Ellen; Goulding, David; Sanders, Mandy; Lefebvre, Paul A; Pei, Jimin; Grishin, Nick V; Vanderlaan, Gary; Billker, Oliver; Snell, William J

    2013-05-15

    Fertilization is a crucial yet poorly characterized event in eukaryotes. Our previous discovery that the broadly conserved protein HAP2 (GCS1) functioned in gamete membrane fusion in the unicellular green alga Chlamydomonas and the malaria pathogen Plasmodium led us to exploit the rare biological phenomenon of isogamy in Chlamydomonas in a comparative transcriptomics strategy to uncover additional conserved sexual reproduction genes. All previously identified Chlamydomonas fertilization-essential genes fell into related clusters based on their expression patterns. Out of several conserved genes in a minus gamete cluster, we focused on Cre06.g280600, an ortholog of the fertilization-related Arabidopsis GEX1. Gene disruption, cell biological, and immunolocalization studies show that CrGEX1 functions in nuclear fusion in Chlamydomonas. Moreover, CrGEX1 and its Plasmodium ortholog, PBANKA_113980, are essential for production of viable meiotic progeny in both organisms and thus for mosquito transmission of malaria. Remarkably, we discovered that the genes are members of a large, previously unrecognized family whose first-characterized member, KAR5, is essential for nuclear fusion during yeast sexual reproduction. Our comparative transcriptomics approach provides a new resource for studying sexual development and demonstrates that exploiting the data can lead to the discovery of novel biology that is conserved across distant taxa.

  6. Acetate versus sulfur deprivation role in creating anaerobiosis in light for hydrogen production by Chlamydomonas reinhardtii and Spirulina platensis: two different organisms and two different mechanisms.

    Science.gov (United States)

    Morsy, Fatthy Mohamed

    2011-01-01

    This work was devoted to separate acetate role in creating anaerobiosis from that of sulfur deprivation. Chlamydomonas reinhardtii grown in TAP (Tris-acetate-phosphate) medium was resuspended in sulfur-replete or -deprived medium in sealed or nonsealed cultures. Sulfur deprivation was substantial for starch accumulation and hydrogen evolution; however, acetate induced anaerobiosis in the presence or absence of sulfur in only sealed cultures. In nonsealed cultures, Chlamydomonas did not lose its photosynthetic activity; however, it was arrested in anoxia with no photosynthetic activity as long as the culture was sealed. The sealed cultures resumed photosynthesis upon unsealing overnight unless the cells died by anoxia at late stage of the experiment. These results indicate that the enhanced oxygen consumption for the enormous acetate respiration and inhibition of the external oxygen supply in sealed cultures of Chlamydomonas are the main reasons for the steady anaerobic conditions. Although acetate was substantial for creating anaerobiosis in Chlamydomonas, sulfur deprivation alone could create anaerobiosis in Spirulina platensis grown autotrophically. Hydrogen evolution and glycogen accumulation were induced under such conditions. Severely reduced phycocyanin, chlorophyll and photosynthesis, while respiration had increased, induced anaerobiosis in Spirulina. This study reports for the first time anaerobiosis under autotrophic conditions in a cyanobacterium.

  7. Overexpressing Ferredoxins in Chlamydomonas reinhardtii Increase Starch and Oil Yields and Enhance Electric Power Production in a Photo Microbial Fuel Cell

    Directory of Open Access Journals (Sweden)

    Li-Fen Huang

    2015-08-01

    Full Text Available Ferredoxins (FDX are final electron carrier proteins in the plant photosynthetic pathway, and function as major electron donors in diverse redox-driven metabolic pathways. We previously showed that overexpression of a major constitutively expressed ferredoxin gene PETF in Chlamydomonas decreased the reactive oxygen species (ROS level and enhanced tolerance to heat stress. In addition to PETF, an endogenous anaerobic induced FDX5 was overexpressed in transgenic Chlamydomonas lines here to address the possible functions of FDX5. All the independent FDX transgenic lines showed decreased cellular ROS levels and enhanced tolerance to heat and salt stresses. The transgenic Chlamydomonas lines accumulated more starch than the wild-type line and this effect increased almost three-fold in conditions of nitrogen depletion. Furthermore, the lipid content was higher in the transgenic lines than in the wild-type line, both with and without nitrogen depletion. Two FDX-overexpressing Chlamydomonas lines were assessed in a photo microbial fuel cell (PMFC; power density production by the transgenic lines was higher than that of the wild-type cells. These findings suggest that overexpression of either PETF or FDX5 can confer tolerance against heat and salt stresses, increase starch and oil production, and raise electric power density in a PMFC.

  8. Taxonomic revision of Chlamydomonas subg. Amphichloris (Volvocales, Chlorophyceae), with resurrection of the genus Dangeardinia and descriptions of Ixipapillifera gen. nov. and Rhysamphichloris gen. nov.

    Science.gov (United States)

    Nakada, Takashi; Tomita, Masaru; Wu, Jiunn-Tzong; Nozaki, Hisayoshi

    2016-04-01

    Chlamydomonas (Cd.) is one of the largest but most polyphyletic genera of freshwater unicellular green algae. It consists of 400-600 morphological species and requires taxonomic revision. Toward reclassification, each morphologically defined classical subgenus (or subgroup) should be examined using culture strains. Chlamydomonas subg. Amphichloris is characterized by a central nucleus between two axial pyrenoids, however, the phylogenetic structure of this subgenus has yet to be examined using molecular data. Here, we examined 12 strains including six newly isolated strains, morphologically identified as Chlamydomonas subg. Amphichloris, using 18S rRNA gene phylogeny, light microscopy, and mitochondria fluorescent microscopy. Molecular phylogenetic analyses revealed three independent lineages of the subgenus, separated from the type species of Chlamydomonas, Cd. reinhardtii. These three lineages were further distinguished from each other by light and fluorescent microscopy-in particular by the morphology of the papillae, chloroplast surface, stigmata, and mitochondria-and are here assigned to three genera: Dangeardinia emend., Ixipapillifera gen. nov., and Rhysamphichloris gen. nov. Based on the molecular and morphological data, two to three species were recognized in each genus, including one new species, I. pauromitos. In addition, Cd. deasonii, which was previously assigned to subgroup "Pleiochloris," was included in the genus Ixipapillifera as I. deasonii comb. nov.

  9. Productivity and selective accumulation of carotenoids of the novel extremophile microalga Chlamydomonas acidophila grown with different carbon sources in batch systems

    NARCIS (Netherlands)

    Cuaresma, M.; Casal, C.; Forján, E.; Vílchez, C.

    2011-01-01

    Cultivation of extremophile microorganisms has attracted interest due to their ability to accumulate highvalue compounds. Chlamydomonas acidophila is an acidophile green microalga isolated by our group from Tinto River, an acidic river that flows down from the mining area in Huelva, Spain. This micr

  10. Mutant p53 protein localized in the cytoplasm inhibits autophagy.

    Science.gov (United States)

    Morselli, Eugenia; Tasdemir, Ezgi; Maiuri, Maria Chiara; Galluzzi, Lorenzo; Kepp, Oliver; Criollo, Alfredo; Vicencio, José Miguel; Soussi, Thierry; Kroemer, Guido

    2008-10-01

    The knockout, knockdown or chemical inhibition of p53 stimulates autophagy. Moreover, autophagy-inducing stimuli such as nutrient depletion, rapamycin or lithium cause the depletion of cytoplasmic p53, which in turn is required for the induction of autophagy. Here, we show that retransfection of p53(-/-) HCT 116 colon carcinoma cells with wild type p53 decreases autophagy down to baseline levels. Surprisingly, one third among a panel of 22 cancer-associated p53 single amino acid mutants also inhibited autophagy when transfected into p53(-/-) cells. Those variants of p53 that preferentially localize to the cytoplasm effectively repressed autophagy, whereas p53 mutants that display a prominently nuclear distribution failed to inhibit autophagy. The investigation of a series of deletion mutants revealed that removal of the DNA-binding domain from p53 fails to interfere with its role in the regulation of autophagy. Altogether, these results identify the cytoplasmic localization of p53 as the most important feature for p53-mediated autophagy inhibition. Moreover, the structural requirements for the two biological activities of extranuclear p53, namely induction of apoptosis and inhibition of autophagy, are manifestly different.

  11. Atrial fibrillation-linked germline GJA5/connexin40 mutants showed an increased hemichannel function.

    Directory of Open Access Journals (Sweden)

    Yiguo Sun

    Full Text Available Mutations in GJA5 encoding the gap junction protein connexin40 (Cx40 have been linked to lone atrial fibrillation. Some of these mutants result in impaired gap junction function due to either abnormal connexin localization or impaired gap junction channels, which may play a role in promoting atrial fibrillation. However, the effects of the atrial fibrillation-linked Cx40 mutants on hemichannel function have not been studied. Here we investigated two atrial fibrillation-linked germline Cx40 mutants, V85I and L221I. These two mutants formed putative gap junction plaques at cell-cell interfaces, with similar gap junction coupling conductance as that of wild-type Cx40. Connexin deficient HeLa cells expressing either one of these two mutants displayed prominent propidium iodide-uptake distinct from cells expressing wild-type Cx40 or other atrial fibrillation-linked Cx40 mutants, I75F, L229M, and Q49X. Propidium iodide-uptake was sensitive to [Ca2+]o and the hemichannel blockers, carbenoxolone, flufenamic acid and mefloquine, but was not affected by the pannexin 1 channel blocking agent, probenecid, indicating that uptake is most likely mediated via connexin hemichannels. A gain-of-hemichannel function in these two atrial fibrillation-linked Cx40 mutants may provide a novel mechanism underlying the etiology of atrial fibrillation.

  12. Mutant p53 and mTOR/PKM2 regulation in cancer cells.

    Science.gov (United States)

    Dando, Ilaria; Cordani, Marco; Donadelli, Massimo

    2016-09-01

    Mutations of TP53 gene are the most common feature in aggressive malignant cells. In addition to the loss of the tumor suppressive role of wild-type p53, hotspot mutant p53 isoforms display oncogenic proprieties notoriously referred as gain of functions (GOFs) which result in chemoresistance to therapies, genomic instability, aberrant deregulation of cell cycle progression, invasiveness and enhanced metastatic potential, and finally, in patient poor survival rate. The identification of novel functional oncogenic pathways regulated by mutant p53 represent and intriguing topic for emerging therapies against a broad spectrum of cancer types bearing mutant TP53 gene. Mammalian target of rapamycin (mTOR), as well as pyruvate kinase isoform M2 (PKM2) are master regulators of cancer growth, metabolism, and cell proliferation. Herein, we report that GOF mutant R175H and R273H p53 proteins trigger PKM2 phosphorylation on Tyr 105 through the involvement of mTOR signaling. Our data, together with the newly discovered connection between mutant p53 and mTOR stimulation, raise important implications for the potential therapeutic use of synthetic drugs inhibiting mTOR/PKM2 axis in cancer cells bearing mutant TP53 gene. We further hypothesize that mTOR/PKM2 pathway stimulation serves to sustain the oncogenic activity of mutant p53 through both the enhancement of chemoresistance and of aerobic glycolysis of cancer cells. © 2016 IUBMB Life, 68(9):722-726, 2016.

  13. A Genetic Screen for Mutants with Supersized Lipid Droplets in Caenorhabditis elegans

    Science.gov (United States)

    Li, Shiwei; Xu, Shibin; Ma, Yanli; Wu, Shuang; Feng, Yu; Cui, Qingpo; Chen, Lifeng; Zhou, Shuang; Kong, Yuanyuan; Zhang, Xiaoyu; Yu, Jialei; Wu, Mengdi; Zhang, Shaobing O.

    2016-01-01

    To identify genes that regulate the dynamics of lipid droplet (LD) size, we have used the genetically tractable model organism Caenorhabditis elegans, whose wild-type LD population displays a steady state of size with an upper limit of 3 μm in diameter. From a saturated forward genetic screen of 6.7 × 105 mutagenized haploid genomes, we isolated 118 mutants with supersized intestinal LDs often reaching 10 μm. These mutants define nine novel complementation groups, in addition to four known genes (maoc-1, dhs-28, daf-22, and prx-10). The nine groups are named drop (lipid droplet abnormal) and categorized into four classes. Class I mutants drop-5 and drop-9, similar to prx-10, are up-regulated in ACS-22-DGAT-2-dependent LD growth, resistant to LD hydrolysis, and defective in peroxisome import. Class II mutants drop-2, drop-3, drop-6, and drop-7 are up-regulated in LD growth, are resistant to LD hydrolysis, but are not defective in peroxisome import. Class III mutants drop-1 and drop-8 are neither up-regulated in LD growth nor resistant to LD hydrolysis, but seemingly up-regulated in LD fusion. Class IV mutant drop-4 is cloned as sams-1 and, different to the other three classes, is ACS-22-independent and hydrolysis-resistant. These four classes of supersized LD mutants should be valuable for mechanistic studies of LD cellular processes including growth, hydrolysis, and fusion. PMID:27261001

  14. A Genetic Screen for Mutants with Supersized Lipid Droplets in Caenorhabditis elegans

    Directory of Open Access Journals (Sweden)

    Shiwei Li

    2016-08-01

    Full Text Available To identify genes that regulate the dynamics of lipid droplet (LD size, we have used the genetically tractable model organism Caenorhabditis elegans, whose wild-type LD population displays a steady state of size with an upper limit of 3 μm in diameter. From a saturated forward genetic screen of 6.7 × 105 mutagenized haploid genomes, we isolated 118 mutants with supersized intestinal LDs often reaching 10 μm. These mutants define nine novel complementation groups, in addition to four known genes (maoc-1, dhs-28, daf-22, and prx-10. The nine groups are named drop (lipid droplet abnormal and categorized into four classes. Class I mutants drop-5 and drop-9, similar to prx-10, are up-regulated in ACS-22-DGAT-2-dependent LD growth, resistant to LD hydrolysis, and defective in peroxisome import. Class II mutants drop-2, drop-3, drop-6, and drop-7 are up-regulated in LD growth, are resistant to LD hydrolysis, but are not defective in peroxisome import. Class III mutants drop-1 and drop-8 are neither up-regulated in LD growth nor resistant to LD hydrolysis, but seemingly up-regulated in LD fusion. Class IV mutant drop-4 is cloned as sams-1 and, different to the other three classes, is ACS-22-independent and hydrolysis-resistant. These four classes of supersized LD mutants should be valuable for mechanistic studies of LD cellular processes including growth, hydrolysis, and fusion.

  15. Systems Biology Approach in Chlamydomonas Reveals Connections between Copper Nutrition and Multiple Metabolic Steps[C][W][OA

    Science.gov (United States)

    Castruita, Madeli; Casero, David; Karpowicz, Steven J.; Kropat, Janette; Vieler, Astrid; Hsieh, Scott I.; Yan, Weihong; Cokus, Shawn; Loo, Joseph A.; Benning, Christoph; Pellegrini, Matteo; Merchant, Sabeeha S.

    2011-01-01

    In this work, we query the Chlamydomonas reinhardtii copper regulon at a whole-genome level. Our RNA-Seq data simulation and analysis pipeline validated a 2-fold cutoff and 10 RPKM (reads per kilobase of mappable length per million mapped reads) (~1 mRNA per cell) to reveal 63 CRR1 targets plus another 86 copper-responsive genes. Proteomic and immunoblot analyses captured 25% of the corresponding proteins, whose abundance was also dependent on copper nutrition, validating transcriptional regulation as a major control mechanism for copper signaling in Chlamydomonas. The impact of copper deficiency on the expression of several O2-dependent enzymes included steps in lipid modification pathways. Quantitative lipid profiles indicated increased polyunsaturation of fatty acids on thylakoid membrane digalactosyldiglycerides, indicating a global impact of copper deficiency on the photosynthetic apparatus. Discovery of a putative plastid copper chaperone and a membrane protease in the thylakoid suggest a mechanism for blocking copper utilization in the chloroplast. We also found an example of copper sparing in the N assimilation pathway: the replacement of copper amine oxidase by a flavin-dependent backup enzyme. Forty percent of the targets are previously uncharacterized proteins, indicating considerable potential for new discovery in the biology of copper. PMID:21498682

  16. Construction of modular tandem expression vectors for the green alga Chlamydomonas reinhardtii using the Cre/lox-system.

    Science.gov (United States)

    Heitzer, Markus; Zschoernig, Barbara

    2007-09-01

    The successful expression of foreign genes mainly depends on both a reliable method for transformation and a suitable promoter sequence. We created a series of modular plasmids that facilitate the rapid construction of large tandem vectors for transgene expression under the control of different promoter sequences in Chlamydomonas reinhardtii. Tandem vectors carrying expression cassettes for Renilla luciferase and a metabolic selection marker (ARG7) were manufactured by fusing two plasmids in vitro using Cre/lox site-specific recombination. Supercoiled and linear plasmids were used to transform an arginine auxotrophic Chlamydomonas strain, and rates of co-expression as well as levels of luciferase activity were monitored for frequently used promoters (HSP70A, LHCB1, PSAD, and the chimeric HSP70A/RBCS2). Linearized tandem vectors generally increased the co-expression frequency (up to 77%) compared with standard cotransformation protocols. Most transformants showed a single and complete integration event confirming the close linkage of active selectable marker and reporter gene within the nuclear genome. The analysis of luciferase activity showed expression levels within three orders of magnitude for the promoters used, with the artificial HSP70A/RRBCS2 being the most active. For 69% of all luminescent transformants carrying the HSP70A promoter luciferase expression was enhanced by heatshock, indicating physiological promoter function in a transgenic context.

  17. Effect of Cd on GSH and GSH-related enzymes of Chlamydomonas sp. ICE-L existing in Antarctic ice

    Institute of Scientific and Technical Information of China (English)

    DING Yu; MIAO Jin-lai; LI Guang-you; WANG Quan-fu; KAN Guang-feng; WANG Guo-dong

    2005-01-01

    Glutathione(GSH) and GSH-related enzymes play a great role in protecting organisms from oxidative damage. The GSH level and GSH-related enzymes activities were investigated as well as the growth yield and malonyldialdehyde(MDA) content in the Antarctic ice microalga Chlamydomonas sp. ICE-L exposure to the different cadmium concentration in this paper. The results showed that the higher concentration Cd inhibited the growth of ICE-L significantly and Cd would induce formation of MDA. At the same time, it is clear that GSH level, glutathione peroxidases(GPx) activity and glutathione S-transferases(GST), activity were higher in ICE-L exposed to Cd than the control. But GR activity dropped notably when ICE-L were cultured in the medium containing Cd. Increase of GSH level, GPx and GST activities acclimate to oxidative stress induced by Cd and protect Antarctic ice microalga Chlamydomonas sp. ICE-L from toxicity caused by Cd exposure. These parameters may be used to assess the biological impact of Cd in the Antarctic pole region environment.

  18. Ecophysiology, secondary pigments and ultrastructure of Chlainomonas sp. (Chlorophyta) from the European Alps compared with Chlamydomonas nivalis forming red snow.

    Science.gov (United States)

    Remias, Daniel; Pichrtová, Martina; Pangratz, Marion; Lütz, Cornelius; Holzinger, Andreas

    2016-04-01

    Red snow is a well-known phenomenon caused by microalgae thriving in alpine and polar regions during the melting season. The ecology and biodiversity of these organisms, which are adapted to low temperatures, high irradiance and freeze-thaw events, are still poorly understood. We compared two different snow habitats containing two different green algal genera in the European Alps, namely algae blooming in seasonal rock-based snowfields (Chlamydomonas nivalis) and algae dominating waterlogged snow bedded over ice (Chlainomonassp.). Despite the morphological similarity of the red spores found at the snow surface, we found differences in intracellular organization investigated by light and transmission electron microscopy and in secondary pigments investigated by chromatographic analysis in combination with mass spectrometry. Spores ofChlainomonassp. show clear differences fromChlamydomonas nivalisin cell wall arrangement and plastid organization. Active photosynthesis at ambient temperatures indicates a high physiological activity, despite no cell division being present. Lipid bodies containing the carotenoid astaxanthin, which produces the red color, dominate cells of both species, but are modified differently. While inChlainomonassp. astaxanthin is mainly esterified with two fatty acids and is more apolar, inChamydomonas nivalis, in contrast, less apolar monoesters prevail.

  19. Changes of proteins in the Antarctic ice microalga Chlamydomonas sp. cultured under UV-B radiation stress

    Institute of Scientific and Technical Information of China (English)

    KAN Guangfeng; MIAO Jinlai; SHI Cuijuan; LI Guangyou

    2006-01-01

    Antarctic ice microalga Chlamydomonas sp. can thrive undisturbed under high UV radiation in the Antarctic ice layer. However, it is unknown that the initial adaptation mechanisms in protein level occurring in response to high UV radiation. Global-expression profiling of proteins in response to stress was analyzed by two-dimensional electrophoresis (2-DE) and image analysis. In the 2-DE analysis,protein preparation is the key step. Three different protein extract methods were compared, and the results showed that the trichloroacetic acid (TCA)-acetone fractional precipitation method was the fittest one. At the same time, the proteins in Chlamydomonas sp. were compared in 2-DE way, and the synthesis of seven protein spots was found disappeared and 18 decreased after exposed to UV-B radiation. In addition, 14 protein spots were enhanced or induced, among which two new peptides (20 and 21 kDa) appeared whose isoelectric point (pI) was 7.05 and 4.60 respectively. These changed proteins might act as key role in the acclimation of Antarctic ice microalga to UV-B radiation

  20. Using single cell cultivation system for on-chip monitoring of the interdivision timer in Chlamydomonas reinhardtii cell cycle

    Directory of Open Access Journals (Sweden)

    Soloviev Mikhail

    2010-09-01

    Full Text Available Abstract Regulation of cell cycle progression in changing environments is vital for cell survival and maintenance, and different regulation mechanisms based on cell size and cell cycle time have been proposed. To determine the mechanism of cell cycle regulation in the unicellular green algae Chlamydomonas reinhardtii, we developed an on-chip single-cell cultivation system that allows for the strict control of the extracellular environment. We divided the Chlamydomonas cell cycle into interdivision and division phases on the basis of changes in cell size and found that, regardless of the amount of photosynthetically active radiation (PAR and the extent of illumination, the length of the interdivision phase was inversely proportional to the rate of increase of cell volume. Their product remains constant indicating the existence of an 'interdivision timer'. The length of the division phase, in contrast, remained nearly constant. Cells cultivated under light-dark-light conditions did not divide unless they had grown to twice their initial volume during the first light period. This indicates the existence of a 'commitment sizer'. The ratio of the cell volume at the beginning of the division phase to the initial cell volume determined the number of daughter cells, indicating the existence of a 'mitotic sizer'.

  1. Optokinetic behavior is reversed in achiasmatic mutant zebrafish larvae.

    Science.gov (United States)

    Rick, J M; Horschke, I; Neuhauss, S C

    2000-05-18

    The vertebrate optokinetic nystagmus (OKN) is a compensatory oculomotor behavior that is evoked by movement of the visual environment. It functions to stabilize visual images on the retina. The OKN can be experimentally evoked by rotating a drum fitted with stripes around the animal and has been studied extensively in many vertebrate species, including teleosts. This simple behavior has earlier been used to screen for mutations affecting visual system development in the vertebrate model organism zebrafish. In such a screen, we have found a significant number of homozygous belladonna (bel) mutant larvae to be defective in the correct execution of the OKN [1]. We now show that about 40% of homozygous bel larvae display a curious reversal of the OKN upon visual stimulation. Monocular stimulation leads to primary activation of ipsilateral eye movements in larvae that behave like the wild type. In contrast, affected larvae display contralateral activation of eye movements upon monocular stimulation. Anatomical analysis of retinal ganglion cell axon projections reveal a morphological basis for the observed behavioral defect. All animals with OKN reversal are achiasmatic. Further behavioral examination of affected larvae show that OKN-reversed animals execute this behavior in a stimulus-velocity-independent manner. Our data support a parsimonious model of optokinetic reversal by the opening of a controlling feedback loop at the level of the optic chiasm that is solely responsible for the observed behavioral abnormality in mutant belladonna larvae.

  2. BES Monitoring & Displaying System

    Institute of Scientific and Technical Information of China (English)

    MengWANG; BingyunZHANG; 等

    2001-01-01

    BES1 Monitoring & Displaying System(BESMDS)is projected to monitor and display the running status of DAQ and Slow Control systems of BES through the Web for worldwide accessing.It provides a real-time remote means of monitoring as well as an approach to study the environmental influence upon physical data taking.The system collects real-time data separately from BES online subsystems by network sockets and stores the data into a database.People can access the system through its web site.which retrieves data on request from the database and can display results in dynamically created images.Its web address in http:// besmds,ihep.ac.cn/

  3. Engineering antibodies by yeast display.

    Science.gov (United States)

    Boder, Eric T; Raeeszadeh-Sarmazdeh, Maryam; Price, J Vincent

    2012-10-15

    Since its first application to antibody engineering 15 years ago, yeast display technology has been developed into a highly potent tool for both affinity maturing lead molecules and isolating novel antibodies and antibody-like species. Robust approaches to the creation of diversity, construction of yeast libraries, and library screening or selection have been elaborated, improving the quality of engineered molecules and certainty of success in an antibody engineering campaign and positioning yeast display as one of the premier antibody engineering technologies currently in use. Here, we summarize the history of antibody engineering by yeast surface display, approaches used in its application, and a number of examples highlighting the utility of this method for antibody engineering.

  4. PROGRAMMABLE DISPLAY PUSHBUTTON LEGEND EDITOR

    Science.gov (United States)

    Busquets, A. M.

    1994-01-01

    The Programmable Display Pushbutton (PDP) is a pushbutton device available from Micro Switch which has a programmable 16 x 35 matrix of LEDs on the pushbutton surface. Any desired legends can be displayed on the PDPs, producing user-friendly applications which greatly reduce the need for dedicated manual controls. Because the PDP can interact with the operator, it can call for the correct response before transmitting its next message. It is both a simple manual control and a sophisticated programmable link between the operator and the host system. The Programmable Display Pushbutton Legend Editor, PDPE, is used to create the LED displays for the pushbuttons. PDPE encodes PDP control commands and legend data into message byte strings sent to a Logic Refresh and Control Unit (LRCU). The LRCU serves as the driver for a set of four PDPs. The legend editor (PDPE) transmits to the LRCU user specified commands that control what is displayed on the LED face of the individual pushbuttons. Upon receiving a command, the LRCU transmits an acknowledgement that the message was received and executed successfully. The user then observes the effect of the command on the PDP displays and decides whether or not to send the byte code of the message to a data file so that it may be called by an applications program. The PDPE program is written in FORTRAN for interactive execution. It was developed on a DEC VAX 11/780 under VMS. It has a central memory requirement of approximately 12800 bytes. It requires four Micro Switch PDPs and two RS-232 VAX 11/780 terminal ports. The PDPE program was developed in 1985.

  5. Computational multi-projection display.

    Science.gov (United States)

    Moon, Seokil; Park, Soon-Gi; Lee, Chang-Kun; Cho, Jaebum; Lee, Seungjae; Lee, Byoungho

    2016-04-18

    A computational multi-projection display is proposed by employing a multi-projection system combining with compressive light field displays. By modulating the intensity of light rays from a spatial light modulator inside a single projector, the proposed system can offer several compact views to observer. Since light rays are spread to all directions, the system can provide flexible positioning of viewpoints without stacking projectors in vertical direction. Also, if the system is constructed properly, it is possible to generate view images with inter-pupillary gap and satisfy the super multi-view condition. We explain the principle of the proposed system and verify its feasibility with simulations and experimental results.

  6. Display standards for commercial flight decks

    Science.gov (United States)

    Lamberth, Larry S.; Penn, Cecil W.

    1994-06-01

    SAE display standards are used as guidelines for certifying commercial airborne electronic displays. The SAE document generation structure and approval process is described. The SAE committees that generate display standards are described. Three SAE documents covering flat panel displays (AS-8034, ARP-4256, and ARP-4260) are discussed with their current status. Head-Up Display documents are also in work.

  7. Biochemical and histological characterization of tomato mutants

    Directory of Open Access Journals (Sweden)

    Carolina C. Monteiro

    2012-06-01

    Full Text Available Biochemical responses inherent to antioxidant systems as well morphological and anatomical properties of photomorphogenic, hormonal and developmental tomato mutants were investigated. Compared to the non-mutant Micro-Tom (MT, we observed that the malondialdehyde (MDA content was enhanced in the diageotropica (dgt and lutescent (l mutants, whilst the highest levels of hydrogen peroxide (H2O2 were observed in high pigment 1 (hp1 and aurea (au mutants. The analyses of antioxidant enzymes revealed that all mutants exhibited reduced catalase (CAT activity when compared to MT. Guaiacol peroxidase (GPOX was enhanced in both sitiens (sit and notabilis (not mutants, whereas in not mutant there was an increase in ascorbate peroxidase (APX. Based on PAGE analysis, the activities of glutathione reductase (GR isoforms III, IV, V and VI were increased in l leaves, while the activity of superoxide dismutase (SOD isoform III was reduced in leaves of sit, epi, Never ripe (Nr and green flesh (gf mutants. Microscopic analyses revealed that hp1 and au showed an increase in leaf intercellular spaces, whereas sit exhibited a decrease. The au and hp1 mutants also exhibited a decreased in the number of leaf trichomes. The characterization of these mutants is essential for their future use in plant development and ecophysiology studies, such as abiotic and biotic stresses on the oxidative metabolism.Neste trabalho, analisamos as respostas bioquímicas inerentes ao sistema antioxidante, assim como propriedades morfológicas e anatômicas de mutantes fotomorfogenéticos e hormonais de tomateiro. Comparados ao não mutante Micro-Tom (MT, observamos que o conteúdo de malondialdeído (MDA aumentou nos mutantes diageotropica (dgt e lutescent (l, enquanto os maiores níveis de H2O2 foram encontrados nos mutantes high pigment 1 (hp1 e aurea (au. Análises de enzimas antioxidantes mostraram que todos os mutantes reduziram a atividade de catalase (CAT quando comparado a MT. A

  8. Photobiological hydrogen production with the unicellular green alga Chlamydomonas reinhardtii under process engineering aspects; Photobiologische Wasserstoffproduktion mit der einzelligen Gruenalge Chlamydomonas reinhardtii unter verfahrenstechnischen Aspekten

    Energy Technology Data Exchange (ETDEWEB)

    Geier, Stephanie

    2011-07-01

    Hydrogen is of high interest as a clean and environmentally friendly energy source as its combustion only emits water and energy. However, currently hydrogen is produced in energy demanding processes by the consumption of fossil fuels. An alternative way of sustainable and non-polluting hydrogen production could be provided by use of photosynthetic active microalgae. Within this work, the photobiological hydrogen production with the unicellular green algae Chlamydomonas reinhardtii is investigated under the aspects of bioprocess-engineering and economics. Objectives are, besides the increase of the photochemical efficiency, the cultivation of the algae and subsequent hydrogen production under cost-free sunlight. It could be demonstrated that outdoor cultivation of C. reinhardtii is possible in Central Europe throughout the year by using e.g. waste heat. Similar cell numbers in the range from 1,2.10{sup 7} cells ml{sup -1} to 1,7.10{sup 7} cells ml{sup -1} could be achieved in closed photobioreactors of the type Photobioreactor Screening Module under controlled laboratory conditions and both continuous illumination (200 {mu}mol.m{sup -2}.s{sup -1}) and simulated outdoor conditions according to the light intensity of idealized summer day as well as in outdoor experiments (up to 2000 {mu}mol.m{sup -2}.s{sup -1}).The use of 10 % CO{sub 2} corresponding to the CO{sub 2} content in flue gas led to a doubling of cell numbers under continuous illumination to 4,2.10{sup 7} cells ml{sup -1}, compared to the reference culture bubbled with 3 % CO{sub 2}. A significant increase of cell numbers under the light profiles of an idealized summer day could not be achieved. The cultivation under the light profile of a winter day at 25 C reduced cell growth to 54 %, compared to the summer simulation. In open 30 L outdoor ponds, only 0,26.10{sup 7} cells ml{sup -1} could be achieved under photoheterotrophic conditions during the summer months, which corresponds to 20 % of the cell

  9. Photobiological hydrogen production with the unicellular green alga Chlamydomonas reinhardtii under process engineering aspects; Photobiologische Wasserstoffproduktion mit der einzelligen Gruenalge Chlamydomonas reinhardtii unter verfahrenstechnischen Aspekten

    Energy Technology Data Exchange (ETDEWEB)

    Geier, Stephanie

    2011-07-01

    Hydrogen is of high interest as a clean and environmentally friendly energy source as its combustion only emits water and energy. However, currently hydrogen is produced in energy demanding processes by the consumption of fossil fuels. An alternative way of sustainable and non-polluting hydrogen production could be provided by use of photosynthetic active microalgae. Within this work, the photobiological hydrogen production with the unicellular green algae Chlamydomonas reinhardtii is investigated under the aspects of bioprocess-engineering and economics. Objectives are, besides the increase of the photochemical efficiency, the cultivation of the algae and subsequent hydrogen production under cost-free sunlight. It could be demonstrated that outdoor cultivation of C. reinhardtii is possible in Central Europe throughout the year by using e.g. waste heat. Similar cell numbers in the range from 1,2.10{sup 7} cells ml{sup -1} to 1,7.10{sup 7} cells ml{sup -1} could be achieved in closed photobioreactors of the type Photobioreactor Screening Module under controlled laboratory conditions and both continuous illumination (200 {mu}mol.m{sup -2}.s{sup -1}) and simulated outdoor conditions according to the light intensity of idealized summer day as well as in outdoor experiments (up to 2000 {mu}mol.m{sup -2}.s{sup -1}).The use of 10 % CO{sub 2} corresponding to the CO{sub 2} content in flue gas led to a doubling of cell numbers under continuous illumination to 4,2.10{sup 7} cells ml{sup -1}, compared to the reference culture bubbled with 3 % CO{sub 2}. A significant increase of cell numbers under the light profiles of an idealized summer day could not be achieved. The cultivation under the light profile of a winter day at 25 C reduced cell growth to 54 %, compared to the summer simulation. In open 30 L outdoor ponds, only 0,26.10{sup 7} cells ml{sup -1} could be achieved under photoheterotrophic conditions during the summer months, which corresponds to 20 % of the cell

  10. Autostereoscopic display with eye tracking

    Science.gov (United States)

    Tomono, Takao; Hoon, Kyung; Ha, Yong Soo; Kim, Sung-Sik; Son, Jung-Young

    2002-05-01

    Auto-stereoscopic 21-inch display with eye tracking having wide viewing zone and bright image was fabricated. The image of display is projected to retinal through several optical components. We calculated optical system for wider viewing zone by using Inverse-Ray Trace Method. The viewing zone of first model is 155mm (theoretical value: 161mm). We could widen viewing zone by controlling paraxial radius of curvature of spherical mirror, the distance between lenses and so on. The viewing zone of second model is 208mm. We used two spherical mirrors to obtain twice brightness. We applied eye-tracking system to the display system. Eye recognition is based on neural network card based on ZICS technology. We fabricated Auto-stereoscopic 21-inch display with eye tracking. We measured viewing zone based on illumination area. The viewing zone was 206mm, which was close to theoretical value. We could get twice brightness also. We could see 3D image according to position without headgear.

  11. Graphics Display of Foreign Scripts.

    Science.gov (United States)

    Abercrombie, John R.

    1987-01-01

    Describes Graphics Project for Foreign Language Learning at the University of Pennsylvania, which has developed ways of displaying foreign scripts on microcomputers. Character design on computer screens is explained; software for graphics, printing, and language instruction is discussed; and a text editor is described that corrects optically…

  12. Real Time Sonic Boom Display

    Science.gov (United States)

    Haering, Ed

    2014-01-01

    This presentation will provide general information about sonic boom mitigation technology to the public in order to supply information to potential partners and licensees. The technology is a combination of flight data, atmospheric data and terrain information implemented into a control room real time display for flight planning. This research is currently being performed and as such, any results and conclusions are ongoing.

  13. Verbal Modification via Visual Display

    Science.gov (United States)

    Richmond, Edmun B.; Wallace-Childers, La Donna

    1977-01-01

    The inability of foreign language students to produce acceptable approximations of new vowel sounds initiated a study to devise a real-time visual display system whereby the students could match vowel production to a visual pedagogical model. The system used amateur radio equipment and a standard oscilloscope. (CHK)

  14. Information retrieval and display system

    Science.gov (United States)

    Groover, J. L.; King, W. L.

    1977-01-01

    Versatile command-driven data management system offers users, through simplified command language, a means of storing and searching data files, sorting data files into specified orders, performing simple or complex computations, effecting file updates, and printing or displaying output data. Commands are simple to use and flexible enough to meet most data management requirements.

  15. Colour displays for categorical images

    NARCIS (Netherlands)

    Glasbey, C.; Heijden, van der G.W.A.M.; Toh, V.F.K.; Gray, A.J.

    2007-01-01

    We propose a method for identifying a set of colours for displaying 2D and 3D categorical images when the categories are unordered labels. The principle is to find maximally distinct sets of colours. We either generate colours sequentially, to maximize the dissimilarity or distance between a new col

  16. Book Display as Adult Service.

    Science.gov (United States)

    Moore, Matthew S.

    1997-01-01

    Defines book display as an adult service as choosing and positioning adult books from the library collection to increase their circulation. The author contrasts bookstore arrangement for sales versus library arrangement for access, including contrasting missions, genre grouping, weeding, problems, and dimensions. (Author/LRW)

  17. Enhanced Ascorbate Regeneration Via Dehydroascorbate Reductase Confers Tolerance to Photo-Oxidative Stress in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Lin, Shu-Tseng; Chiou, Chih-Wen; Chu, Yen-Lin; Hsiao, Yu; Tseng, Yu-Fei; Chen, Yi-Chun; Chen, Hsien-Jung; Chang, Hsin-Yang; Lee, Tse-Min

    2016-10-01

    The role of ascorbate (AsA) recycling via dehydroascorbate reductase (DHAR) in the tolerance of Chlamydomonas reinhardtii to photo-oxidative stress was examined. The activity of DHAR and the abundance of the CrDHAR1 (Cre10.g456750) transcript increased after moderate light (ML; 750 µmol m(-2) s(-1)) or high light (HL; 1,800 µmol m(-2) s(-1)) illumination, accompanied by dehydroascorbate (DHA) accumulation, decreased AsA redox state, photo-inhibition, lipid peroxidation, H2O2 overaccumulation, growth inhibition and cell death. It suggests that DHAR and AsA recycling is limiting under high-intensity light stress. The CrDHAR1 gene was cloned and its recombinant CrDHAR1 protein was a monomer (25 kDa) detected by Western blot that exhibits an enzymatic activity of 965 µmol min(-1)( )mg(-1) protein. CrDHAR1 was overexpressed driven by a HSP70A:RBCS2 fusion promoter or down-regulated by artificial microRNA (amiRNA) to examine whether DHAR-mediated AsA recycling is critical for the tolerance of C. reinahartii cells to photo-oxidative stress. The overexpression of CrDHAR1 increased DHAR protein abundance and enzyme activity, AsA pool size, AsA:DHA ratio and the tolerance to ML-, HL-, methyl viologen- or H2O2-induced oxidative stress. The CrDHAR1-knockdown amiRNA lines that have lower DHAR expression and AsA recycling ability were sensitive to high-intensity illumination and oxidative stress. The glutathione pool size, glutathione:oxidized glutathione ratio and glutathione reductase and ascorbate peroxidase activities were increased in CrDHAR1-overexpressing cells and showed a further increase after high-intensity illumination but decreased in wild-type cells after light stress. The present results suggest that increasing AsA regeneration via enhanced DHAR activity modulates the ascorbate-glutathione cycle activity in C. reinhardtii against photo-oxidative stress.

  18. Interactive effects of copper oxide nanoparticles and light to green alga Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Cheloni, Giulia; Marti, Elodie; Slaveykova, Vera I., E-mail: vera.slaveykova@unige.ch

    2016-01-15

    Highlights: • Comparable stability of CuO-NP suspensions under different light conditions. • UVR* inhibits growth, bleaches chlorophyll fluorescence and damages membrane. • Below 1 mg L{sup −1} CuO-NPs do not attenuate light in algal suspension. • SNL enhances significantly the effect of 0.8 mg L{sup −1} CuO-NPs on microalgae. • Synergistic interactions between UVR* and CuO-NPs. - Abstract: The present study explores the effect of light with different spectral composition on the stability of CuO-nanoparticle (CuO-NP) dispersions and their effects to green alga Chlamydomonas reinhardtii. The results showed that simulated natural light (SNL) and light with enhanced UVB radiation (UVR*) do not affect the dissolution of CuO-NPs as compared to light irradiation conditions typically used in laboratory incubator (INC). Comparable values of ζ-potential and hydrodynamic size during 24 h were found under all studied conditions. Concentrations of CuO-NPs below 1 mg L{sup −1} do not attenuate the light penetration in the algal suspensions in comparison with NP-free system. Exposure to a combination of 8 μg L{sup −1} or 0.8 mg L{sup −1} CuO-NPs and INC or SNL has no significant effect on the algal growth inhibition, algal fluorescence and membrane integrity under short-term exposure. However, an enhancement of the percentage of cells experiencing oxidative stress was observed upon exposure to 0.8 mg L{sup −1} CuO-NPs and SNL for 4 and 8 h. Combination of UVR* and 0.8 mg L{sup −1} CuO-NPs resulted in synergistic effects for all biological endpoints. Despite the photocatalytic properties of CuO-NPs no significant increase in abiotic reactive oxygen species (ROS) production under simulated solar radiation was observed suggesting that the synergistic effect observed might be correlated to other factors than CuO-NP-mediated ROS photoproduction. Tests performed with CuSO{sub 4} confirmed the important role of dissolution as toxicity driving force for lower

  19. Harvesting microalgae cultures with superabsorbent polymers: desulfurization of Chlamydomonas reinhardtii for hydrogen production.

    Science.gov (United States)

    Martín del Campo, Julia S; Patiño, Rodrigo

    2013-12-01

    It is presented in this work a new methodology to harvest fresh water microalgae cultures by extracting the culture medium with superabsorbent polymers (SAPs). The microalgae Chlamydomonas reinhardtii were grown in the Sueoka culture medium, harvested with polyacrylic SAPs and re-suspended in the culture medium tris-acetate-potassium without sulfur (TAP-S) to generate hydrogen (H2 ) under anoxic conditions. The H2 production as an alternative fuel is relevant since this gas has high-energy recovery without involving carbon. Before microalgae harvesting, a number of range diameters (1-7 mm) for SAPs spherical particles were tested, and the initial rate (V0 ) and the maximal capacity (Qmax ) were determined for the Sueoka medium absorption. The SAP particles with the diameter range 2.0-2.5 mm performed the best and these were employed for the rest of the experiments. The Sueoka medium has a high salt content and the effect of the ionic strength was also studied for different medium concentrations (0-400%). The SAPs were reused in consecutive absorption/desorption cycles, maintaining their absorption capacity. Although the Sueoka medium reduces the SAPs absorption capacity to 40% compared with deionized water, the use of SAPs was very significant for the desulfurization process of C. reihardtii. The presence of C. reinhardtii at different concentrations does not affect the absorption capacity of the Sueoka culture medium by the SAPs. In order to reduce the time of the process, an increase of the SAPs concentration was tested, being 20 g of SAP per liter of medium, a condition to harvest the microalgae culture in 4 h. There were no evident cell ruptures during the harvesting process and the cells remained alive. Finally, the harvested biomass was re-suspended in TAP-S medium and kept under anaerobic conditions and illumination to produce H2 that was monitored by a PEM fuel cell. The use of SAPs for microalgae harvesting is a feasible non-invasive procedure to obtain

  20. CMPD: cancer mutant proteome database.

    Science.gov (United States)

    Huang, Po-Jung; Lee, Chi-Ching; Tan, Bertrand Chin-Ming; Yeh, Yuan-Ming; Julie Chu, Lichieh; Chen, Ting-Wen; Chang, Kai-Ping; Lee, Cheng-Yang; Gan, Ruei-Chi; Liu, Hsuan; Tang, Petrus

    2015-01-01

    Whole-exome sequencing, which centres on the protein coding regions of disease/cancer associated genes, represents the most cost-effective method to-date for deciphering the association between genetic alterations and diseases. Large-scale whole exome/genome sequencing projects have been launched by various institutions, such as NCI, Broad Institute and TCGA, to provide a comprehensive catalogue of coding variants in diverse tissue samples and cell lines. Further functional and clinical interrogation of these sequence variations must rely on extensive cross-platforms integration of sequencing information and a proteome database that explicitly and comprehensively archives the corresponding mutated peptide sequences. While such data resource is a critical for the mass spectrometry-based proteomic analysis of exomic variants, no database is currently available for the collection of mutant protein sequences that correspond to recent large-scale genomic data. To address this issue and serve as bridge to integrate genomic and proteomics datasets, CMPD (http://cgbc.cgu.edu.tw/cmpd) collected over 2 millions genetic alterations, which not only facilitates the confirmation and examination of potential cancer biomarkers but also provides an invaluable resource for translational medicine research and opportunities to identify mutated proteins encoded by mutated genes.

  1. Study on culturing Trichodema mutants

    Institute of Scientific and Technical Information of China (English)

    CHEN Jian-ai; WANG Wei-ming

    2004-01-01

    @@ Trichodema mutants strains T5, T0803, T1010, T1003were cultured in different conditions and media, also in the presence of fungicides at 40 mg/kg (CK or procymidone + chlorothalonil, or maneb or phosethyl-Al) . The pH values of media were 5, 6, 7 and 8 and hyphae were grown at temperatures of 15, 20, 25 and 30 ℃. After being cultured for 3, 4, 5, or 6 days, the strains were transferred at a lower temperature to sporulate (20℃) Obtained data were analyzed statistically, with the orthogonal array and ranges (R) differing dependes on the treatments (R = 40.0,42.4, 48.0, 62.8,107.0). The results indicated that the most important factor was the nature of the strain (R =107.0), while the change in temperature and time of cultivation produced the lowest effect (R =40.0). Each factor variance was significant and A3B4C2D1E3 was the optimum combined condition, in which strain T1010 grew more quickly and sporulated most.

  2. Effect of Temperature and light intensity on growth and Photosynthetic Activity of Chlamydomonas reinhard II; Efecto de la temperatura e intensidad luminosa sobre el crecimiento y actividad fotosintetica del alga Chlamydomonas Reinhardt II

    Energy Technology Data Exchange (ETDEWEB)

    Alfonsel Jaen, M.; Fernandez Gonzalez, J.

    1985-07-01

    The effect of five temperatures (15,20,25,30 and 35 degree centigree) and two levels of illumination on growth and photosynthetic activity of Chlamydomonas reinhard II has been studied. The growth of the cultures was evaluated by optical density. Photosynthetic activity has been carried out studying either the assimilation rate of C0{sub 2} labelled with C-14 or the oxygen evolution by means of polarographic measurements. The maximum photosynthetic rate has been obtained at 25 degree centigree for the lower level of illumination (2400 lux) and at 35 degree centigree for the higher one (13200 lux) and at 35 degree centigree for the higher ono (13200 lux). These results suggest an interaction of temperature and illumination on photosynthetic activity. (Author) 37 refs.

  3. Game engines and immersive displays

    Science.gov (United States)

    Chang, Benjamin; Destefano, Marc

    2014-02-01

    While virtual reality and digital games share many core technologies, the programming environments, toolkits, and workflows for developing games and VR environments are often distinct. VR toolkits designed for applications in visualization and simulation often have a different feature set or design philosophy than game engines, while popular game engines often lack support for VR hardware. Extending a game engine to support systems such as the CAVE gives developers a unified development environment and the ability to easily port projects, but involves challenges beyond just adding stereo 3D visuals. In this paper we outline the issues involved in adapting a game engine for use with an immersive display system including stereoscopy, tracking, and clustering, and present example implementation details using Unity3D. We discuss application development and workflow approaches including camera management, rendering synchronization, GUI design, and issues specific to Unity3D, and present examples of projects created for a multi-wall, clustered, stereoscopic display.

  4. Characterization of the rotating display.

    Science.gov (United States)

    Keyes, J W; Fahey, F H; Harkness, B A; Eggli, D F; Balseiro, J; Ziessman, H A

    1988-09-01

    The rotating display is a useful method for reviewing single photon emission computed tomography (SPECT) data. This study evaluated the requirements for a subjectively pleasing and useful implementation of this technique. Twelve SPECT data sets were modified and viewed by several observers who recorded the minimum framing rates for apparent smooth rotation, 3D effect, effects of image size, and other parameters. The results showed that a minimum of 16 frames was needed for a useful display. Smaller image sizes and more frames were preferred. The recommended minimal framing rate for a 64-frame study is 16-17 frames per second and for a 32-frame study, 12-13 frames per second. Other enhancements also were useful.

  5. Displays for future intermediate UAV

    Science.gov (United States)

    Desjardins, Daniel; Metzler, James; Blakesley, David; Rister, Courtney; Nuhu, Abdul-Razak

    2008-04-01

    The Dedicated Autonomous Extended Duration Airborne Long-range Utility System (DAEDALUS) is a prototype Unmanned Aerial Vehicle (UAV) that won the 2007 AFRL Commander's Challenge. The purpose of the Commander's Challenge was to find an innovative solution to urgent warfighter needs by designing a UAV with increased persistence for tactical employment of sensors and communication systems. DAEDALUS was chosen as a winning prototype by AFRL, AFMC and SECAF. Follow-on units are intended to fill an intermediate role between currently fielded Tier I and Tier II UAV's. The UAV design discussed in this paper, including sensors and displays, will enter Phase II for Rapid Prototype Development with the intent of developing the design for eventual production. This paper will discuss the DAEDALUS UAV prototype system, with particular focus on its communications, to include the infrared sensor and electro-optical camera, but also displays, specifically man-portable.

  6. Interactive display of polygonal data

    Energy Technology Data Exchange (ETDEWEB)

    Wood, P.M.

    1977-10-01

    Interactive computer graphics is an excellent approach to many types of applications. It is an exciting method of doing geographic analysis when desiring to rapidly examine existing geographically related data or to display specially prepared data and base maps for publication. One such program is the interactive thematic mapping system called CARTE, which combines polygonal base maps with statistical data to produce shaded maps using a variety of shading symbolisms on a variety of output devices. A polygonal base map is one where geographic entities are described by points, lines, or polygons. It is combined with geocoded data to produce special subject or thematic maps. Shading symbolisms include texture shading for areas, varying widths for lines, and scaled symbols for points. Output devices include refresh and storage CRTs and auxiliary Calcomp or COM hardcopy. The system is designed to aid in the quick display of spatial data and in detailed map design.

  7. Striations in Plasma Display Panel

    Institute of Scientific and Technical Information of China (English)

    OUYANG Ji-Ting; CAO Jing; MIAO Jin-Song

    2005-01-01

    @@ The phenomenon of striation has been investigated experimentally in a macroscopic ac-plasma display panel (PDP). The relationship between the characteristics of striation and the operation conditions including voltage, frequency, rib, and electrode configuration, etc is obtained experimentally. The origin of the striations is considered to be the ionization waves in the transient positive column near the dielectric surface in the anode area during the discharge, and the perturbation is caused by resonance kinetic effects in inert gas.

  8. Modern Display Technologies and Applications

    Science.gov (United States)

    1982-01-01

    conventional tubes, LSI circuitry offers the possibility of correcting some of the deficiencies in electron-optic perform- ance and may lead to acceptable...certain ceramic materials such as PLZT (lead lanthanum zirconate titanate) can be utilized for display applications. PLZT is transparent in the visible...consuming power (3.8.12). 3.8.4.2 State of development, Magnetic particles have been made of polyethylene with powdered Strontium ferrite as a filler

  9. GridOrbit public display

    DEFF Research Database (Denmark)

    Ramos, Juan David Hincapie; Tabard, Aurélien; Bardram, Jakob

    2010-01-01

    We introduce GridOrbit, a public awareness display that visualizes the activity of a community grid used in a biology laboratory. This community grid executes bioin-formatics algorithms and relies on users to donate CPU cycles to the grid. The goal of GridOrbit is to create a shared awareness about...... people comment on projects. Our work explores the usage of interactive technologies as enablers for the appropriation of an otherwise invisible infrastructure....

  10. Proof nets for display logic

    CERN Document Server

    Moot, Richard

    2007-01-01

    This paper explores several extensions of proof nets for the Lambek calculus in order to handle the different connectives of display logic in a natural way. The new proof net calculus handles some recent additions to the Lambek vocabulary such as Galois connections and Grishin interactions. It concludes with an exploration of the generative capacity of the Lambek-Grishin calculus, presenting an embedding of lexicalized tree adjoining grammars into the Lambek-Grishin calculus.

  11. Multiview synthesis for autostereoscopic displays

    Science.gov (United States)

    Dane, Gökçe.; Bhaskaran, Vasudev

    2013-09-01

    Autostereoscopic (AS) displays spatially multiplex multiple views, providing a more immersive experience by enabling users to view the content from different angles without the need of 3D glasses. Multiple views could be captured from multiple cameras at different orientations, however this could be expensive, time consuming and not applicable to some applications. The goal of multiview synthesis in this paper is to generate multiple views from a stereo image pair and disparity map by using various video processing techniques including depth/disparity map processing, initial view interpolation, inpainting and post-processing. We specifically emphasize the need for disparity processing when there is no depth information is available that is associated with the 2D data and we propose a segmentation based disparity processing algorithm to improve disparity map. Furthermore we extend the texture based 2D inpainting algorithm to 3D and further improve the hole-filling performance of view synthesis. The benefit of each step of the proposed algorithm is demonstrated with comparison to state of the art algorithms in terms of visual quality and PSNR metric. Our system is evaluated in an end-to-end multi view synthesis framework where only stereo image pair is provided as input to the system and 8 views are outputted and displayed in 8-view Alioscopy AS display.

  12. Nuclear protein import is reduced in cells expressing nuclear envelopathy-causing lamin A mutants

    Energy Technology Data Exchange (ETDEWEB)

    Busch, Albert; Kiel, Tilman; Heupel, Wolfgang-M. [University of Wuerzburg, Institute of Anatomy and Cell Biology, Koellikerstrasse 6, 97070 Wuerzburg (Germany); Wehnert, Manfred [Institute of Human Genetics, University of Greifswald, Greifswald (Germany); Huebner, Stefan, E-mail: stefan.huebner@mail.uni-wuerzburg.de [University of Wuerzburg, Institute of Anatomy and Cell Biology, Koellikerstrasse 6, 97070 Wuerzburg (Germany)

    2009-08-15

    Lamins, which form the nuclear lamina, not only constitute an important determinant of nuclear architecture, but additionally play essential roles in many nuclear functions. Mutations in A-type lamins cause a wide range of human genetic disorders (laminopathies). The importance of lamin A (LaA) in the spatial arrangement of nuclear pore complexes (NPCs) prompted us to study the role of LaA mutants in nuclear protein transport. Two mutants, causing prenatal skin disease restrictive dermopathy (RD) and the premature aging disease Hutchinson Gilford progeria syndrome, were used for expression in HeLa cells to investigate their impact on the subcellular localization of NPC-associated proteins and nuclear protein import. Furthermore, dynamics of the LaA mutants within the nuclear lamina were studied. We observed affected localization of NPC-associated proteins, diminished lamina dynamics for both LaA mutants and reduced nuclear import of representative cargo molecules. Intriguingly, both LaA mutants displayed similar effects on nuclear morphology and functions, despite their differences in disease severity. Reduced nuclear protein import was also seen in RD fibroblasts and impaired lamina dynamics for the nucleoporin Nup153. Our data thus represent the first study of a direct link between LaA mutant expression and reduced nuclear protein import.

  13. Influence of Operating Conditions on the Removal Cd Ions from Aqueous Media by Adsorption Using Chlamydomonas Reinhardtii

    Institute of Scientific and Technical Information of China (English)

    Jiang Yongbin; Zhu Yi; Ji Hongbing

    2010-01-01

    Chlamydomonas reinhardtii(C.reinhardtii)was used to study adsorption of cadmium(Cd)from aqueous media within various experimental conditions.Results showed that the adsorption process was very fast,with most of the adsorption occurring within30 min of contact time and the equilibrium state was reached in about 60 min.The adsorption ability of the algae increases with the increasing adsorptions sites on cells.Maximum adsorption was observed at the initial Cd concentration of 100 mg/L and pH 6.0.The adsorption was markedly inhibited in the presence of calcium and magnesium ions at 10 mM and the Cd removal efficiency was reduced by 16.54% and 14.99% respectively.This study would be a finding of note with regard to practical wastewater treatment.

  14. Transcriptomic and Physiological Responses of the Green Microalga Chlamydomonas reinhardtii during Short-Term Exposure to Subnanomolar Methylmercury Concentrations.

    Science.gov (United States)

    Beauvais-Flück, Rebecca; Slaveykova, Vera I; Cosio, Claudia

    2016-07-05

    The effects of short-term exposure to subnanomolar methyl-mercury (MeHg) concentrations, representative of contaminated environments, on the microalga Chlamydomonas reinhardtii were assessed using both physiological end points and gene expression analysis. MeHg bioaccumulated and induced significant increase of the photosynthesis efficiency, while the algal growth, oxidative stress, and chlorophyll fluorescence were unaffected. At the molecular level, MeHg significantly dysregulated the expression of genes involved in motility, energy metabolism, lipid metabolism, metal transport, and antioxidant enzymes. Data suggest that the cells were able to cope with subnanomolar MeHg exposure, but this tolerance resulted in a significant cost to the cell energy and reserve metabolism as well as ample changes in the nutrition and motility of C. reinhardtii. The present results allowed gaining new insights on the effects and uptake mechanisms of MeHg at subnanomolar concentrations in aquatic primary producers.

  15. Gestures to Intuitively Control Large Displays

    NARCIS (Netherlands)

    Fikkert, F.W.; Vet, van der P.E.; Rauwerda, H.; Breit, T.; Nijholt, A.; Sales Dias, M.; Gibet, S.; Wanderley, M.W.; Bastos, R.

    2009-01-01

    Large displays are highly suited to support discussions in empirical science. Such displays can display project results on a large digital surface to feed the discussion. This paper describes our approach to closely involve multidisciplinary omics scientists in the design of an intuitive display con

  16. Shewanella oneidensis: a new and efficient System for Expression and Maturation of heterologous [Fe-Fe] Hydrogenase from Chlamydomonas reinhardtii

    Directory of Open Access Journals (Sweden)

    Sybirna Kateryna

    2008-09-01

    Full Text Available Abstract Background The eukaryotic green alga, Chlamydomonas reinhardtii, produces H2 under anaerobic conditions, in a reaction catalysed by a [Fe-Fe] hydrogenase HydA1. For further biochemical and biophysical studies a suitable expression system of this enzyme should be found to overcome its weak expression in the host organism. Two heterologous expression systems used up to now have several advantages. However they are not free from some drawbacks. In this work we use bacterium Shewanella oneidensis as a new and efficient system for expression and maturation of HydA1 from Chlamydomonas reinhardtii. Results Based on codon usage bias and hydrogenase maturation ability, the bacterium S. oneidensis, which possesses putative [Fe-Fe] and [Ni-Fe] hydrogenase operons, was selected as the best potential host for C. reinhardtii [Fe-Fe] hydrogenase expression. Hydrogen formation by S. oneidensis strain AS52 (ΔhydAΔhyaB transformed with a plasmid bearing CrHydA1 and grown in the presence of six different substrates for anaerobic respiration was determined. A significant increase in hydrogen evolution was observed for cells grown in the presence of trimethylamine oxide, dimethylsulfoxide and disodium thiosulfate, showing that the system of S. oneidensis is efficient for heterologous expression of algal [Fe-Fe] hydrogenase. Conclusion In the present work a new efficient system for heterologous expression and maturation of C. reinhardtii hydrogenase has been developed. HydA1 of C. reinhardtii was purified and shown to contain 6 Fe atoms/molecule of protein, as expected. Using DMSO, TMAO or thiosulfate as substrates for anaerobic respiration during the cell growth, 0.4 – 0.5 mg l-1(OD600 = 1 of catalytically active HydA1 was obtained with hydrogen evolution rate of ~700 μmol H2 mg-1 min-1.

  17. Review of Defense Display Research Programs

    Science.gov (United States)

    2001-01-01

    Programs Flat Panel Autostereoscopic N-perspective 3D High Definition DMD Digital Projector Light Piping & Quantum Cavity Displays Solid State Laser...Megapixel Displays • Size Commonality • 67 % Weight Reduction • > 200 sq. in. per Display 20-20 Vision Simulators True 3D , sparse symbols Foldable Display...megapixel 2D and True 3D Display Technology 25M & T3D FY02-FY06 New service thrusts

  18. Recent Trend in Development of Olfactory Displays

    Science.gov (United States)

    Yanagida, Yasuyuki

    An olfactory display is a device that generates scented air with desired concentration of aroma, and delivers it to the user's olfactory organ. In this article, the nature of olfaction is briefly described from the view point of how to configure olfactory displays. Next, component technologies to compose olfactory displays, i.e., making scents and delivering scents, are categorized. Several existing olfactory display systems are introduced to show the current status of research and development of olfactory displays.

  19. Acute effects of a prooxidant herbicide on the microalga Chlamydomonas reinhardtii: Screening cytotoxicity and genotoxicity endpoints

    Energy Technology Data Exchange (ETDEWEB)

    Esperanza, Marta; Cid, Ángeles; Herrero, Concepción; Rioboo, Carmen, E-mail: carmen.rioboo@udc.es

    2015-08-15

    Highlights: • Mitochondrial membrane potential constituted the most sensitive parameter assayed. • Several genotoxicity methods were applied for first time in ecotoxicological studies. • Oxidative DNA base damage (8-OHdG) was induced by paraquat exposure. • Cells with DNA strand breakage and subG1-nuclei increased in treated cultures. • Typical apoptosis hallmarks were observed in microalgal cells exposed to paraquat. - Abstract: Since recent evidence has demonstrated that many types of chemicals exhibit oxidative and/or genotoxic potential on living organisms, reactive oxygen species (ROS) formation and DNA damage are currently the best accepted paradigms to assess the potential hazardous biological effects of a wide range of contaminants. The goal of this study was to evaluate the sensitivity of different cytotoxicity and genotoxicity responses on the model microalga Chlamydomonas reinhardtii exposed to the prooxidant herbicide paraquat. In addition to the growth endpoint, cell viability, mitochondrial membrane potential and presence of reactive oxygen species (ROS) were assayed as potential markers of cytotoxicity using flow cytometry (FCM). To study the effects of paraquat on C. reinhardtii DNA, several genotoxicity approaches were implemented for the first time in an ecotoxicological study on microalgae. Oxidative DNA base damage was analysed by measuring the oxidative DNA lesion 8-OHdG by FCM. DNA fragmentation was analysed by different methods: comet assay, and cell cycle analysis by FCM, with a particular focus on the presence of subG1-nuclei. Finally, effects on morphology of nuclei were monitored through DAPI staining. The evaluation of these endpoints showed that several physiological and biochemical parameters reacted to oxidative stress disturbances with greater sensitivity than integrative parameters such as growth rates or cell viability. The experiments revealed concentration-dependent cytotoxicity (ROS formation, depolarization of

  20. New recombinant cyclohexylamine oxidase variants for deracemization of secondary amines by orthogonally assaying designed mutants with structurally diverse substrates

    Science.gov (United States)

    Li, Guangyue; Yao, Peiyuan; Cong, Peiqian; Ren, Jie; Wang, Lei; Feng, Jinhui; Lau, Peter C. K.; Wu, Qiaqing; Zhu, Dunming

    2016-05-01

    To further expand the substrate range of the cyclohexylamine oxidase (CHAO) from Brevibacterium oxydans, a library of diverse mutants was created and assayed toward a group of structurally diverse substrates. Among them, mutants T198A and M226A exhibited enhanced activity relative to wt CHAO for most (S)-enantiomers of primary amines and some secondary amines. While mutants T198I, L199I, L199F, M226I and M226T were more active than wt CHAO toward the primary amines, mutants T198F, L199T, Y321A, Y321T, Y321I and Y321F enhanced the enzyme activity toward the secondary amines. In particular, mutant Y321I displayed an enhanced catalytic efficiency toward 1-(4-methoxybenzyl)-1, 2, 3, 4, 5, 6, 7, 8-octahydroisoquinoline (13). Whereas a double mutant, Y321I/M226T, acted on (S)-N-(prop-2-yn-1-yl)-2, 3-dihydro-1H-inden-1-amine [(S)-8]. Since (R)-8 is an irreversible inhibitor of monoamine oxidase and (S)-13 is an intermediate of dextromethorphan, a cough suppressant drug, deracemizations of 8 and 13 were carried out with crude enzyme extracts of the respective mutants. This resulted in 51% and 78% isolated yields of (R)-8 and (S)-13, respectively, each with high enantiomeric excess (93% and 99% ee). The results demonstrated the application potential of the evolved CHAO mutants in drug synthesis requiring chiral secondary amines.

  1. 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.

  2. Optical display for radar sensing

    Science.gov (United States)

    Szu, Harold; Hsu, Charles; Willey, Jefferson; Landa, Joseph; Hsieh, Minder; Larsen, Louis V.; Krzywicki, Alan T.; Tran, Binh Q.; Hoekstra, Philip; Dillard, John T.; Krapels, Keith A.; Wardlaw, Michael; Chu, Kai-Dee

    2015-05-01

    Boltzmann headstone S = kB Log W turns out to be the Rosette stone for Greek physics translation optical display of the microwave sensing hieroglyphics. The LHS is the molecular entropy S measuring the degree of uniformity scattering off the sensing cross sections. The RHS is the inverse relationship (equation) predicting the Planck radiation spectral distribution parameterized by the Kelvin temperature T. Use is made of the conservation energy law of the heat capacity of Reservoir (RV) change T Δ S = -ΔE equals to the internal energy change of black box (bb) subsystem. Moreover, an irreversible thermodynamics Δ S > 0 for collision mixing toward totally larger uniformity of heat death, asserted by Boltzmann, that derived the so-called Maxwell-Boltzmann canonical probability. Given the zero boundary condition black box, Planck solved a discrete standing wave eigenstates (equation). Together with the canonical partition function (equation) an average ensemble average of all possible internal energy yielded the celebrated Planck radiation spectral (equation) where the density of states (equation). In summary, given the multispectral sensing data (equation), we applied Lagrange Constraint Neural Network (LCNN) to solve the Blind Sources Separation (BSS) for a set of equivalent bb target temperatures. From the measurements of specific value, slopes and shapes we can fit a set of Kelvin temperatures T's for each bb targets. As a result, we could apply the analytical continuation for each entropy sources along the temperature-unique Planck spectral curves always toward the RGB color temperature display for any sensing probing frequency.

  3. Simulated monitor display for CCTV

    Energy Technology Data Exchange (ETDEWEB)

    Steele, B.J.

    1982-01-01

    Two computer programs have been developed which generate a two-dimensional graphic perspective of the video output produced by a Closed Circuit Television (CCTV) camera. Both programs were primarily written to produce a graphic display simulating the field-of-view (FOV) of a perimeter assessment system as seen on a CCTV monitor. The original program was developed for use on a Tektronix 4054 desktop computer; however, the usefulness of this graphic display program led to the development of a similar program for a Hewlett-Packard 9845B desktop computer. After entry of various input parameters, such as, camera lens and orientation, the programs automatically calculate and graphically plot the locations of various items, e.g., fences, an assessment zone, running men, and intrusion detection sensors. Numerous special effects can be generated to simulate such things as roads, interior walls, or sides of buildings. Other objects can be digitized and entered into permanent memory similar to the running men. With this type of simulated monitor perspective, proposed camera locations with respect to fences and a particular assessment zone can be rapidly evaluated without the costly time delays and expenditures associated with field evaluation.

  4. Boc modifies the holoprosencephaly spectrum of Cdo mutant mice

    Directory of Open Access Journals (Sweden)

    Wei Zhang

    2011-05-01

    Holoprosencephaly (HPE is caused by a failure to form the midline of the forebrain and/or midface. It is one of the most common human birth defects, but clinical expression is extremely variable. HPE is associated with mutations in the sonic hedgehog (SHH pathway. Mice lacking the Shh pathway regulator Cdo (also called Cdon display HPE with strain-dependent penetrance and expressivity, implicating silent modifier genes as one cause of the variability. However, the identities of potential HPE modifiers of this type are unknown. We report here that whereas mice lacking the Cdo paralog Boc do not have HPE, Cdo;Boc double mutants on a largely Cdo-resistant genetic background have lobar HPE with strong craniofacial anomalies and defects in Shh target gene expression in the developing forebrain. Boc is therefore a silent HPE modifier gene in mice. Furthermore, Cdo and Boc have specific, selective roles in Shh signaling in mammals, because Cdo;Boc double-mutant mice do not display the most severe HPE phenotype seen in Shh-null mice, nor do they have major defects in digit patterning or development of vertebrae, which are also Shh-dependent processes. This is in contrast to reported observations in Drosophila, where genetic removal of the Cdo and Boc orthologs Ihog and Boi results in a complete loss of response to the hedgehog ligand. Therefore, there is evolutionary divergence between mammals and insects in the requirement of the hedgehog pathway for Cdo/Ihog family members, with mammalian development involving additional factors and/or distinct mechanisms at this level of pathway regulation.

  5. Águas com predominância de Eutreptia lanowi steuer e Chlamydomonas reinhardi dangeard no plancton, na enseada de Inhauma, Baía de Guanabara

    Directory of Open Access Journals (Sweden)

    Lejeune P. H. de Oliveira

    1962-03-01

    Full Text Available In brackish waters of a creek of Guanabara Bay, the author points by the first time the presence of Chlamydomonas reinhardi, Eutreptia lanowi, Oscillatoria putrida, O. limosa, O. chlorina that were unknown in our waters; such biologic indicators proved themselves pollutional conditions, so bad a stark-mesosaprobic regime. Other news are plankton analysis by the Standar methods, of two most expressive samples of water masses;also the mobility of the plankters are measured in micra by second.

  6. Targeting ESR1-Mutant Breast Cancer

    Science.gov (United States)

    2015-09-01

    AWARD NUMBER: W81XWH-14-1-0359 TITLE: Targeting ESR1-Mutant Breast Cancer PRINCIPAL INVESTIGATOR: Dr. Sarat Chandarlapaty CONTRACTING...31 Aug 2015 4. TITLE AND SUBTITLE Targeting ESR1-Mutant Breast Cancer 5a. CONTRACT NUMBER 5b. GRANT NUMBER W81XWH-14-1-0359 5c. PROGRAM ELEMENT...mutations found in breast cancer using both structural and cell based assays. We have now have evidence for the effects of the most recurrent

  7. Targeting ESR1-Mutant Breast Cancer

    Science.gov (United States)

    2015-09-01

    Introduction Approximately 70% of ER+ breast cancers harbor expression of the estrogen receptor and are dependent upon its activity for various aspects of the...resistance to current FDA approved ER antagonists, but that more potent and selective estrogen receptor antagonists will be sufficiently active to...antagonists and their potency against ER mutants both in vitro and in vivo . Targeting ESR1-Mutant Breast Cancer W81XWH-14-1-0359 9 4. Impact A) Impact

  8. Automated quantification of locomotion, social interaction, and mate preference in Drosophila mutants

    OpenAIRE

    Iyengar, Atulya; Imoehl, Jordan; Ueda, Atsushi; Nirschl, Jeffery; Wu, Chun-Fang

    2012-01-01

    Automated tracking methods facilitate screening for and characterization of abnormal locomotion or more complex behaviors in Drosophila. We developed the Iowa Fly Locomotion and Interaction Tracker (IowaFLI Tracker), a MATLAB based video analysis system, to identify and track multiple flies in a small arena. We report altered motor activity in the K+ and Na+ channel mutants, Hk1 and parats1, which had previously been shown to display abnormal larval locomotion. Environmental factors influenci...

  9. von Hippel-Lindau tumor suppressor mutants faithfully model pathological hypoxia-driven angiogenesis and vascular retinopathies in zebrafish

    NARCIS (Netherlands)

    van Rooijen, E.; Voest, E.E.; Logister, I.; Bussmann, J.; Korving, J.; van Eeden, F.J.; Giles, R.H.; Schulte-Merker, S.

    2010-01-01

    Biallelic inactivation of the von Hippel-Lindau (VHL) tumor suppressor gene predisposes human patients to the development of highly vascularized neoplasms in multiple organ systems. We show that zebrafish vhl mutants display a marked increase in blood vessel formation throughout the embryo, starting

  10. Restoration of self-sustained circadian rhythmicity by the mutant Clock allele in mice in constant illumination

    NARCIS (Netherlands)

    Spoelstra, K; Oklejewicz, M; Daan, S

    2002-01-01

    Mice mutant for the Clock gene display abnormal circadian behavior characterized by long circadian periods and a tendency to become rapidly arrhythmic in constant darkness (DID). To investigate whether this result is contingent on the absence of light, the authors studied the circadian behavior of h

  11. Rolling adhesion of alphaL I domain mutants decorrelated from binding affinity.

    Science.gov (United States)

    Pepper, Lauren R; Hammer, Daniel A; Boder, Eric T

    2006-06-30

    Activated lymphocyte function-associated antigen-1 (LFA-1, alphaLbeta2 integrin) found on leukocytes facilitates firm adhesion to endothelial cell layers by binding to intercellular adhesion molecule-1 (ICAM-1), which is up-regulated on endothelial cells at sites of inflammation. Recent work has shown that LFA-1 in a pre-activation, low-affinity state may also be involved in the initial tethering and rolling phase of the adhesion cascade. The inserted (I) domain of LFA-1 contains the ligand-binding epitope of the molecule, and a conformational change in this region during activation increases ligand affinity. We have displayed wild-type I domain on the surface of yeast and validated expression using I domain specific antibodies and flow cytometry. Surface display of I domain supports yeast rolling on ICAM-1-coated surfaces under shear flow. Expression of a locked open, high-affinity I domain mutant supports firm adhesion of yeast, while yeast displaying intermediate-affinity I domain mutants exhibit a range of rolling phenotypes. We find that rolling behavior for these mutants fails to correlate with ligand binding affinity. These results indicate that unstressed binding affinity is not the only molecular property that determines adhesive behavior under shear flow.

  12. Simplified Night Sky Display System

    Science.gov (United States)

    Castellano, Timothy P.

    2010-01-01

    A document describes a simple night sky display system that is portable, lightweight, and includes, at most, four components in its simplest configuration. The total volume of this system is no more than 10(sup 6) cm(sup 3) in a disassembled state, and weighs no more than 20 kilograms. The four basic components are a computer, a projector, a spherical light-reflecting first surface and mount, and a spherical second surface for display. The computer has temporary or permanent memory that contains at least one signal representing one or more images of a portion of the sky when viewed from an arbitrary position, and at a selected time. The first surface reflector is spherical and receives and reflects the image from the projector onto the second surface, which is shaped like a hemisphere. This system may be used to simulate selected portions of the night sky, preserving the appearance and kinesthetic sense of the celestial sphere surrounding the Earth or any other point in space. These points will then show motions of planets, stars, galaxies, nebulae, and comets that are visible from that position. The images may be motionless, or move with the passage of time. The array of images presented, and vantage points in space, are limited only by the computer software that is available, or can be developed. An optional approach is to have the screen (second surface) self-inflate by means of gas within the enclosed volume, and then self-regulate that gas in order to support itself without any other mechanical support.

  13. OZ: An Innovative Primary Flight Display Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The proposed SBIR project will develop OZ, an innovative primary flight display for aircraft. The OZ display, designed from "first principles" of vision science,...

  14. Holographic Waveguided See-Through Display Project

    Data.gov (United States)

    National Aeronautics and Space Administration — To address the NASA need for lightweight, space suit-mounted displays, Luminit proposes a novel Holographic Waveguided See-Through Display. Our proposed Holographic...

  15. ProPath Display Process Overview

    Data.gov (United States)

    Department of Veterans Affairs — This API displays an overview of the process including the description, goals, associated roles (linked to their detailed information). It also displays all of the...

  16. Projection/Reflection Heads-up Display

    Data.gov (United States)

    National Aeronautics and Space Administration — To address the NASA need for an EVA information display device, Physical Optics Corporation (POC) proposes to develop a new Projection/Reflection Heads-up Display...

  17. Mutational biosynthesis of neomycin analogs by a mutant of neomycin-producing Streptomyces fradiae.

    Science.gov (United States)

    Shi, Guanying; Zhang, Xingang; Wu, Lang; Xie, Jin; Tao, Ke; Hou, Taiping

    2011-11-01

    Neomycin, produced by Streptomyces fradiae, has been widely used for the treatment of bacterial infections in clinical and agricultural applications. In this study, a neomycin nonproducing mutant of S. fradiae was obtained by gene disruption technique for mutational biosynthesis. A crucial gene neoC (neo7) which encodes 2-deoxystreptamine (2-DOS) synthases was disrupted. The mutant could resume producing neomycin in the presence of 2-DOS. Salen derivatives of 2-DOS were synthesized and individually added to cultures of the mutant. Antibacterial activity of the mutasynthesis products against Staphylococcus aureus and four plant pathogenic bacteria (Pseudomonas solanacarum, Erwinia carotovora, Xanthomonas oryzae, and Xanthomonas campestris) was detected quantitatively by Oxford cup method. It is suggested that all 2-DOS derivatives were incorporated by the mutant into new active neomycin analogs except for 2-DOS derivative 2d ((1R,2r,3S,4R,6S)-4,6-bis((E)-3,5-di-tert-butyl-2-hydroxybenzylideneamino)cyclohexane-1,2,3-triol). Neomycin analogs produced by feeding 2-DOS derivative 2a ((1R,2r,3S,4R,6S)-4,6-bis((E)-2 hydroxybenzylideneamino)cyclohexane-1,2,3-triol) to cultures of the mutant displayed a similar antibacterial activity with neomycin produced by wild strain.

  18. Neurobehavioral performances and brain regional metabolism in Dab1(scm) (scrambler) mutant mice.

    Science.gov (United States)

    Jacquelin, C; Lalonde, R; Jantzen-Ossola, C; Strazielle, C

    2013-09-01

    As disabled-1 (DAB1) protein acts downstream in the reelin signaling pathway modulating neuronal migration, glutamate neurotransmission, and cytoskeletal function, the disabled-1 gene mutation (scrambler or Dab1(scm) mutation) results in ataxic mice displaying dramatic neuroanatomical defects similar to those observed in the reeler gene (Reln) mutation. By comparison to non-ataxic controls, Dab1(scm) mutants showed severe motor coordination impairments on stationary beam, coat-hanger, and rotorod tests but were more active in the open-field. Dab1(scm) mutants were also less anxious in the elevated plus-maze but with higher latencies in the emergence test. In mutants versus controls, changes in regional brain metabolism as measured by cytochrome oxidase (COX) activity occurred mainly in structures intimately connected with the cerebellum, in basal ganglia, in limbic regions, particularly hippocampus, as well as in visual and parietal sensory cortices. Although behavioral results characterized a major cerebellar disorder in the Dab1(scm) mutants, motor activity impairments in the open-field were associated with COX activity changes in efferent basal ganglia structures such as the substantia nigra, pars reticulata. Metabolic changes in this structure were also associated with the anxiety changes observed in the elevated plus-maze and emergence test. These results indicate a crucial participation of the basal ganglia in the functional phenotype of ataxic Dab1(scm) mutants.

  19. Genetic analysis and preliminary gene mapping of a novel seeding yellow leaf mutant in rice

    Directory of Open Access Journals (Sweden)

    GE Shaoxing

    2013-02-01

    Full Text Available A novel seeding yellow-leaf mutant, syl11(seedling yellow leaf 11,was found in rice mutant pool from japonica variety,Nipponbare,treated by 60Coγ-radiation.In comparison with the wild-type parent (Nipponbare,the mutant displayed the phenotype of yellow on the 2nd and 3rd leaf and began to turn green from its top before they completely expanded besides the significantly decreased contents of photosynthetic pigments,but all leaves become normal green after the 4-leaf-old stage.The genetic analysis indicated that the mutant trait was controlled by a single recessive nuclear gene.In addition,with the help of SSR,new developed InDel markers and F2 segregating population derived from the cross between the syl11 mutant and Peiai64S,the syl11 was mapped between RM26652(on the long armand ID11974(closing to the centromereon chromosome 11,with a genetic distance of 0.5cM and 0.7cM,respectively.

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

    Directory of Open Access Journals (Sweden)

    Baier Margarete

    2010-06-01

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

  1. A Novel Two-Step Method for Screening Shade Tolerant Mutant Plants via Dwarfism

    Science.gov (United States)

    Li, Wei; Katin-Grazzini, Lorenzo; Krishnan, Sanalkumar; Thammina, Chandra; El-Tanbouly, Rania; Yer, Huseyin; Merewitz, Emily; Guillard, Karl; Inguagiato, John; McAvoy, Richard J.; Liu, Zongrang; Li, Yi

    2016-01-01

    When subjected to shade, plants undergo rapid shoot elongation, which often makes them more prone to disease and mechanical damage. Shade-tolerant plants can be difficult to breed; however, they offer a substantial benefit over other varieties in low-light areas. Although perennial ryegrass (Lolium perenne L.) is a popular species of turf grasses because of their good appearance and fast establishment, the plant normally does not perform well under shade conditions. It has been reported that, in turfgrass, induced dwarfism can enhance shade tolerance. Here we describe a two-step procedure for isolating shade tolerant mutants of perennial ryegrass by first screening for dominant dwarf mutants, and then screening dwarf plants for shade tolerance. The two-step screening process to isolate shade tolerant mutants can be done efficiently with limited space at early seedling stages, which enables quick and efficient isolation of shade tolerant mutants, and thus facilitates development of shade tolerant new cultivars of turfgrasses. Using the method, we isolated 136 dwarf mutants from 300,000 mutagenized seeds, with 65 being shade tolerant (0.022%). When screened directly for shade tolerance, we recovered only four mutants from a population of 150,000 (0.003%) mutagenized seeds. One shade tolerant mutant, shadow-1, was characterized in detail. In addition to dwarfism, shadow-1 and its sexual progeny displayed high degrees of tolerance to both natural and artificial shade. We showed that endogenous gibberellin (GA) content in shadow-1 was higher than wild-type controls, and shadow-1 was also partially GA insensitive. Our novel, simple and effective two-step screening method should be applicable to breeding shade tolerant cultivars of turfgrasses, ground covers, and other economically important crop plants that can be used under canopies of existing vegetation to increase productivity per unit area of land. PMID:27752260

  2. Functional Rescue of Trafficking-Impaired ABCB4 Mutants by Chemical Chaperones.

    Directory of Open Access Journals (Sweden)

    Raquel Gordo-Gilart

    Full Text Available Multidrug resistance protein 3 (MDR3, ABCB4 is a hepatocellular membrane protein that mediates biliary secretion of phosphatidylcholine. Null mutations in ABCB4 gene give rise to severe early-onset cholestatic liver disease. We have previously shown that the disease-associated mutations p.G68R, p.G228R, p.D459H, and p.A934T resulted in retention of ABCB4 in the endoplasmic reticulum, thus failing to target the plasma membrane. In the present study, we tested the ability of two compounds with chaperone-like activity, 4-phenylbutyrate and curcumin, to rescue these ABCB4 mutants by assessing their effects on subcellular localization, protein maturation, and phospholipid efflux capability. Incubation of transfected cells at a reduced temperature (30°C or exposure to pharmacological doses of either 4-PBA or curcumin restored cell surface expression of mutants G228R and A934T. The delivery of these mutants to the plasma membrane was accompanied by a switch in the ratio of mature to inmature protein forms, leading to a predominant expression of the mature protein. This effect was due to an improvement in the maturation rate and not to the stabilization of the mature forms. Both mutants were also functionally rescued, displaying bile salt-dependent phospholipid efflux activity after addition of 4-PBA or curcumin. Drug-induced rescue was mutant specific, given neither 4-PBA nor curcumin had an effect on the ABCB4 mutants G68R and A934T. Collectively, these data indicate that the functionality of selected trafficking-defective ABCB4 mutants can be recovered by chemical chaperones through restoration of membrane localization, suggesting a potential treatment for patients carrying such mutations.

  3. Reconfigurable Full-Page Braille Displays

    Science.gov (United States)

    Garner, H. Douglas

    1994-01-01

    Electrically actuated braille display cells of proposed type arrayed together to form full-page braille displays. Like other braille display cells, these provide changeable patterns of bumps driven by digitally recorded text stored on magnetic tapes or in solid-state electronic memories. Proposed cells contain electrorheological fluid. Viscosity of such fluid increases in strong electrostatic field.

  4. 27 CFR 6.83 - Product displays.

    Science.gov (United States)

    2010-04-01

    ... industry member of giving or selling product displays to a retailer does not constitute a means to induce... costs are excluded. (2) All product displays must bear conspicuous and substantial advertising matter on... address of the retailer may appear on the product displays. (3) The giving or selling of such...

  5. Emerging Large-Screen Display Technology

    Science.gov (United States)

    1992-11-01

    1255, Santa Clara, CA. 25. Williams, R. D., and F. Garcia, 1988, "A Real Time Autostereoscopic Multiplanar 3D Display System," Society for Information...K. Miyaji, 1989, " 3D Display using Laser and Moving Screen, Japan Display 1989, Paper P3-5. 27. Sterling, R. D., R. D. TeKolste, J. M. Haggerty, T. C

  6. Phanerochaete mutants with enhanced ligninolytic activity

    Energy Technology Data Exchange (ETDEWEB)

    Kakar, S.N.; Perez, A.; Gonzales, J.

    1993-06-01

    In addition to lignin, the white rot fungus Phanerochaete chrysosporium has the ability to degrade a wide spectrum of recalcitrant organopollutants in soils and aqueous media. Although some of the organic compounds are degraded under nonligninolytic conditions, most are degraded under ligninolytic conditions with the involvement of the extracellular enzymes, lignin peroxidases, and manganese-dependent peroxidases, which are produced as secondary metabolites triggered by conditions of nutrient starvation (e.g., nitrogen limitation). The fungus and its enzymes can thus provide alternative technologies for bioremediation, biopulping, biobleaching, and other industrial applications. The efficiency and effectiveness of the fungus can be enhanced by increasing production and secretion of the important enzymes in large quantities and as primary metabolites under enriched conditions. One way this can be achieved is through isolation of mutants that are deregulated or are hyperproducers or supersecretors of key enzymes under enriched conditions. Through ultraviolet-light and gamma-rays mutagenesis we have isolated a variety of mutants, some of which produce key enzymes of the ligninolytic system under high-nitrogen growth conditions. One of the mutants produced 272 units (U) of lignin peroxidases enzyme activity per liter after nine days under high nitrogen. The mutant and the parent strains produced up to 54 U/L and 62 U/L, respectively, of the enzyme activity under low-nitrogen growth conditions during this period. In some experiments the mutant showed 281 U/L of enzyme activity under high nitrogen after 17 days.

  7. New steps toward understanding the regulation of photosystem i biogenesis in chlamydomonas reinhardtii

    OpenAIRE

    Douchi, Damien

    2016-01-01

    Cette thèse décrit l’identification de MAC1, une protéine nouvellement identifiée encodée par le génome nucléaire, impliquée dans la stabilisation de l’ARN messager de psaC, sous-unité du photosystème I. L’identification de ce gène a été possible suite à l’isolation d’un mutant d’insertion dont le génotypage par PCR inverse qui a caractérisé le gène affecté. Ce gène code pour MAC1 qui est une protéine de 100 kDa environ contenant des répétitions de type « Tetratricopeptide repeats », TPR ou H...

  8. Disrupting ER-associated protein degradation suppresses the abscission defect of a weak hae hsl2 mutant in Arabidopsis

    Science.gov (United States)

    Baer, John; Taylor, Isaiah; Walker, John C.

    2016-01-01

    In Arabidopsis thaliana, the process of abscission, or the shedding of unwanted organs, is mediated by two genes, HAESA (HAE) and HAESA-LIKE 2 (HSL2), encoding receptor-like protein kinases (RLKs). The double loss-of-function mutant hae-3 hsl2-3 is completely deficient in floral abscission, but, interestingly, the hae-3 hsl2-9 mutant displays a less severe defect. This mutant was chosen for an ethyl methanesulfonate (EMS) screen to isolate enhancer and suppressor mutants, and two such suppressors are the focus of this study. Pooled DNA from the F2 generation of a parental backcross was analyzed by genome sequencing to reveal candidate genes, two of which complement the suppressor phenotype. These genes, EMS-MUTAGENIZED BRI1 SUPPRESSOR 3 (EBS3) and EBS4, both encode mannosyltransferases involved in endoplasmic reticulum (ER)-associated degradation (ERAD) of proteins. Further analysis of these suppressor lines revealed that suppressor mutations are acting solely on the partially functional hsl2-9 mutant receptor to modify the abscission phenotype. Expressing a hsl2-9–yellow fluorescent protein (YFP) transgene in ebs3 mutants yields a higher fluorescent signal than in EBS3/ebs3, suggesting that these mutants restore abscission by disrupting ERAD to allow accumulation of the hsl2-9 receptor, which probably escapes degradation to be trafficked to the plasma membrane to regain signaling. PMID:27566817

  9. CERN students display their work

    CERN Multimedia

    Anaïs Schaeffer

    2011-01-01

    The first poster session by students working on the LHC experiments, organised by the LPCC, was a great success. Showcasing the talents of over a hundred young physicists from all over the world, it was an opportunity for everyone at CERN to check out the wide range of research work being done by the new generation of physicists at CERN.   At 5.30 p.m. on Wednesday 23 March, the first poster session by CERN students took place in Restaurant No.1, where no fewer than 87 posters went on public display. The students were split into 8 groups according to their research field* and all were on hand to answer the questions of an inquisitive audience. TH Department's Michelangelo Mangano, who is head of the LHC Physics Centre at CERN (LPCC) and is responsible for the initiative, confirms that nothing was left to chance, even the choice of date: "We wanted to make the most of the general enthusiasm around the winter conferences and the meeting of the LHC Experiments Committee to present the stud...

  10. Citizenship displayed by disabled people

    Directory of Open Access Journals (Sweden)

    Eliana Prado Carlino

    2010-12-01

    Full Text Available By investigating the processes by which successful teachers become activate citizens and by listening to the diversity and richness of their life and formation stories, this work became possible. Its aim is to display some of the utterances of two Down Syndrome individuals and their active-citizenship activities. Their stories were told in the reports of two teachers when describing their personal and professional history, and were considered to be an integral part of it. Thus, some of the utterances and perceptions with which these two individuals elaborate their references, their worldview and their active-citizenship activity are evidenced in this paper. This article is based on the language conceptions of Vygotsky and Bakhtin who defend the idea that the group and the social mentality are ingrain in the individual. Hence, the history of one person reveals that of many others, since there is a deep link between the individual and the social in the formation of a subjective worldview. As a result, it can be easily seen that the utterances expressed by the participants in this research cannot be considered strictly individual because enunciation is social in nature. Despite the fact that the utterances are those of individuals, they manifest a collective reality. This demonstrates the real advantages and possibilities that deficient people get from their participation and intervention in society.

  11. ABI domain-containing proteins contribute to surface protein display and cell division in Staphylococcus aureus.

    Science.gov (United States)

    Frankel, Matthew B; Wojcik, Brandon M; DeDent, Andrea C; Missiakas, Dominique M; Schneewind, Olaf

    2010-10-01

    The human pathogen Staphylococcus aureus requires cell wall anchored surface proteins to cause disease. During cell division, surface proteins with YSIRK signal peptides are secreted into the cross-wall, a layer of newly synthesized peptidoglycan between separating daughter cells. The molecular determinants for the trafficking of surface proteins are, however, still unknown. We screened mutants with non-redundant transposon insertions by fluorescence-activated cell sorting for reduced deposition of protein A (SpA) into the staphylococcal envelope. Three mutants, each of which harboured transposon insertions in genes for transmembrane proteins, displayed greatly reduced envelope abundance of SpA and surface proteins with YSIRK signal peptides. Characterization of the corresponding mutations identified three transmembrane proteins with abortive infectivity (ABI) domains, elements first described in lactococci for their role in phage exclusion. Mutations in genes for ABI domain proteins, designated spdA, spdB and spdC (surface protein display), diminish the expression of surface proteins with YSIRK signal peptides, but not of precursor proteins with conventional signal peptides. spdA, spdB and spdC mutants display an increase in the thickness of cross-walls and in the relative abundance of staphylococci with cross-walls, suggesting that spd mutations may represent a possible link between staphylococcal cell division and protein secretion.

  12. Limitations of yeast surface display in engineering proteins of high thermostability.

    Science.gov (United States)

    Park, Sheldon; Xu, Yao; Stowell, Xiaoran Fu; Gai, Feng; Saven, Jeffery G; Boder, Eric T

    2006-05-01

    Engineering proteins that can fold to unique structures remains a challenge. Protein stability has previously been engineered via the observed correlation between thermal stability and eukaryotic secretion level. To explore the limits of an expression-based approach, variants of the highly thermostable three-helix bundle protein alpha3D were studied using yeast surface display. A library of alpha3D mutants was created to explore the possible correlation of protein stability and fold with expression level. Five efficiently expressed mutants were then purified and further studied biochemically. Despite their differences in stability, most mutants expressed at levels comparable with that of wild-type alpha3D. Two other related sequences (alpha3A and alpha3B) that form collapsed, stable molten globules but lack a uniquely folded structure were similarly expressed at high levels by yeast display. Together these observations suggest that the quality control system in yeast is unable to discriminate between well-folded proteins of high stability and molten globules. The present study, therefore, suggests that an optimization of the surface display efficiency on yeast may yield proteins that are thermally and chemically stable yet are poorly folded.

  13. Enhanced cellulase production in mutants of Thermomonospora

    Energy Technology Data Exchange (ETDEWEB)

    Fennington, G.; Lupo, D.; Stutzenberger, F.

    1982-01-01

    Thermomonospora curvata, a thermophilic actinomycete, secretes multiple forms of endo-beta, 1-4-glucanase (EG) when grown on cellulose-mineral salts liquid medium. The EG activity (measured as carboxymethyl cellulose hydrolysis) was separated by ion exchange chromatography into three distinct components which differed in their kinetic properties. Exposure of T. curvata to ultraviolet light, N-nitrosoguanidine, or ethane methyl sulfonate produced mutants with enhanced EG production. Selection of colonies which cleared cellulose agar plants containing 2-deoxyglucose or glycerol yielded mutants having 1.5 to 2.6 times the extracellular EG and saccharifying activity (measured by filter-paper and cotton-fiber hydrolysis). The secretion of extracellular protein was increased proportionally in mutant cultures. (Refs. 40).

  14. Aging Kit mutant mice develop cardiomyopathy.

    Directory of Open Access Journals (Sweden)

    Lei Ye

    Full Text Available Both bone marrow (BM and myocardium contain progenitor cells expressing the c-Kit tyrosine kinase. The aims of this study were to determine the effects of c-Kit mutations on: i. myocardial c-Kit(+ cells counts and ii. the stability of left ventricular (LV contractile function and structure during aging. LV structure and contractile function were evaluated (echocardiography in two groups of Kit mutant (W/Wv and W41/W42 and in wild type (WT mice at 4 and 12 months of age and the effects of the mutations on LV mass, vascular density and the numbers of proliferating cells were also determined. In 4 month old Kit mutant and WT mice, LV ejection fractions (EF and LV fractional shortening rates (FS were comparable. At 12 months of age EF and FS were significantly decreased and LV mass was significantly increased only in W41/W42 mice. Myocardial vascular densities and c-Kit(+ cell numbers were significantly reduced in both mutant groups when compared to WT hearts. Replacement of mutant BM with WT BM at 4 months of age did not prevent these abnormalities in either mutant group although they were somewhat attenuated in the W/Wv group. Notably BM transplantation did not prevent the development of cardiomyopathy in 12 month W41/W42 mice. The data suggest that decreased numbers and functional capacities of c-Kit(+ cardiac resident progenitor cells may be the basis of the cardiomyopathy in W41/W42 mice and although defects in mutant BM progenitor cells may prove to be contributory, they are not causal.

  15. Behavioral characterization of system xc- mutant mice.

    Science.gov (United States)

    McCullagh, Elizabeth A; Featherstone, David E

    2014-05-15

    The slc7a11 gene encodes xCT, an essential component of 'system xc-', a plasma membrane exchanger that imports cystine and exports glutamate. Slc7a11 is expressed primarily in the brain, but its role there is not clear. We performed behavioral tests on two different strains of homozygous slc7a11 mutant mice ('sut' and 'xCT'), as well as heteroallelic offspring of these two strains ('xCT/sut') and their associated genetic backgrounds. Homozygous sut mutant males showed reduced spontaneous alternation in spontaneous alternation tasks as well as reduced movement in an open field maze, but xCT and xCT/sut strains did not show significant changes in these tasks compared to appropriate controls. Neither xCT nor sut mutants showed differences from controls in rotarod tests. Female behavioral phenotypes were independent of estrus cycle stage. To ensure that homozygous xCT, sut, and xCT/sut strains all represent protein null alleles, we measured whole brain xCT protein levels using immunoblots. xCT, sut and xCT/sut strains showed no detectable xCT protein expression, confirming them as null alleles. Previously published microdialysis experiments showed reduced striatal glutamate in xCT mutants. Using the same methods, we measured reduced interstitial glutamate levels in the striatum but not cerebellum of sut mutants. However, we detected no glutamate change in the striatum or cerebellum of sut/xCT mice. We detected no changes in whole brain EAAT-1, -2, or -3 expression. We conclude that the behavioral and chemical differences exist between slc7a11 mutant strains, but we were unable to definitively attribute any of these differences to loss of system xc-.

  16. Class II integrase mutants with changes in putative nuclear localization signals are primarily blocked at a postnuclear entry step of human immunodeficiency virus type 1 replication.

    Science.gov (United States)

    Lu, Richard; Limón, Ana; Devroe, Eric; Silver, Pamela A; Cherepanov, Peter; Engelman, Alan

    2004-12-01

    Integrase has been implicated in human immunodeficiency virus type 1 (HIV-1) nuclear import. Integrase analyses, however, can be complicated by the pleiotropic nature of mutations: whereas class I mutants are integration defective, class II mutants display additional assembly and/or reverse transcription defects. We previously determined that HIV-1(V165A), originally reported as defective for nuclear import, was a class II mutant. Here we analyzed mutants containing changes in other putative nuclear localization signals, including (186)KRK(188)/(211)KELQKQITK(219) and Cys-130. Previous work established HIV-1(K186Q), HIV-1(Q214L/Q216L), and HIV-1(C130G) as replication defective, but phenotypic classification was unclear and nuclear import in nondividing cells was not addressed. Consistent with previous reports, most of the bipartite mutants studied here were replication defective. These mutants as well as HIV-1(V165A) synthesized reduced cDNA levels, but a normal fraction of mutant cDNA localized to dividing and nondividing cell nuclei. Somewhat surprisingly, recombinant class II mutant proteins were catalytically active, and class II Vpr-integrase fusion proteins efficiently complemented class I mutant virus. Since a class I Vpr-integrase mutant efficiently complemented class II mutant viruses under conditions in which class II Vpr-integrases failed to function, we conclude that classes I and II define two distinct complementation groups and suggest that class II mutants are primarily defective at a postnuclear entry step of HIV-1 replication. HIV-1(C130G) was also defective for reverse transcription, but Vpr-integrase(C130G) did not efficiently complement class I mutant HIV-1. Since HIV-1(C130A) grew like the wild type, we conclude that Cys-130 is not essential for replication and speculate that perturbation of integrase structure contributed to the pleiotropic HIV-1(C130G) phenotype.

  17. Military display market segment: wearable and portable

    Science.gov (United States)

    Desjardins, Daniel D.; Hopper, Darrel G.

    2003-09-01

    The military display market (MDM) is analyzed in terms of one of its segments, wearable and portable displays. Wearable and portable displays are those embedded in gear worn or carried by warfighters. Categories include hand-mobile (direct-view and monocular/binocular), palm-held, head/helmet-mounted, body-strapped, knee-attached, lap-born, neck-lanyard, and pocket/backpack-stowed. Some 62 fielded and developmental display sizes are identified in this wearable/portable MDM segment. Parameters requiring special consideration, such as weight, luminance ranges, light emission, viewing angles, and chromaticity coordinates, are summarized and compared. Ruggedized commercial versus commercial off-the-shelf designs are contrasted; and a number of custom displays are also found in this MDM category. Display sizes having aggregate quantities of 5,000 units or greater or having 2 or more program applications are identified. Wearable and portable displays are also analyzed by technology (LCD, LED, CRT, OLED and plasma). The technical specifications and program history of several high-profile military programs are discussed to provide a systems context for some representative displays and their function. As of August 2002 our defense-wide military display market study has documented 438,882 total display units distributed across 1,163 display sizes and 438 weapon systems. Wearable and portable displays account for 202,593 displays (46% of total DoD) yet comprise just 62 sizes (5% of total DoD) in 120 weapons systems (27% of total DoD). Some 66% of these wearable and portable applications involve low information content displays comprising just a few characters in one color; however, there is an accelerating trend towards higher information content units capable of showing changeable graphics, color and video.

  18. Microspheres in Plasma Display Panels

    Science.gov (United States)

    2006-01-01

    Filling small bubbles of molten glass with gases is just as difficult as it sounds, but the technical staff at NASA is not known to shy away from a difficult task. When Microsphere Systems, Inc. (MSI), of Ypsilanti, Michigan, and Imaging Systems Technology, Inc. (IST), of Toledo, Ohio, were trying to push the limits of plasma displays but were having difficulty with the designs, NASA s Glenn Garrett Morgan Commercialization Initiative (GMCI) assembled key personnel at Glenn Research Center and Ohio State University for a brainstorming session to come up with a solution for the companies. They needed a system that could produce hollow, glass micro-sized spheres (microspheres) that could be filled with a variety of gasses. But the extremely high temperature required to force the micro-sized glass bubbles to form at the tip of a metal nozzle resulted in severe discoloration of the microspheres. After countless experiments on various glass-metal combinations, they had turned to the GMCI for help. NASA experts in advanced metals, ceramics, and glass concluded that a new design approach was necessary. The team determined that what was needed was a phosphate glass composition that would remain transparent, and they went to work on a solution. Six weeks later, using the design tips from the NASA team, Tim Henderson, president of MSI, had designed a new system in which all surfaces in contact with the molten glass would be ceramic instead of metal. Meanwhile, IST was able to complete a Phase I Small Business Innovation Research (SBIR) grant supported by the National Science Foundation (NSF) and supply a potential customer with samples of the microspheres for evaluation as filler materials for high-performance insulations.

  19. Changes in Stoichiometry, Cellular RNA, and Alkaline Phosphatase Activity of Chlamydomonas in Response to Temperature and Nutrients

    Science.gov (United States)

    Hessen, Dag O.; Hafslund, Ola T.; Andersen, Tom; Broch, Catharina; Shala, Nita K.; Wojewodzic, Marcin W.

    2017-01-01

    Phytoplankton may respond both to elevated temperatures and reduced nutrients by changing their cellular stoichiometry and cell sizes. Since increased temperatures often cause increased thermal stratification and reduced vertical flux of nutrients into the mixed zone, it is difficult to disentangle these drivers in nature. In this study, we used a factorial design with high and low levels of phosphorus (P) and high and low temperature to assess responses in cellular stoichiometry, levels of RNA, and alkaline phosphatase activity (APA) in the chlorophyte Chlamydomonas reinhardtii. Growth rate, C:P, C:N, N:P, RNA, and APA all responded primarily to P treatment, but except for N:P and APA, also temperature contributed significantly. For RNA, the contribution from temperature was particularly strong with higher cellular levels of RNA at low temperatures, suggesting a compensatory allocation to ribosomes to maintain protein synthesis and growth. These experiments suggest that although P-limitation is the major determinant of growth rate and cellular stoichiometry, there are pronounced effects of temperature also via interaction with P. At the ecosystem level, nutrients and temperature will thus interact, but temperatures would likely exert a stronger impact on these phytoplankton traits indirectly via its force on stratification regimes and vertical nutrient fluxes. PMID:28167934

  20. Flux balance analysis reveals acetate metabolism modulates cyclic electron flow and alternative glycolytic pathways in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Chapman, Stephen P; Paget, Caroline M; Johnson, Giles N; Schwartz, Jean-Marc

    2015-01-01

    Cells of the green alga Chlamydomonas reinhardtii cultured in the presence of acetate perform mixotrophic growth, involving both photosynthesis and organic carbon assimilation. Under such conditions, cells exhibit a reduced capacity for photosynthesis but a higher growth rate, compared to phototrophic cultures. Better understanding of the down regulation of photosynthesis would enable more efficient conversion of carbon into valuable products like biofuels. In this study, Flux Balance Analysis (FBA) and Flux Variability Analysis (FVA) have been used with a genome scale model of C. reinhardtii to examine changes in intracellular flux distribution in order to explain their changing physiology. Additionally, a reaction essentiality analysis was performed to identify which reaction subsets are essential for a given growth condition. Our results suggest that exogenous acetate feeds into a modified tricarboxylic acid (TCA) cycle, which bypasses the CO2 evolution steps, explaining increases in biomass, consistent with experimental data. In addition, reactions of the oxidative pentose phosphate and glycolysis pathways, inactive under phototrophic conditions, show substantial flux under mixotrophic conditions. Importantly, acetate addition leads to an increased flux through cyclic electron flow (CEF), but results in a repression of CO2 fixation via Rubisco, explaining the down regulation of photosynthesis. However, although CEF enhances growth on acetate, it is not essential-impairment of CEF results in alternative metabolic pathways being increased. We have demonstrated how the reactions of photosynthesis interconnect with carbon metabolism on a global scale, and how systems approaches play a viable tool in understanding complex relationships at the scale of the organism.

  1. Molecular toxicity of cerium oxide nanoparticles to the freshwater alga Chlamydomonas reinhardtii is associated with supra-environmental exposure concentrations.

    Science.gov (United States)

    Taylor, Nadine S; Merrifield, Ruth; Williams, Tim D; Chipman, J Kevin; Lead, Jamie R; Viant, Mark R

    2016-01-01

    Ceria nanoparticles (NPs) are widely used as fuel catalysts and consequently are likely to enter the environment. Their potential impacts on. biota at environmentally relevant concentrations, including uptake and toxicity, remain to be elucidated and quantitative data on which to assess risk are sparse. Therefore, a definitive assessment of the molecular and phenotypic effects of ceria NPs was undertaken, using well-characterised mono-dispersed NPs as their toxicity is likely to be higher, enabling a conservative hazard assessment. Unbiased transcriptomics and metabolomics approaches were used to investigate the potential toxicity of tightly constrained 4-5 nm ceria NPs to the unicellular green alga, Chlamydomonas reinhardtii, a sentinel freshwater species. A wide range of exposure concentrations were investigated from predicted environmental levels, to support hazard assessment, to supra-environmental levels to provide insight into molecular toxicity pathways. Ceria NPs were internalised into intracellular vesicles within C. reinhardtii, yet caused no significant effect on algal growth at any exposure concentration. Molecular perturbations were only detected at supra-environmental ceria NP-concentrations, primarily down-regulation of photosynthesis and carbon fixation with associated effects on energy metabolism. For acute exposures to small mono-dispersed particles, it can be concluded there should be little concern regarding their dispersal into the environment for this trophic level.

  2. Effects of sodium sulfate on the freshwater microalga Chlamydomonas moewusii: implications for the optimization of algal culture media.

    Science.gov (United States)

    Mera, Roi; Torres, Enrique; Abalde, Julio

    2016-02-01

    The study of the microalgal growth kinetics is an indispensable tool in all fields of phycology. Knowing the optimal nutrient concentration is an important issue that will help to develop efficient growth systems for these microorganisms. Although nitrogen and phosphorus are well studied for this purpose, sulfur seems to be less investigated. Sulfate is a primary sulfur source used by microalgae; moreover, the concentration of this compound is increasing in freshwater systems due to pollution. The aim of this study was to investigate the effects of different sodium sulfate concentrations in the culture medium on growth and growth kinetics of the freshwater microalga Chlamydomonas moewusii. Production of biomass, chl content, kinetic equations, and a mathematical model that describe the microalgal growth in relation with the concentration of sodium sulfate were obtained. The lowest concentration of sodium sulfate allowing optimal growth was 0.1 mM. Concentrations higher than 3 mM generated a toxic effect. This work demonstrates that this toxic effect was not directly due to the excess of sulfate ion but by the elevation of the ionic strength. An inhibition model was successfully used to simulate the relationship between specific growth rate and sodium sulfate in this microalga.

  3. Green algal cytochrome b6-f complexes: isolation and characterization from Dunaliella saline, Chlamydomonas reinhardtii and Scenedesmus obliquus.

    Science.gov (United States)

    Wynn, R M; Bertsch, J; Bruce, B D; Malkin, R

    1988-09-14

    Cytochrome b6-f complexes have been isolated from Chlamydomonas reinhardtii, Dunaliella saline and Scenedesmus obliquus. Each complex is essentially free of chlorophyll and carotenoids and contains cytochrome b6 and cytochrome f hemes in a 2:1 molar ratio. C. reinhardtii and S. obliquus complexes contain the Rieske iron-sulfur protein (present in approx 1:1 molar ratio to cytochrome f) and each catalyzes a DBMIB- and DNP-INT-sensitive electron transfer from duroquinol to spinach plastocyanin. Immunological assays using antibodies to the peptides from the spinach cytochrome complex show varying cross-reactivity patterns except for the complete absence of binding to the Rieske proteins in any of the three complexes, suggesting little structural similarity between the Rieske proteins of algae with those from higher plants. One complex (D. salina) has been uniformly labeled by growth in NaH14CO3 to determine stoichiometries of constituent polypeptide subunits. Results from these studies indicate that all functionally active cytochrome b6-f complexes contain four subunits which occur in equimolar amounts.

  4. A factor related to pseudouridine synthases is required for chloroplast group II intron trans-splicing in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Perron, K; Goldschmidt-Clermont, M; Rochaix, J D

    1999-11-15

    In Chlamydomonas reinhardtii, the psaA mRNA is assembled by a process involving two steps of trans-splicing that remove two group II introns and give rise to the mature mRNA. The products of at least 14 nuclear genes and one chloroplast gene (tscA) are necessary for this process. We have cloned Maa2, one of the nuclear genes involved in trans-splicing of the second intron. Maa2 encodes a protein with similarity to conserved domains of pseudouridine synthases, but mutagenesis of putative catalytic residues showed that this activity may not be required for trans-splicing of psaA RNA. Although it is not clear whether the pseudouridine synthase activity has been maintained in Maa2, it is possible that this enzyme was recruited during evolution as an RNA chaperone for folding or stabilizing the psaA intron. The Maa2 protein appears to be associated through ionic interactions with a low density membrane system in the chloroplast that also contains RNA-binding proteins involved in translation.

  5. Relationships between H{sub 2} photoproduction and different electron transport pathways in sulfur-deprived Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Antal, Taras K.; Volgusheva, Alena A.; Kukarskih, Galina P.; Krendeleva, Tatyana E.; Rubin, Andrej B. [Faculty of Biology, Moscow State University, Vorobyevi Gory 119992, Moscow (Russian Federation)

    2009-11-15

    In this study the relationships between photosystem (PS) II dependent and independent pathways of H{sub 2} photoproduction, cyclic electron transport around PS I, chloro- and mitorespiration, and transmembrane {delta}pH were examined by inhibitor analysis in S deprived Chlamydomonas reinhardtii. The rate of non-photochemical reduction of plastoquinones in photosynthetic membranes was significantly diminished under starvation which may explain the minor contribution of the PS II independent pathway of H{sub 2} photoproduction in starved cells. The suppressive effect of the herbicide 3-(3,4-dichlorophenyl)-1,1-dimethylurea on the long-term H{sub 2} photoproduction was shown to be entirely attributed to the inhibition of electron transport in PS II, whereas non-specific interactions did not take place. Ferredoxin-quinone reductase - dependent cyclic electron transport around PS I slowed down H{sub 2} photoproduction more than two fold. This result was related to the competition between ferredoxin-quinone reductase and hydrogenase for the reduced ferredoxin and to the decrease in transmembrane {delta}pH induced by the cyclic electron flow. The {delta}pH gradient was shown to down regulate the PS II independent pathway of H{sub 2} photoproduction in starved cells. (author)

  6. Origin of pronounced differences in 77 K fluorescence of the green alga Chlamydomonas reinhardtii in state 1 and 2.

    Science.gov (United States)

    Ünlü, Caner; Polukhina, Iryna; van Amerongen, Herbert

    2016-04-01

    In response to changes in the reduction state of the plastoquinone pool in its thylakoid membrane, the green alga Chlamydomonas reinhardtti is performing state transitions: remodelling of its thylakoid membrane leads to a redistribution of excitations over photosystems I and II (PSI and PSII). These transitions are accompanied by marked changes in the 77 K fluorescence spectrum, which form the accepted signature of state transitions. The changes are generally thought to reflect a redistribution of light-harvesting complexes (LHCs) over PSII (fluorescing below 700 nm) and PSI (fluorescing above 700 nm). Here we studied the picosecond fluorescence properties of C. reinhardtti over a broad range of wavelengths with very low excitation intensities (0.2 nJ per laser pulse). Cells were directly used for time-resolved fluorescence measurements at 77 K without further treatment, such as medium exchange with glycerol. It is observed that upon going from state 1 (relatively more fluorescence below 700 nm) to state 2 (relatively more fluorescence above 700 nm), a large part of the fluorescence of LHC/PSII becomes substantially quenched in concurrence with LHC detachment from PSII, whereas the absolute amount of PSI fluorescence hardly changes. These results are in agreement with the recent proposal that the amount of LHC moving from PSII to PSI upon going from state 1 to state 2 is rather limited (Unlu et al. Proc Natl Acad Sci USA 111 (9):3460-3465, 2014).

  7. Complete elimination of maternal mitochondrial DNA during meiosis resulting in the paternal inheritance of the mitochondrial genome in Chlamydomonas species.

    Science.gov (United States)

    Aoyama, H; Hagiwara, Y; Misumi, O; Kuroiwa, T; Nakamura, S

    2006-09-01

    The non-Mendelian inheritance of organellar DNA is common in most plants and animals. In the isogamous green alga Chlamydomonas species, progeny inherit chloroplast genes from the maternal parent, as paternal chloroplast genes are selectively eliminated in young zygotes. Mitochondrial genes are inherited from the paternal parent. Analogically, maternal mitochondrial DNA (mtDNA) is thought to be selectively eliminated. Nevertheless, it is unclear when this selective elimination occurs. Here, we examined the behaviors of maternal and paternal mtDNAs by various methods during the period between the beginning of zygote formation and zoospore formation. First, we observed the behavior of the organelle nucleoids of living cells by specifically staining DNA with the fluorochrome SYBR Green I and staining mitochondria with 3,3'-dihexyloxacarbocyanine iodide. We also examined the fate of mtDNA of male and female parental origin by real-time PCR, nested PCR with single zygotes, and fluorescence in situ hybridization analysis. The mtDNA of maternal origin was completely eliminated before the first cell nuclear division, probably just before mtDNA synthesis, during meiosis. Therefore, the progeny inherit the remaining paternal mtDNA. We suggest that the complete elimination of maternal mtDNA during meiosis is the primary cause of paternal mitochondrial inheritance.

  8. Application of proton exchange membrane fuel cells for the monitoring and direct usage of biohydrogen produced by Chlamydomonas reinhardtii

    Science.gov (United States)

    Oncel, S.; Vardar-Sukan, F.

    Photo-biologically produced hydrogen by Chlamydomonas reinhardtii is integrated with a proton exchange (PEM) fuel cell for online electricity generation. To investigate the fuel cell efficiency, the effect of hydrogen production on the open circuit fuel cell voltage is monitored during 27 days of batch culture. Values of volumetric hydrogen production, monitored by the help of the calibrated water columns, are related with the open circuit voltage changes of the fuel cell. From the analysis of this relation a dead end configuration is selected to use the fuel cell in its best potential. After the open circuit experiments external loads are tested for their effects on the fuel cell voltage and current generation. According to the results two external loads are selected for the direct usage of the fuel cell incorporating with the photobioreactors (PBR). Experiments with the PEM fuel cell generate a current density of 1.81 mA cm -2 for about 50 h with 10 Ω load and 0.23 mA cm -2 for about 80 h with 100 Ω load.

  9. Application of proton exchange membrane fuel cells for the monitoring and direct usage of biohydrogen produced by Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Oncel, S.; Vardar-Sukan, F. [Department of Bioengineering, Faculty of Engineering, Ege University, 35100 Bornova, Izmir (Turkey)

    2011-01-01

    Photo-biologically produced hydrogen by Chlamydomonas reinhardtii is integrated with a proton exchange (PEM) fuel cell for online electricity generation. To investigate the fuel cell efficiency, the effect of hydrogen production on the open circuit fuel cell voltage is monitored during 27 days of batch culture. Values of volumetric hydrogen production, monitored by the help of the calibrated water columns, are related with the open circuit voltage changes of the fuel cell. From the analysis of this relation a dead end configuration is selected to use the fuel cell in its best potential. After the open circuit experiments external loads are tested for their effects on the fuel cell voltage and current generation. According to the results two external loads are selected for the direct usage of the fuel cell incorporating with the photobioreactors (PBR). Experiments with the PEM fuel cell generate a current density of 1.81 mA cm{sup -2} for about 50 h with 10 {omega} load and 0.23 mA cm{sup -2} for about 80 h with 100 {omega} load. (author)

  10. Comparison of tubular and panel type photobioreactors for biohydrogen production utilizing Chlamydomonas reinhardtii considering mixing time and light intensity.

    Science.gov (United States)

    Oncel, S; Kose, A

    2014-01-01

    Two different photobioreactor designs; tubular and panel, were investigated for the biohydrogen production utilizing a green microalgae Chlamydomonas reinhardtii strain CC124 following the two stage protocol. Mixing time and light intensity of the systems were adjusted to compare the productivity of both aerobic culture phase and the following anaerobic biohydrogen production phase. The results showed there was an effect on both phases related with the design. During the aerobic phase bigger illumination area serving more energy, tubular photobioreactor reached higher biomass productivity of 31.8±2.1 mg L(-1) h(-1) which was about 11% higher than the panel photobioreactor. On the other hand biohydrogen productivity in the panel photobioreactor reached a value of 1.3±0.05 mL L(-1) h(-1) based on the efficient removal of biohydrogen gas. According to the results it would be a good approach to utilize tubular design for aerobic phase and panel for biohydrogen production phase.

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

    Science.gov (United States)

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

    1998-06-01

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

  12. Assessment of a Novel Algal Strain Chlamydomonas debaryana NIREMACC03 for Mass Cultivation, Biofuels Production and Kinetic Studies.

    Science.gov (United States)

    Mishra, Sanjeev; Singh, Neetu; Sarma, Anil Kumar

    2015-08-01

    A novel microalgae strain Chlamydomonas debaryana (KJ210856) was isolated from a freshwater lake of Punjab, India, and cultivated considering climatic sustainability and inherent adaptability concern. C. debaryana was grown in a 30-L indoor photobioreactor to study the mass cultivation prospect and biofuel potential. Physicochemical characterization of biomass and the lipid was performed with effect to nitrogen stress. It showed a higher biomass yield (1.58 ± 0.02 g L(-1), dry weight) and twofold increase in lipid yield (552.78 ± 9 mg L(-1)) with 34.2 ± 0.19 % lipid content under nitrogen deficient condition. Strikingly, increase in triglycerides achieved with nitrogen depletion containing over 96 % of total fatty acids (C 14, C 16, and C 18). Proximate and ultimate analysis suggested the presence of relatively higher volatile matter and carbon-hydrogen ratio. Furthermore, lower moisture and ash content signified C. debaryana biomass has promising features towards biofuel applications. The pyrolytic behavior of the whole biomass was also studied using thermogravimetric analyzer (TGA) and kinetic parameters were estimated using different methods. Promising growth rate and lipid yield leading to feasible biofuel feed stock production in indoor photobioreactor along with autosediment potential of cells validates C. debaryana NIREMACC03, a potential strain for mass cultivation.

  13. Refactoring the Six-Gene Photosystem II Core in the Chloroplast of the Green Algae Chlamydomonas reinhardtii.

    Science.gov (United States)

    Gimpel, Javier A; Nour-Eldin, Hussam H; Scranton, Melissa A; Li, Daphne; Mayfield, Stephen P

    2016-07-15

    Oxygenic photosynthesis provides the energy to produce all food and most of the fuel on this planet. Photosystem II (PSII) is an essential and rate-limiting component of this process. Understanding and modifying PSII function could provide an opportunity for optimizing photosynthetic biomass production, particularly under specific environmental conditions. PSII is a complex multisubunit enzyme with strong interdependence among its components. In this work, we have deleted the six core genes of PSII in the eukaryotic alga Chlamydomonas reinhardtii and refactored them in a single DNA construct. Complementation of the knockout strain with the core PSII synthetic module from three different green algae resulted in reconstitution of photosynthetic activity to 85, 55, and 53% of that of the wild-type, demonstrating that the PSII core can be exchanged between algae species and retain function. The strains, synthetic cassettes, and refactoring strategy developed for this study demonstrate the potential of synthetic biology approaches for tailoring oxygenic photosynthesis and provide a powerful tool for unraveling PSII structure-function relationships.

  14. Expression of a clostridial [FeFe]-hydrogenase in Chlamydomonas reinhardtii prolongs photo-production of hydrogen from water splitting

    Energy Technology Data Exchange (ETDEWEB)

    Noone, Seth; Ratcliff, Kathleen; Davis, ReAnna; Subramanian, Venkataramanan; Meuser, Jonathan; Posewitz, Matthew C.; King, Paul W.; Ghirardi, Maria L.

    2017-03-01

    The high oxygen (O2) sensitivity of green algal [FeFe]-hydrogenases is a significant limitation for the sustained production of hydrogen gas (H2) from photosynthetic water splitting. To address this limitation we replaced the native [FeFe]-hydrogenases with a more O2-tolerant clostridial [FeFe]-hydrogenase CaI in Chlamydomonas reinhardtii strain D66..delta..HYD (hydA1-hydA2-) that contains insertionally inactivated [FeFe]-hydrogenases genes. Expression and translocation of CaI in D66..delta..HYD led to the recovery of H2 photoproduction at ~ 20% of the rates of the wild-type parent strain D66. We show for the first time that a bacterial [FeFe]-hydrogenase can be expressed, localized and matured to a catalytically active form that couples to photosynthetic electron transport in the green alga C. reinhardtii. The lower rates of O2 inactivation of CaI led to more sustained H2 photoproduction when cultures were challenged with O2 or kept under prolonged illumination at solar intensities. These results provide new insights into the requisites for attaining photobiological H2 production from water splitting using a more O2-tolerant hydrogenase.

  15. Intact anthracene inhibits photosynthesis in algal cells: a fluorescence induction study on Chlamydomonas reinhardtii cw92 strain.

    Science.gov (United States)

    Aksmann, Anna; Tukaj, Zbigniew

    2008-12-01

    Short-term (24h) experiments were performed to examine the effect of anthracene (ANT) on Chlamydomonas reinhardtii cw92 grown in a batch culture system aerated with 2.5% CO(2). At concentrations ranging from 0.7 to 5.6 microM, ANT inhibited the growth of population in a concentration-dependent manner and EC(50) calculated amounted to 1.6 microM. At concentrations from 0.7 to 4.2 microM ANT stimulated respiration and inhibited the intensity of photosynthesis but did not affect chlorophyll content in the cells. ANT influenced chlorophyll a fluorescence parameters, measured by OJIP test (O, J, I and P are the different steps of fluorescence induction curve). ANT diminished the performance index (PI), the yield of primary photochemistry (phi(Po)), the yield of electron transport (phi(Epsilonomicron), the efficiency of moving the electron beyond Qa(-) (Psi(0)) and the fraction of active oxygen evolving complexes (OEC). The fraction of active PS II reaction centres in the treated samples dramatically dropped. The most pronounced changes in ANT-treated cells were observed in the stimulation of energy dissipation parameter (DI(0)/RC). The only OJIP parameter that was not influenced by ANT was energy absorption by photosynthetic antennae (ABS). The results lead to a conclusion that the inhibition of photosynthesis may be a consequence of unspecific ANT-membrane interaction, resulting from hydrophobic character of this hydrocarbon.

  16. A dominant semi dwarf mutant in rice

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    @@ In the winter of 1997, a semi dwarf mutant was found in the F6 population of M9056/ R8018 xuan in Hainan Province. In the spring of 1998, the seeds were sown in Hefei, Anhui Province and the plant height of the population was measured at maturity.

  17. Nicotinamide ribosyl uptake mutants in Haemophilus influenzae.

    Science.gov (United States)

    Herbert, Mark; Sauer, Elizabeta; Smethurst, Graeme; Kraiss, Anita; Hilpert, Anna-Karina; Reidl, Joachim

    2003-09-01

    The gene for the nicotinamide riboside (NR) transporter (pnuC) was identified in Haemophilus influenzae. A pnuC mutant had only residual NR uptake and could survive in vitro with high concentrations of NR, but could not survive in vivo. PnuC may represent a target for the development of inhibitors for preventing H. influenzae disease.

  18. Colored HOMFLY polynomials can distinguish mutant knots

    CERN Document Server

    Nawata, Satoshi; Singh, Vivek Kumar

    2015-01-01

    We illustrate from the viewpoint of braiding operations on WZNW conformal blocks how colored HOMFLY polynomials with multiplicity structure can detect mutations. As an example, we explicitly evaluate the (2,1)-colored HOMFLY polynomials that distinguish a famous mutant pair, Kinoshita-Terasaka and Conway knot.

  19. Arabidopsis DNA polymerase lambda mutant is mildly sensitive to DNA double strand breaks but defective in integration ofa transgene.

    Directory of Open Access Journals (Sweden)

    Tomoyuki eFurukawa

    2015-05-01

    Full Text Available The DNA double-strand break (DSB is a critical type of damage, and can be induced by both endogenous sources (e.g. errors of oxidative metabolism, transposable elements, programmed meiotic breaks, or perturbation of the DNA replication fork and exogenous sources (e.g. ionizing radiation or radiomimetic chemicals. Although higher plants, like mammals, are thought to preferentially repair DSBs via nonhomologous end joining (NHEJ, much remains unclear about plant DSB repair pathways. Our reverse genetic approach suggests that DNA polymerase λ is involved in DSB repair in Arabidopsis. The Arabidopsis T-DNA insertion mutant (atpolλ-1 displayed sensitivity to both gamma-irradiation and treatment with radiomimetic reagents, but not to other DNA damaging treatments. The atpolλ-1 mutant showed a moderate sensitivity to DSBs, while Arabidopsis Ku70 and DNA ligase 4 mutants (atku70-3 and atlig4-2, both of which play critical roles in NHEJ, exhibited a hypersensitivity to these treatments. The atpolλ-1/atlig4-2 double mutant exhibited a higher sensitivity to DSBs than each single mutant, but the atku70/atpolλ-1 showed similar sensitivity to the atku70-3 mutant. We showed that transcription of the DNA ligase 1, DNA ligase 6, and Wee1 genes was quickly induced by BLM in several NHEJ deficient mutants in contrast to wild-type. Finally, the T-DNA transformation efficiency dropped in NHEJ deficient mutants and the lowest transformation efficiency was scored in the atpolλ-1/atlig4-2 double mutant. These results imply that AtPolλ is involved in both DSB repair and DNA damage response pathway.

  20. Evaluating Ambient Displays in the Wild

    DEFF Research Database (Denmark)

    Messeter, Jörn; Molenaar, Daryn

    A prominent issue for evaluating ambient displays has been the conflict between the relative intrusiveness of evaluation methods and the intention to keep the display at the periphery of the user’s attention. There is a general lack of research discussing the difficulties of evaluating ambient...... displays in the wild, and in particular social aspects of use has received little attention. This paper presents a case study of an ambient light display designed for a public setting. Based on results from a non-intrusive in situ evaluation, we argue that viewing ambient displays as features of a broader...... social setting may aid our understanding of issues regarding the evaluation of ambient displays in the wild....

  1. Laser-driven polyplanar optic display

    Energy Technology Data Exchange (ETDEWEB)

    Veligdan, J.T.; Biscardi, C.; Brewster, C.; DeSanto, L. [Brookhaven National Lab., Upton, NY (United States). Dept. of Advanced Technology; Beiser, L. [Leo Beiser Inc., Flushing, NY (United States)

    1998-01-01

    The Polyplanar Optical Display (POD) is a unique display screen which can be used with any projection source. This display screen is 2 inches thick and has a matte-black face which allows for high contrast images. The prototype being developed is a form, fit and functional replacement display for the B-52 aircraft which uses a monochrome ten-inch display. The new display uses a 200 milliwatt green solid-state laser (532 nm) as its optical source. In order to produce real-time video, the laser light is being modulated by a Digital Light Processing (DLP) chip manufactured by Texas Instruments, Inc. A variable astigmatic focusing system is used to produce a stigmatic image on the viewing face of the POD. In addition to the optical design, the authors discuss the DLP chip, the optomechanical design and viewing angle characteristics.

  2. Spectroradiometric characterization of autostereoscopic 3D displays

    Science.gov (United States)

    Rubiño, Manuel; Salas, Carlos; Pozo, Antonio M.; Castro, J. J.; Pérez-Ocón, Francisco

    2013-11-01

    Spectroradiometric measurements have been made for the experimental characterization of the RGB channels of autostereoscopic 3D displays, giving results for different measurement angles with respect to the normal direction of the plane of the display. In the study, 2 different models of autostereoscopic 3D displays of different sizes and resolutions were used, making measurements with a spectroradiometer (model PR-670 SpectraScan of PhotoResearch). From the measurements made, goniometric results were recorded for luminance contrast, and the fundamental hypotheses have been evaluated for the characterization of the displays: independence of the RGB channels and their constancy. The results show that the display with the lower angle variability in the contrast-ratio value and constancy of the chromaticity coordinates nevertheless presented the greatest additivity deviations with the measurement angle. For both displays, when the parameters evaluated were taken into account, lower angle variability consistently resulted in the 2D mode than in the 3D mode.

  3. Conceptual Design of Industrial Process Displays

    DEFF Research Database (Denmark)

    Pedersen, C.R.; Lind, Morten

    1999-01-01

    by a simple example from a plant with batch processes. Later the method is applied to develop a supervisory display for a condenser system in a nuclear power plant. The differences between the continuous plant domain of power production and the batch processes from the example are analysed and broad...... categories of display types are proposed. The problems involved in specification and invention of a supervisory display are analysed and conclusions from these problems are made. It is concluded that the design method proposed provides a framework for the progress of the display design and is useful in pin......-pointing the actual problems. The method was useful in reducing the number of existing displays that could fulfil the requirements of the supervision task. The method provided at the same time a framework for dealing with the problems involved in inventing new displays based on structured analysis. However...

  4. Three-dimensional Imaging, Visualization, and Display

    CERN Document Server

    Javidi, Bahram; Son, Jung-Young

    2009-01-01

    Three-Dimensional Imaging, Visualization, and Display describes recent developments, as well as the prospects and challenges facing 3D imaging, visualization, and display systems and devices. With the rapid advances in electronics, hardware, and software, 3D imaging techniques can now be implemented with commercially available components and can be used for many applications. This volume discusses the state-of-the-art in 3D display and visualization technologies, including binocular, multi-view, holographic, and image reproduction and capture techniques. It also covers 3D optical systems, 3D display instruments, 3D imaging applications, and details several attractive methods for producing 3D moving pictures. This book integrates the background material with new advances and applications in the field, and the available online supplement will include full color videos of 3D display systems. Three-Dimensional Imaging, Visualization, and Display is suitable for electrical engineers, computer scientists, optical e...

  5. Deficiency of antinociception and excessive grooming induced by acute immobilization stress in Per1 mutant mice.

    Science.gov (United States)

    Zhang, Jing; Wu, Zhouqiao; Zhou, Linglin; Li, Huili; Teng, Huajing; Dai, Wei; Wang, Yongqing; Sun, Zhong Sheng

    2011-01-14

    Acute stressors induce changes in numerous behavioral parameters through activation of the hypothalamic-pituitary-adrenal (HPA) axis. Several important hormones in paraventricular nucleus of the hypothalamus (PVN) play the roles in these stress-induced reactions. Corticotropin-releasing hormone (CRH), arginine-vasopressin (AVP) and corticosterone are considered as molecular markers for stress-induced grooming behavior. Oxytocin in PVN is an essential modulator for stress-induced antinociception. The clock gene, Per1, has been identified as an effecter response to the acute stresses, but its function in neuroendocrine stress systems remains unclear. In the present study we observed the alterations in grooming and nociceptive behaviors induced by acute immobilization stress in Per1 mutant mice and other genotypes (wild types and Per2 mutant). The results displayed that stress elicited a more robust effect on grooming behavior in Per1 mutant mice than in other genotypes. Subsequently, the obvious stress-induced antinociception was observed in the wild-type and Per2 mutant mice, however, in Per1 mutant, this antinociceptive effects were partially-reversed (mechanical sensitivity), or over-reversed to hyperalgesia (thermal sensitivity). The real-time qPCR results showed that in PVN, there were stress-induced up-regulations of Crh, Avp and c-fos in all of genotypes; moreover, the expression change of Crh in Per1 mutant mice was much larger than in others. Another hormonal gene, Oxt, was up-regulated induced by stress in wild-type and Per2 mutant but not in Per1 mutant. In addition, the stress significantly elevated the serum corticosterone levels without genotype-dependent differences, and accordingly the glucocorticoid receptor gene, Nr3c1, expressed with a similar pattern in PVN of all strains. Taken together, the present study indicated that in acute stress treated Per1 mutant mice, there are abnormal hormonal responses in PVN, correlating with the aberrant

  6. A human vitamin D receptor mutant activated by cholecalciferol.

    Science.gov (United States)

    Ousley, Amanda M; Castillo, Hilda S; Duraj-Thatte, Anna; Doyle, Donald F; Azizi, Bahareh

    2011-07-01

    The human vitamin D receptor (hVDR) is a member of the nuclear receptor superfamily, involved in calcium and phosphate homeostasis; hence implicated in a number of diseases, such as Rickets and Osteoporosis. This receptor binds 1α,25-dihydroxyvitamin D(3) (also referred to as 1,25(OH)(2)D(3)) and other known ligands, such as lithocholic acid. Specific interactions between the receptor and ligand are crucial for the function and activation of this receptor, as implied by the single point mutation, H305Q, causing symptoms of Type II Rickets. In this work, further understanding of the significant and essential interactions between the ligand and the receptor was deciphered, through a combination of rational and random mutagenesis. A hVDR mutant, H305F, was engineered with increased sensitivity towards lithocholic acid, with an EC(50) value of 10 μM and 40±14 fold activation in mammalian cell assays, while maintaining wild-type activity with 1,25(OH)(2)D(3). Furthermore, via random mutagenesis, a hVDR mutant, H305F/H397Y, was discovered to bind a novel small molecule, cholecalciferol, a precursor in the 1α,25-dihydroxyvitamin D(3) biosynthetic pathway, which does not activate wild-type hVDR. This variant, H305F/H397Y, binds and activates in response to cholecalciferol concentrations as low as 100 nM, with an EC(50) value of 300 nM and 70±11 fold activation in mammalian cell assays. In silico docking analysis of the variant displays a dramatic conformational shift of cholecalciferol in the ligand binding pocket in comparison to the docked analysis of cholecalciferol with wild-type hVDR. This shift is hypothesized to be due to the introduction of two bulkier residues, suggesting that the addition of these bulkier residues introduces molecular interactions between the ligand and receptor, leading to activation with cholecalciferol.

  7. GAMPMS: Genetic algorithm managed peptide mutant screening.

    Science.gov (United States)

    Long, Thomas; McDougal, Owen M; Andersen, Tim

    2015-06-30

    The prominence of endogenous peptide ligands targeted to receptors makes peptides with the desired binding activity good molecular scaffolds for drug development. Minor modifications to a peptide's primary sequence can significantly alter its binding properties with a receptor, and screening collections of peptide mutants is a useful technique for probing the receptor-ligand binding domain. Unfortunately, the combinatorial growth of such collections can limit the number of mutations which can be explored using structure-based molecular docking techniques. Genetic algorithm managed peptide mutant screening (GAMPMS) uses a genetic algorithm to conduct a heuristic search of the peptide's mutation space for peptides with optimal binding activity, significantly reducing the computational requirements of the virtual screening. The GAMPMS procedure was implemented and used to explore the binding domain of the nicotinic acetylcholine receptor (nAChR) α3β2-isoform with a library of 64,000 α-conotoxin (α-CTx) MII peptide mutants. To assess GAMPMS's performance, it was compared with a virtual screening procedure that used AutoDock to predict the binding affinity of each of the α-CTx MII peptide mutants with the α3β2-nAChR. The GAMPMS implementation performed AutoDock simulations for as few as 1140 of the 64,000 α-CTx MII peptide mutants and could consistently identify a set of 10 peptides with an aggregated binding energy that was at least 98% of the aggregated binding energy of the 10 top peptides from the exhaustive AutoDock screening.

  8. Matrix-addressable electrochromic display cell

    Science.gov (United States)

    Beni, G.; Schiavone, L. M.

    1981-04-01

    We report an electrochromic display cell with intrinsic matrix addressability. The cell, based on a sputtered iridium oxide film (SIROF) and a tantalum-oxide hysteretic counterelectrode, has electrochromic parameters (i.e., response times, operating voltages, and contrast) similar to those of other SIROF display devices, but in addition, has short-circuit memory and voltage threshold. Memory and threshold are sufficiently large to allow, in principle, multiplexing of electrochromic display panels of large-screen TV pixel size.

  9. Laser Based 3D Volumetric Display System

    Science.gov (United States)

    1993-03-01

    Literature, Costa Mesa, CA July 1983. 3. "A Real Time Autostereoscopic Multiplanar 3D Display System", Rodney Don Williams, Felix Garcia, Jr., Texas...8217 .- NUMBERS LASER BASED 3D VOLUMETRIC DISPLAY SYSTEM PR: CD13 0. AUTHOR(S) PE: N/AWIU: DN303151 P. Soltan, J. Trias, W. Robinson, W. Dahlke 7...laser generated 3D volumetric images on a rotating double helix, (where the 3D displays are computer controlled for group viewing with the naked eye

  10. Characterization of zebrafish mutants with defects in bone calcification during development.

    Science.gov (United States)

    Xi, Yang; Chen, Dongyan; Sun, Lei; Li, Yuhao; Li, Lei

    2013-10-11

    Using the fluorescent dyes calcein and alcian blue, we stained the F3 generation of chemically (ENU) mutagenized zebrafish embryos and larvae, and screened for mutants with defects in bone development. We identified a mutant line, bone calcification slow (bcs), which showed delayed axial vertebra calcification during development. Before 4-5 days post-fertilization (dpf), the bcs embryos did not display obvious abnormalities in bone development (i.e., normal number, size and shape of cartilage and vertebrae). At 5-6 dpf, when vertebrae calcification starts, bcs embryos began to show defects. At 7 dpf, for example, in most of the bcs embryos examined, calcein staining revealed no signals of vertebrae mineralization, whereas during the same developmental stages, 2-14 mineralized vertebrae were observed in wild-type animals. Decreases in the number of calcified vertebrae were also observed in bcs mutants when examined at 9 and 11 dpf, respectively. Interestingly, by 13 dpf the defects in bcs mutants were no longer evident. There were no significant differences in the number of calcified vertebrae between wild-type and mutant animals. We examined the expression of bone development marker genes (e.g., Sox9b, Bmp2b, and Cyp26b1, which play important roles in bone formation and calcification). In mutant fish, we observed slight increases in Sox9b expression, no alterations in Bmp2b expression, but significant increases in Cyp26b1 expression. Together, the data suggest that bcs delays axial skeletal calcification, but does not affect bone formation and maturation.

  11. Differential reactivity of mouse monoclonal anti-HBs antibodies with recombinant mutant HBs antigens

    Institute of Scientific and Technical Information of China (English)

    Azam Roohi; Yaghoub Yazdani; Jalal Khoshnoodi; Seyed Mohammad Jazayeri; William F Carman; Mahmood Chamankhah; Manley Rashedan; Fazel Shokri

    2006-01-01

    AIM: To investigate the reactivity of a panel of 8 mouse anti-hepatitis B surface antigen (HBsAg) monoclonal antibodies (mAbs) using a collection of 9 recombinant HBsAg mutants with a variety of amino acid substitutions mostly located within the "a" region.METHODS: The entire HBs genes previously cloned into a mammalian expression vector were transiently transfected into COS7 cells. Two standard unmutated sequences of the ayw and adw subtypes served as controls. Secreted mutant proteins were collected and measured by three commercial diagnostic immunoassays to assess transfection efficiency. Reactivity of anti-HBs mAbs with mutated HBsAgs was determined by sandwich enzyme-linked immunosorbent assay (ELISA).RESULTS: Reactivity of anti-HBs mAbs with mutated HBsAgs revealed different patterns. While three mutants reacted strongly with all mAbs, two mutants reacted weakly with only two mAbs and the remaining proteins displayed variable degrees of reactivity towards different mAbs. Accordingly, four groups of mAbs with different but overlapping reactivity patterns could be envisaged. One group consisting of two mAbs (37C5-S7 and 35C6-S11) was found to recognize stable linear epitopes conserved in all mutants. Mutations outside the "a"determinant at positions 120 (P→S), 123(T→N) and 161(M→T) were found to affect reactivity of these mAbs.CONCLUSION: Our findings could have important implications for biophysical studies, vaccination strategies and immunotherapy of hepatitis B virus (HBV) mutants.

  12. Volumetric Three-Dimensional Display Systems

    Science.gov (United States)

    Blundell, Barry G.; Schwarz, Adam J.

    2000-03-01

    A comprehensive study of approaches to three-dimensional visualization by volumetric display systems This groundbreaking volume provides an unbiased and in-depth discussion on a broad range of volumetric three-dimensional display systems. It examines the history, development, design, and future of these displays, and considers their potential for application to key areas in which visualization plays a major role. Drawing substantially on material that was previously unpublished or available only in patent form, the authors establish the first comprehensive technical and mathematical formalization of the field, and examine a number of different volumetric architectures. System level design strategies are presented, from which proposals for the next generation of high-definition predictable volumetric systems are developed. To ensure that researchers will benefit from work already completed, they provide: * Descriptions of several recent volumetric display systems prepared from material supplied by the teams that created them * An abstract volumetric display system design paradigm * An historical summary of 90 years of development in volumetric display system technology * An assessment of the strengths and weaknesses of many of the systems proposed to date * A unified presentation of the underlying principles of volumetric display systems * A comprehensive bibliography Beautifully supplemented with 17 color plates that illustrate volumetric images and prototype displays, Volumetric Three-Dimensional Display Systems is an indispensable resource for professionals in imaging systems development, scientific visualization, medical imaging, computer graphics, aerospace, military planning, and CAD/CAE.

  13. New ultraportable display technology and applications

    Science.gov (United States)

    Alvelda, Phillip; Lewis, Nancy D.

    1998-08-01

    MicroDisplay devices are based on a combination of technologies rooted in the extreme integration capability of conventionally fabricated CMOS active-matrix liquid crystal display substrates. Customized diffraction grating and optical distortion correction technology for lens-system compensation allow the elimination of many lenses and systems-level components. The MicroDisplay Corporation's miniature integrated information display technology is rapidly leading to many new defense and commercial applications. There are no moving parts in MicroDisplay substrates, and the fabrication of the color generating gratings, already part of the CMOS circuit fabrication process, is effectively cost and manufacturing process-free. The entire suite of the MicroDisplay Corporation's technologies was devised to create a line of application- specific integrated circuit single-chip display systems with integrated computing, memory, and communication circuitry. Next-generation portable communication, computer, and consumer electronic devices such as truly portable monitor and TV projectors, eyeglass and head mounted displays, pagers and Personal Communication Services hand-sets, and wristwatch-mounted video phones are among the may target commercial markets for MicroDisplay technology. Defense applications range from Maintenance and Repair support, to night-vision systems, to portable projectors for mobile command and control centers.

  14. Microencapsulated Electrophoretic Films for Electronic Paper Displays

    Science.gov (United States)

    Amundson, Karl

    2003-03-01

    Despite the dominance of liquid crystal displays, they do not perform some functions very well. While backlit liquid crystal displays can offer excellent color performance, they wash out in bright lighting and suffer from high power consumption. Reflective liquid crystal displays have limited brightness, making these devices challenging to read for long periods of time. Flexible liquid crystal displays are difficult to manufacture and keep stable. All of these attributes (long battery lifetime, bright reflective appearance, compatibility with flexible substrates) are traits that would be found in an ideal electronic paper display - an updateable substitute for paper that could be employed in electronic books, newspapers, and other applications. I will discuss technologies that are being developed for electronic-paper-like displays, and especially on particle-based technologies. A microencapsulated electrophoretic display technology is being developed at the E Ink corporation. This display film offers offer high brightness and an ink-on-paper appearance, compatibility with flexible substrates, and image stability that can lead to very low power consumption. I will present some of the physical and chemical challenges associated with making display films with high performance.

  15. Pengaruh Display Produk pada Keputusan Pembelian Konsumen

    Directory of Open Access Journals (Sweden)

    Ina Melati

    2012-10-01

    Full Text Available Most of ritel outlet recently using product display as a one of their best marketing strategy, the reason is quiet easy to be understood, since consumers are too easy to be teased by those kind of beautiful product display that is being displayed by the retail outlet. The good retail outlets are trying their best to design and make the very good product display, so they can attract more consumers and make them not thinking twice to visit their store and purchase lots of thing. Clearly seeing that an attractive product design is able to influence a consumer to make a buying decision.

  16. Framework for effective use of multiple displays

    Science.gov (United States)

    Liu, Qiong; Kimber, Don; Zhao, Frank; Huang, Jeffrey

    2005-10-01

    Meeting environments, such as conference rooms, executive briefing centers, and exhibition spaces, are now commonly equipped with multiple displays, and will become increasingly display-rich in the future. Existing authoring/presentation tools such as PowerPoint, however, provide little support for effective utilization of multiple displays. Even using advanced multi-display enabled multimedia presentation tools, the task of assigning material to displays is tedious and distracts presenters from focusing on content. This paper describes a framework for automatically assigning presentation material to displays, based on a model of the quality of views of audience members. The framework is based on a model of visual fidelity which takes into account presentation content, audience members' locations, the limited resolution of human eyes, and display location, orientation, size, resolution, and frame rate. The model can be used to determine presentation material placement based on average or worst case audience member view quality, and to warn about material that would be illegible. By integrating this framework with a previous system for multi-display presentation [PreAuthor, others], we created a tool that accepts PowerPoint and/or other media input files, and automatically generates a layout of material onto displays for each state of the presentation. The tool also provides an interface allowing the presenter to modify the automatically generated layout before or during the actual presentation. This paper discusses the framework, possible application scenarios, examples of the system behavior, and our experience with system use.

  17. Refreshable Braille Displays Using EAP Actuators

    Science.gov (United States)

    Bar-Cohen, Yoseph

    2010-01-01

    Refreshable Braille can help visually impaired persons benefit from the growing advances in computer technology. The development of such displays in a full screen form is a great challenge due to the need to pack many actuators in small area without interferences. In recent years, various displays using actuators such as piezoelectric stacks have become available in commercial form but most of them are limited to one line Braille code. Researchers in the field of electroactive polymers (EAP) investigated methods of using these materials to form full screen displays. This manuscript reviews the state of the art of producing refreshable Braille displays using EAP-based actuators..

  18. Refreshable Braille displays using EAP actuators

    Science.gov (United States)

    Bar-Cohen, Yoseph

    2010-04-01

    Refreshable Braille can help visually impaired persons benefit from the growing advances in computer technology. The development of such displays in a full screen form is a great challenge due to the need to pack many actuators in small area without interferences. In recent years, various displays using actuators such as piezoelectric stacks have become available in commercial form but most of them are limited to one line Braille code. Researchers in the field of electroactive polymers (EAP) investigated methods of using these materials to form full screen displays. This manuscript reviews the state of the art of producing refreshable Braille displays using EAP-based actuators.

  19. Characterization of a mutant strain of a filamentous fungus Cladosporium phlei for the mass production of the secondary metabolite phleichrome.

    Science.gov (United States)

    Yi, Min-Hee; Kim, Jung-Ae; Kim, Jung-Mi; Park, Jin-Ah; Kim, Beom-Tae; Park, Seung-Moon; Yang, Moon-Sik; Hwang, Ki-Jun; Kim, Dae-Hyuk

    2011-08-01

    UV-mutagenesis was performed to obtain mutant strains that demonstrate altered production of phleichrome, a secondary metabolite of Cladosporium phlei. Among fifty mutants selected, based on the increased area and intensity of the purple pigment surrounding the colonies, the strain M0035 showed the highest production of phleichrome, more than seven fold over wild type. Plate cultures of the M0035 strain resulted in a total of 592 mg phleichrome consisting of 146 mg and 446 mg from the mycelia and agar media, respectively. The M0035 strain displayed a growth rate and a mycelial mass comparable to the parental strain but had significantly reduced asexual sporulation.

  20. Molecular cloning and bioinformatics analysis of the S-adenosyl homocysteine hydrolase(SAHH)gene in the Antarctic ice alga Chlamydomonas sp.ICE-L(Chlamydomonadales, Chlamydomonas)%南极冰藻Chlamydomonas sp.ICE-L S-腺苷同型半胱氨酸水解酶基因的克隆及其生物信息学分析

    Institute of Scientific and Technical Information of China (English)

    王金慧; 丁燏; 简纪常; 吴灶和

    2011-01-01

    S-Adenosyl-L-methionine(SAM)is the most common methyl donor in a multitude of cellular methyla-tion reactions. Numerous methyltransferases transfer the methyl group from SAM to their respective biological acceptors, forming S-adenosyl-L-homocysteine(SAH). SAH is hydrolyzed to adenosine and L-homocysteine by S-adenosyl-L-homocysteine hydrolase(SAHH, EC 3.3.1.1). SAHH is an essential enzyme in all living cells, and has been explored as a potential drug target for many bacteria and parasites. The inhibition of SAHH activity can result in the accumulation of SAH and reduce the S-adenosylmethionine(SAM):SAH ratio in cells. In that case, SAHH can act as a potent feedback inhibitor, blocking the SAM-dependent methylation that is required for the metabolism of a wide variety of biological compounds such as nucleic acids, proteins, phospholipids, and other small molecules. SAHH is found in animals, plants, fungi, and other microorganisms. Antarctic ice microalgae with special characteristics to withstand the extreme environment characterized by low temperatures and ice, high levels of dissolved oxygen, and strong seasonal changes in light intensity have been investigated in recent years. To further understand the intrinsic mechanisms by which the Antarctic sea ice alga Chlamydomonas sp. ICE-L responds to ecological and environmental factors, we cloned and analyzed its SAHH. A cDNA encoding S-adenosyl homocysteine hydrolase(designated as ICE-LSAHH)was cloned from this alga by RT-PCR and RACE-PCR methods. The ICE-LSAHH full-length cDNA sequence was 1844 bp, including a 5'-terminal untranslated region(UTR)of 36 bp, a 3'-terminal UTR of 344 bp with a poly(A)tail, and an open reading frame(ORF)of 1461 bp encoding a polypeptide of 487 amino acids. The predicted molecular weight(MW)of ICE-LSAHH was 53 kD with an estimated pi of 5.16. Using SignalP 3.0 and TMHMM Server v. 2.0 software, it was predicted that the ICE-LSAHH protein did not contain a signal peptide or a transmembranous

  1. Yeast surface display for directed evolution of protein expression, affinity, and stability.

    Science.gov (United States)

    Boder, E T; Wittrup, K D

    2000-01-01

    The described protocols enable thorough screening of polypeptide libraries with high confidence in the isolation of improved clones. It should be emphasized that the protocols have been fashioned for thoroughness, rather than speed. With library plasmid DNA in hand, the time to plated candidate yeast display mutants is typically 2-3 weeks. Each of the experimental approaches required for this method is fairly standard: yeast culture, immunofluorescent labeling, flow cytometry. Protocols that are more rapid could conceivably be developed by using solid substrate separations with magnetic beads, for instance. However, loss of the two-color normalization possible with flow cytometry would remove the quantitative advantage of the method. Yeast display complements existing polypeptide library methods and opens the possibility of examining extracellular eukaryotic proteins, an important class of proteins not generally amenable to yeast two-hybrid or phage display methodologies.

  2. Influence of temperature on glutathione level and glutathione-related enzyme activities of Antarctic ice microalgae Chlamydomonas sp.ICE-L

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    GSH system plays a role in the control of the redox balance state, anti-oxidation and protecting life from injury of ROS (reactive oxygen species).In present paper, the possible GSH system of Chlamydomonas sp.ICE-L has been investigated by evaluating GSH and GSH-related enzymatic responses at different temperatures using speetrophotometer methods.The results showed that the GSH system is correlated positively to low temperature, and other factors but GR are correlated negatively to high temperature.So GSH and GSH-related enzymes play an important role in the adaptation of Antarctic ice microulgae to low temperature.

  3. Study of metabolic pathways for hydrogen production in chlamydomonas reinhardtii and transposition on a torus photo bioreactor; Etude des voies metaboliques de production d'hydrogene chez la microalgue Chlamydomonas reinhardtii et transposition en photobioreacteur

    Energy Technology Data Exchange (ETDEWEB)

    Fouchard, S

    2006-04-15

    Considering the recent increase in energy consumption. aide associated environmental risks, new trails are followed today to develop the use of clean and renewable alternative energies. In this context hydrogen seems to be a serious solution and this study, based on micro-algae photosynthetic capacities exploitation, will allow to devise a process for hydrogen production from only water and solar energy without greenhouse gas release. The sulphur deprivation protocol on TAP medium, known to lead to hydrogen production in Chlamydomonas reinhardtii species was particularly studied. At the metabolic level, two important phenomena are induced under these conditions: an over-accumulation of the intracellular starch reserves and a simultaneous alteration of the PsII activity which leads to anoxia and Fe-hydrogenase induction, an enzyme with a strong specific activity responsible for the hydrogen production. The contribution of the two electron transfer pathways implied in the hydrogen production process (PsII-dependent and PSII-independent) as well as the importance of the previously accumulated starch were highlighted here. We also investigated the potential for designing autotrophic protocols for hydrogen photoproduction. Various protocols, considered to be relevant, were then transposed on a torus photo-bioreactor, specifically developed in this study and which allows the control of culture parameters as well as the precise measurement of gas release kinetics, in order to obtain first estimates of productivity of the system. Integration of the physical; aspects of the pilot and biological aspects of the process in a model, finally opens new prospects for subject development, in particular for a reasoned optimization of hydrogen production via this double physiology/process approach. (author)

  4. Induced protein polymorphisms and nutritional quality of gamma irradiation mutants of sorghum

    Energy Technology Data Exchange (ETDEWEB)

    Mehlo, Luke, E-mail: LMehlo@csir.co.za [CSIR Biosciences, Meiring Naude Road, P.O. Box 395, Pretoria 0001 (South Africa); Mbambo, Zodwa [CSIR Biosciences, Meiring Naude Road, P.O. Box 395, Pretoria 0001 (South Africa); Microbiology Discipline, School of Life Sciences, University of KwaZulu-Natal (Westville Campus), Private Bag X54001, Durban 4000 (South Africa); Bado, Souleymane [Plant Breeding and Genetics Laboratory – Joint FAO/IAEA Agriculture and Biotechnology Laboratory, International Atomic Energy Agency Laboratories, A-2444 Seibersdorf (Austria); Lin, Johnson [Microbiology Discipline, School of Life Sciences, University of KwaZulu-Natal (Westville Campus), Private Bag X54001, Durban 4000 (South Africa); Moagi, Sydwell M.; Buthelezi, Sindisiwe; Stoychev, Stoyan; Chikwamba, Rachel [CSIR Biosciences, Meiring Naude Road, P.O. Box 395, Pretoria 0001 (South Africa)

    2013-09-15

    Highlights: • We analyse kafirin protein polymorphisms induced by gamma irradiation in sorghum. • One mutant with suppressed kafirins in the endosperm accumulated them in the germ. • Kafirin polymorphisms were associated with high levels of free amino acids. • Nutritional value of sorghum can be improved significantly by induced mutations. - Abstract: Physical and biochemical analysis of protein polymorphisms in seed storage proteins of a mutant population of sorghum revealed a mutant with redirected accumulation of kafirin proteins in the germ. The change in storage proteins was accompanied by an unusually high level accumulation of free lysine and other essential amino acids in the endosperm. This mutant further displayed a significant suppression in the synthesis and accumulation of the 27 kDa γ-, 24 kDa α-A1 and the 22 kDa α-A2 kafirins in the endosperm. The suppression of kafirins was counteracted by an upsurge in the synthesis and accumulation of albumins, globulins and other proteins. The data collectively suggest that sorghum has huge genetic potential for nutritional biofortification and that induced mutations can be used as an effective tool in achieving premium nutrition in staple cereals.

  5. Durations and frequencies of free locomotion in wild type and GABAergic mutants of Caenorhabditis elegans.

    Science.gov (United States)

    Shingai, R

    2000-09-01

    We investigated how much time wild-type Caenorhabditis elegans (Bristol N2) nematodes and the GABA-deficient unc25 mutant and the vesicular GABA transporter-deficient unc47 mutant spent moving. The worms were allowed to move freely on the surface of agarose plates either with or without the food bacterium OP50. We identified forward movement, backward movement, resting and turns by watching images on video and computer displays. Forward movement lasted longer and rests were briefer without, than with, bacteria. Frequency distributions except for backward movement fitted a sum of two exponential functions. The duration of backward movement was not strongly influenced by exposure to bacteria, whereas the frequency of backward movements increased in their presence. The duration of forward movement of unc25 nematodes had no long component, thus differing from that of N2 and unc47 strain nematodes in treatments with and without bacteria. The durations of resting in these mutants were much longer than in the N2 strain, especially in the absence of bacteria. The turn frequency of unc47 nematodes had a higher short component than that of the wild type N2 and unc25 nematodes, in the absence of bacteria. A neural network model is discussed in conjunction with the features of mutants and current knowledge of GABAergic neural transmission.

  6. 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.

  7. Co-occurence of filamentation defects and impaired biofilms in Candida albicans protein kinase mutants.

    Science.gov (United States)

    Konstantinidou, Nina; Morrissey, John Patrick

    2015-12-01

    Pathogenicity of Candida albicans is linked with its developmental stages, notably the capacity switch from yeast-like to hyphal growth, and to form biofilms on surfaces. To better understand the cellular processes involved in C. albicans development, a collection of 63 C. albicans protein kinase mutants was screened for biofilm formation in a microtitre plate assay. Thirty-eight mutants displayed some degree of biofilm impairment, with 20 categorised as poor biofilm formers. All the poor biofilm formers were also defective in the switch from yeast to hyphae, establishing it as a primary defect. Five genes, VPS15, IME2, PKH3, PGA43 and CEX1, encode proteins not previously reported to influence hyphal development or biofilm formation. Network analysis established that individual components of some processes, most interestingly MAP kinase pathways, are not required for biofilm formation, most likely indicating functional redundancy. Mutants were also screened for their response to bacterial supernatants and it was found that Pseudomonas aeruginosa supernatants inhibited biofilm formation in all mutants, regardless of the presence of homoserine lactones (HSLs). In contrast, Candida morphology was only affected by supernatant containing HSLs. This confirms the distinct HSL-dependent inhibition of filamentation and the HSL-independent impairment of biofilm development by P. aeruginosa.

  8. Chloroplast dysfunction causes multiple defects in cell cycle progression in the Arabidopsis crumpled leaf mutant.

    Science.gov (United States)

    Hudik, Elodie; Yoshioka, Yasushi; Domenichini, Séverine; Bourge, Mickaël; Soubigout-Taconnat, Ludivine; Mazubert, Christelle; Yi, Dalong; Bujaldon, Sandrine; Hayashi, Hiroyuki; De Veylder, Lieven; Bergounioux, Catherine; Benhamed, Moussa; Raynaud, Cécile

    2014-09-01

    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.

  9. Quantitative Analysis of Mutant Subclones in Chronic Myeloid Leukemia: Comparison of Different Methodological Approaches.

    Science.gov (United States)

    Preuner, Sandra; Barna, Agnes; Frommlet, Florian; Czurda, Stefan; Konstantin, Byrgazov; Alikian, Mary; Machova Polakova, Katerina; Sacha, Tomasz; Richter, Johan; Lion, Thomas; Gabriel, Christian

    2016-04-29

    Identification and quantitative monitoring of mutant BCR-ABL1 subclones displaying resistance to tyrosine kinase inhibitors (TKIs) have become important tasks in patients with Ph-positive leukemias. Different technologies have been established for patient screening. Various next-generation sequencing (NGS) platforms facilitating sensitive detection and quantitative monitoring of mutations in the ABL1-kinase domain (KD) have been introduced recently, and are expected to become the preferred technology in the future. However, broad clinical implementation of NGS methods has been hampered by the limited accessibility at different centers and the current costs of analysis which may not be regarded as readily affordable for routine diagnostic monitoring. It is therefore of interest to determine whether NGS platforms can be adequately substituted by other methodological approaches. We have tested three different techniques including pyrosequencing, LD (ligation-dependent)-PCR and NGS in a series of peripheral blood specimens from chronic myeloid leukemia (CML) patients carrying single or multiple mutations in the BCR-ABL1 KD. The proliferation kinetics of mutant subclones in serial specimens obtained during the course of TKI-treatment revealed similar profiles via all technical approaches, but individual specimens showed statistically significant differences between NGS and the other methods tested. The observations indicate that different approaches to detection and quantification of mutant subclones may be applicable for the monitoring of clonal kinetics, but careful calibration of each method is required for accurate size assessment of mutant subclones at individual time points.

  10. Genetic Analysis and Gene Mapping of Light Brown Spotted Leaf Mutant in Rice

    Institute of Scientific and Technical Information of China (English)

    FENG Bao-hua; YANG Yang; SHI Yong-feng; LIN Lu; CHEN Jie; WEI Yan-lin; Hei LEUNG

    2013-01-01

    A light brown spotted-leaf mutant of rice was isolated from an ethane methyl sulfonate (EMS)induced IR64 mutant bank.The mutant,designated as Ibsl1 (light brown spotted-leaf 1),displayed light brown spot in the whole growth period from the first leaf to the flag leaf under natural summer field conditions.Agronomic traits including plant height,growth duration,number of filled grains per panicle,seed-setting rate and 1000-grain weight of the mutant were significantly affected.Genetic analysis showed that the mutation was controlled by a single recessive gene,tentatively named Ibsl1(t),which was mapped to the short arm of chromosome 6.By developing simple sequence repeat (SSR) markers,the gene was finally delimited to an interval of 130 kb between markers RM586 and RM588.The Ibsl1(t) gene is likely a novel rice spotted-leaf gene since no other similar genes have been identified near the chromosomal region.The genetic data and recombination populations provided will facilitate further fine-mapping and cloning of the gene.

  11. Mutants of Streptomyces cattleya defective in the synthesis of a factor required for thienamycin production.

    Science.gov (United States)

    Buchan, T; Roach, C; Ruby, C; Taylor, D; Preisig, C; Reeves, C

    1994-09-01

    Thienamycin non-producing mutants of Streptomydes cattleya were identified that displayed a cross-feeding relationship. A diffusible product from one of these mutants (RK-11) resulted in restoration of thienamycin production when fed to cultures of another mutant (RK-4). In vivo radiolabeling experiments were conducted to test whether the RK-11 mutant produced a late biosynthetic intermediate which contained a carbapenem ring and a cysteaminyl and/or a hydroxyethyl side chain. Both [35S]cystine and [methyl-3H]methionine were used to label the RK-11 product which was then fed to RK-4 cultures. None of the thienamycin subsequently produced by RK-4 converter cells was labeled, implying the lack of either side chain of the thienamycin molecule in the RK-11 product. Further stability studies suggested that the RK-11 product does not contain a carbapenem ring. Additional feeding experiments with RK-4 cells also ruled out the possibility that the RK-11 product is a co-factor necessary for thienamycin production. It is concluded that the RK-11 product may regulate expression of the thienamycin gene cluster.

  12. Amuvatinib has cytotoxic effects against NRAS-mutant melanoma but not BRAF-mutant melanoma.

    Science.gov (United States)

    Fedorenko, Inna V; Fang, Bin; Koomen, John M; Gibney, Geoffrey T; Smalley, Keiran S M

    2014-10-01

    Effective targeted therapy strategies are still lacking for the 15-20% of melanoma patients whose melanomas are driven by oncogenic NRAS. Here, we report on the NRAS-specific behavior of amuvatinib, a kinase inhibitor with activity against c-KIT, Axl, PDGFRα, and Rad51. An analysis of BRAF-mutant and NRAS-mutant melanoma cell lines showed the NRAS-mutant cohort to be enriched for targets of amuvatinib, including Axl, c-KIT, and the Axl ligand Gas6. Increasing concentrations of amuvatinib selectively inhibited the growth of NRAS-mutant, but not BRAF-mutant melanoma cell lines, an effect associated with induction of S-phase and G2/M-phase cell cycle arrest and induction of apoptosis. Mechanistically, amuvatinib was noted to either inhibit Axl, AKT, and MAPK signaling or Axl and AKT signaling and to induce a DNA damage response. In three-dimensional cell culture experiments, amuvatinib was cytotoxic against NRAS-mutant melanoma cell lines. Thus, we show for the first time that amuvatinib has proapoptotic activity against melanoma cell lines, with selectivity observed for those harboring oncogenic NRAS.

  13. Methods for Selecting Phage Display Antibody Libraries.

    Science.gov (United States)

    Jara-Acevedo, Ricardo; Diez, Paula; Gonzalez-Gonzalez, Maria; Degano, Rosa Maria; Ibarrola, Nieves; Gongora, Rafael; Orfao, Alberto; Fuentes, Manuel

    2016-01-01

    The selection process aims sequential enrichment of phage antibody display library in clones that recognize the target of interest or antigen as the library undergoes successive rounds of selection. In this review, selection methods most commonly used for phage display antibody libraries have been comprehensively described.

  14. Teacher Portfolios: Displaying the Art of Teaching

    Science.gov (United States)

    Reese, Susan

    2004-01-01

    A portfolio can convey a teacher's beliefs, knowledge and skills. An artist uses a portfolio to display artistic talent, and a teacher can use his or her portfolio to display teaching talent and teaching style. A teacher's portfolio may be used to obtain new employment, to document teaching accomplishments in order to receive a promotion or tenure…

  15. Display Developer for Firing Room Applications

    Science.gov (United States)

    Bowman, Elizabeth A.

    2013-01-01

    The firing room at Kennedy Space Center (KSC) is responsible for all NASA human spaceflight launch operations, therefore it is vital that all displays within the firing room be properly tested, up-to-date, and user-friendly during a launch. The Ground Main Propulsion System (GMPS) requires a number of remote displays for Vehicle Integration and Launch (VIL) Operations at KSC. My project is to develop remote displays for the GMPS using the Display Services and Framework (DSF) editor. These remote displays will be based on model images provided by GMPS through PowerPoint. Using the DSF editor, the PowerPoint images can be recreated with active buttons associated with the correct Compact Unique Identifiers (CUIs). These displays will be documented in the Software Requirements and Design Specifications (SRDS) at the 90% GMPS Design Review. In the future, these remote displays will be available for other developers to improve, edit, or add on to so that the display may be incorporated into the firing room to be used for launches.

  16. Interruption and Pausing of Public Display Games

    DEFF Research Database (Denmark)

    Feuchtner, Tiare; Walter, Robert; Müller, Jörg

    2016-01-01

    We present a quantitative and qualitative analysis of interruptions of interaction with a public display game, and explore the use of a manual pause mode in this scenario. In previous public display installations we observed users frequently interrupting their interaction. To explore ways of supp...

  17. Additive and subtractive transparent depth displays

    NARCIS (Netherlands)

    Kooi, F.L.; Toet, A.

    2003-01-01

    Image fusion is the generally preferred method to combine two or more images for visual display on a single screen. We demonstrate that perceptual image separation may be preferable over perceptual image fusion for the combined display of enhanced and synthetic imagery. In this context image separat

  18. Extraction and Analysis of Display Data

    Science.gov (United States)

    Land, Chris; Moye, Kathryn

    2008-01-01

    The Display Audit Suite is an integrated package of software tools that partly automates the detection of Portable Computer System (PCS) Display errors. [PCS is a lap top computer used onboard the International Space Station (ISS).] The need for automation stems from the large quantity of PCS displays (6,000+, with 1,000,000+ lines of command and telemetry data). The Display Audit Suite includes data-extraction tools, automatic error detection tools, and database tools for generating analysis spread sheets. These spread sheets allow engineers to more easily identify many different kinds of possible errors. The Suite supports over 40 independent analyses, 16 NASA Tech Briefs, November 2008 and complements formal testing by being comprehensive (all displays can be checked) and by revealing errors that are difficult to detect via test. In addition, the Suite can be run early in the development cycle to find and correct errors in advance of testing.

  19. 2D/3D switchable displays

    Science.gov (United States)

    Dekker, T.; de Zwart, S. T.; Willemsen, O. H.; Hiddink, M. G. H.; IJzerman, W. L.

    2006-02-01

    A prerequisite for a wide market acceptance of 3D displays is the ability to switch between 3D and full resolution 2D. In this paper we present a robust and cost effective concept for an auto-stereoscopic switchable 2D/3D display. The display is based on an LCD panel, equipped with switchable LC-filled lenticular lenses. We will discuss 3D image quality, with the focus on display uniformity. We show that slanting the lenticulars in combination with a good lens design can minimize non-uniformities in our 20" 2D/3D monitors. Furthermore, we introduce fractional viewing systems as a very robust concept to further improve uniformity in the case slanting the lenticulars and optimizing the lens design are not sufficient. We will discuss measurements and numerical simulations of the key optical characteristics of this display. Finally, we discuss 2D image quality, the switching characteristics and the residual lens effect.

  20. Three-dimensional hologram display system

    Science.gov (United States)

    Mintz, Frederick (Inventor); Chao, Tien-Hsin (Inventor); Bryant, Nevin (Inventor); Tsou, Peter (Inventor)

    2009-01-01

    The present invention relates to a three-dimensional (3D) hologram display system. The 3D hologram display system includes a projector device for projecting an image upon a display medium to form a 3D hologram. The 3D hologram is formed such that a viewer can view the holographic image from multiple angles up to 360 degrees. Multiple display media are described, namely a spinning diffusive screen, a circular diffuser screen, and an aerogel. The spinning diffusive screen utilizes spatial light modulators to control the image such that the 3D image is displayed on the rotating screen in a time-multiplexing manner. The circular diffuser screen includes multiple, simultaneously-operated projectors to project the image onto the circular diffuser screen from a plurality of locations, thereby forming the 3D image. The aerogel can use the projection device described as applicable to either the spinning diffusive screen or the circular diffuser screen.

  1. An interactive multiview 3D display system

    Science.gov (United States)

    Zhang, Zhaoxing; Geng, Zheng; Zhang, Mei; Dong, Hui

    2013-03-01

    The progresses in 3D display systems and user interaction technologies will help more effective 3D visualization of 3D information. They yield a realistic representation of 3D objects and simplifies our understanding to the complexity of 3D objects and spatial relationship among them. In this paper, we describe an autostereoscopic multiview 3D display system with capability of real-time user interaction. Design principle of this autostereoscopic multiview 3D display system is presented, together with the details of its hardware/software architecture. A prototype is built and tested based upon multi-projectors and horizontal optical anisotropic display structure. Experimental results illustrate the effectiveness of this novel 3D display and user interaction system.

  2. Interruption and Pausing of Public Display Games

    DEFF Research Database (Denmark)

    Feuchtner, Tiare; Walter, Robert; Müller, Jörg

    We present a quantitative and qualitative analysis of interruptions of interaction with a public display game, and explore the use of a manual pause mode in this scenario. In previous public display installations we observed users frequently interrupting their interaction. To explore ways...... of supporting such behavior, we implemented a gesture controlled multiuser game with four pausing techniques. We evaluated them in a field study analyzing 704 users and found that our pausing techniques were eagerly explored, but rarely used with the intention to pause the game. Our study shows...... that interactions with public displays are considerably intermissive, and that users mostly interrupt interaction to socialize and mainly approach public displays in groups. We conclude that, as a typical characteristic of public display interaction, interruptions deserve consideration. However, manual pause modes...

  3. The frissonnant mutant mouse, a model of dopamino-sensitive, inherited motor syndrome.

    Science.gov (United States)

    Callizot, N; Guénet, J L; Baillet, C; Warter, J M; Poindron, P

    2001-06-01

    The frissonnant (fri) mutation is an autosomic recessive mutation which spontaneously appeared in the stock of C3H mice. fri mutant mice have locomotor instability and rapid tremor. Since tremor ceases when mutant mice have sleep or are anaesthetized, and because of their obvious stereotyped motor behavior, these mice could represent an inherited Parkinsonian syndrome. We show here that the fri/fri mouse fulfills two out of the three criteria required to validate an experimental model of human disease, that is isomorphism, homology and predictivity. Indeed, fri/fri mice present an important motor deficit accompanying visible tremor and stereotypies. They display some memory deficits as in human Parkinson's desease. l-Dopa and apomorphine (dopaminergic agonists), ropinirole (selective D2 agonist), and selegiline (an monoamino-oxidase B [MAO-B] inhibitor) improve their clinical status. However, neither anatomopathological evidence of nigrostriatal lesion, nor decrease in tyrosine hydroxylase production could be seen.

  4. Pigment content and leaf plastid ultrastructure in the tomato mutant lutescent-2.

    Science.gov (United States)

    Fornasiero, R B; Bonatti, P M

    1985-03-01

    The non-lethal tomato mutant «lutescent-2» shows an early yellowing of normal developed leaves. Its ripe fruits display a yellow colouring, red pigment synthesis being delayed by up to two weeks. Typical pigment synthesis, related to leaf maturation, does not occur in mutant leaves. Both the concentration of chl a and chl b start to decrease very quickly at the end of leaf expansion. Early yellowing of «1-2» leaves appears to be related to the reduced car(470) content, which leads to chlorophyll photooxidation. Structural evidence of a deficiency in car(470) content in young «1- 2» plastids is given by a reduction of stroma thylakoids as well as by a limited grana stacking. The altered balance between the two pigment classes determined an active, even if incomplete, conversion of chloroplasts to chromoplast-like organelles.

  5. Targeting ESR1-Mutant Breast Cancer

    Science.gov (United States)

    2015-09-01

    disease-free survival, the majority of breast cancer patients will present recurrent antiestrogen resistant metastatic lesions following prolonged...exposure to these therapies. By investigating how these lesions become resistant to antiestrogen while maintaining expression of ERα, we found...Retreat. Invited oral presentation. C) Other products Nothing to report. Targeting  ESR1-­‐Mutant  Breast  Cancer   W81XWH-­‐14-­‐1-­‐0360

  6. Characterization of a Legionella micdadei mip mutant

    DEFF Research Database (Denmark)

    O'Connell, W A; Bangsborg, Jette Marie; Cianciotto, N P

    1995-01-01

    The pathogenesis of Legionella micdadei is dependent upon its ability to infect alveolar phagocytes. To better understand the basis of intracellular infection by this organism, we examined the importance of its Mip surface protein. In Legionella pneumophila, Mip promotes infection of both human m...... into the phagocyte. Similarly, the mutant was less able to parasitize Hartmannella amoebae. Taken together, these data argue that Mip specifically potentiates intracellular growth by L. micdadei....

  7. 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.

  8. Improved heterologous protein expression in the chloroplast of Chlamydomonas reinhardtii through promoter and 5' untranslated region optimization.

    Science.gov (United States)

    Rasala, Beth A; Muto, Machiko; Sullivan, Joseph; Mayfield, Stephen P

    2011-08-01

    Microalgae have the potential to be a valuable biotechnological platform for the production of recombinant proteins. However, because of the complex regulatory network that tightly controls chloroplast gene expression, heterologous protein accumulation in a wild-type, photosynthetic-competent algal chloroplast remains low. High levels of heterologous protein accumulation have been achieved using the psbA promoter/5' untranslated region (UTR), but only in a psbA-deficient genetic background, because of psbA/D1-dependent auto-attenuation. Here, we examine the effect of fusing the strong 16S rRNA promoter to the 5' UTR of the psbA and atpA genes on transgene expression in the chloroplast of Chlamydomonas reinhardtii. We show that fusion of the 16S promoter had little impact on protein accumulation from the psbA 5' UTR in a psbA-deficient genetic background. Furthermore, the 16S/psbA promoter/UTR fusion was silenced in the presence of wild-type levels of D1 protein, confirming that the psbA 5' UTR is the primary target for D1-dependent auto-repression. However, fusion of the 16S promoter to the atpA 5' UTR significantly boosts mRNA levels and supports high levels of heterologous protein accumulation in photosynthetic-competent cells. The 16S/atpA promoter/UTR drove LUXCT protein accumulation to levels close to that of psbA in a psbA- background, and drove expression of a human therapeutic protein to levels only twofold lower than the psbA 5' UTR. The 16S/atpA promoter/UTR combination should have utility for heterologous protein production when expression from a photosynthetic-competent microalgal strain is required.

  9. Purification and properties of three NAD(P)+ isozymes of L-glutamate dehydrogenase of Chlamydomonas reinhardtii.

    Science.gov (United States)

    Moyano, E; Cárdenas, J; Muñoz-Blanco, J

    1992-02-13

    Three isozymes of glutamate dehydrogenase (GDH) of Chlamydomonas reinhardtii, induced under different trophic and stress conditions, have been purified about 800-1000-fold to electrophoretic homogeneity. They are hexamers of Mr 266,000-269,000 as deduced from gel filtration and sedimentation coefficient data. GDH1 consisted of six identical subunits of 44 kDa each, whereas both GDH2 and GDH3 consisted of six similar-sized monomers (4 of 44 kDa and 2 of 46 kDa). Optimum pH for the three activities with each pyridine nucleotide was identical (8.5 with NADH; 7.7 with NADPH; and 9.0 with NAD+). The isozymes exhibited similar high optimum temperature values (60-62 degrees C) and isoelectric points (7.9-8.1). Activity was enhanced in vitro by Ca2+ ions and strongly inhibited by pyridoxal 5'-phosphate, KCN, o-phenanthroline and EDTA, and to a lesser extent by pHMB and methylacetimidate. In the aminating reaction the three isozymes were inhibited in a concentration-dependent process by both NADH and NADPH, with apparent Km values for NH4+ ranging from 13-53 mM; 0.36-1.85 mM for 2-oxoglutarate and 0.07-0.78 mM for NADH and NADPH. In the deaminating reaction apparent Km values ranged from 0.64-3.52 mM for L-glutamate and 0.20-0.32 for NAD+. In addition, the three isozymes exhibited a non-hyperbolic kinetics for NAD+ with negative cooperativity (n = 0.8).

  10. Responsibility of regulatory gene expression and repressed protein synthesis for triacylglycerol accumulation on sulfur-starvation in Chlamydomonas reinhardtii.

    Science.gov (United States)

    Sato, Atsushi; Matsumura, Rie; Hoshino, Naomi; Tsuzuki, Mikio; Sato, Norihiro

    2014-01-01

    Triacylglycerol (TG) synthesis is induced for energy and carbon storage in algal cells under nitrogen(N)-starved conditions, and helps prevent reactive oxygen species (ROS) production through fatty acid synthesis that consumes excessive reducing power. Here, the regulatory mechanism for the TG content in sulfur(S)-starved cells of Chlamydomonas reinhardtii was examined, in comparison to that in N- or phosphorus(P)-starved cells. S- and N- starved cells exhibited markedly increased TG contents with up-regulation of mRNA levels of diacylglycerol acyltransferase (DGAT) genes. S-Starvation also induced expression of the genes for phosphatidate synthesis. In contrast, P-starved cells exhibited little alteration of the TG content with almost no induction of these genes. The results implied deficient nutrient-specific regulation of the TG content. An arg9 disruptant defective in arginine synthesis, even without nutritional deficiencies, exhibited an increased TG content upon removal of supplemented arginine, which repressed protein synthesis. Repression of protein synthesis thus seemed crucial for TG accumulation in S- or N- starved cells. Meanwhile, the results of inhibitor experiments involving cells inferred that TG accumulation during S-starvation is supported by photosynthesis and de novo fatty acid synthesis. During S-starvation, sac1 and snrk2.2 disruptants, which are defective in the response to the ambient S-status, accumulated TG at lower and higher levels, respectively, than the wild type. The sac1 and snrk2.2 disruptants showed no or much greater up-regulation of DGAT genes, respectively. In conclusion, TG synthesis would be activated in S-starved cells, through the diversion of metabolic carbon-flow from protein to TG synthesis, and simultaneously through up-regulation of the expression of a particular set of genes for TG synthesis at proper levels through the actions of SAC1 and SNRK2.2.

  11. Responsibility of regulatory gene expression and repressed protein synthesis for triacylglycerol accumulation on sulfur-starvation in Chlamydomonas reinhardtii

    Directory of Open Access Journals (Sweden)

    Atsushi eSato

    2014-09-01

    Full Text Available Triacylglycerol (TG synthesis is induced for energy and carbon storage in algal cells under nitrogen(N-starved conditions, and helps prevent reactive oxygen species production through fatty acid synthesis that consumes excessive reducing power. Here, the regulatory mechanism for the TG content in sulfur(S-starved cells of Chlamydomonas reinhardtii was examined, in comparison to that in N- or phosphorus(P-starved cells. S- and N-starved cells exhibited markedly increased TG contents with up-regulation of mRNA levels of diacylglycerol acyltransferase genes. S-Starvation also induced expression of the genes for phosphatidate synthesis. In contrast, P-starved cells exhibited little alteration of the TG content with almost no induction of these genes. The results implied deficient nutrient-specific regulation of the TG content. An arg9 disruptant defective in arginine synthesis, even without nutritional deficiencies, exhibited an increased TG content upon removal of supplemented arginine, which repressed protein synthesis. Repression of protein synthesis thus seemed crucial for TG accumulation in S- or N-starved cells. Meanwhile, the results of inhibitor experiments involving cells inferred that TG accumulation during S-starvation is supported by photosynthesis and de novo fatty acid synthesis. During S-starvation, sac1 and snrk2.2 disruptants, which are defective in the response to the ambient S-status, accumulated TG at lower and higher levels, respectively, than the wild type. The sac1 and snrk2.2 disruptants showed no or much greater up-regulation of diacylglycerol acyltransferase genes, respectively. In conclusion, TG synthesis would be activated in S-starved cells, through the diversion of metabolic carbon-flow from protein to TG synthesis, and simultaneously through up-regulation of the expression of a particular set of genes for TG synthesis at proper levels through the actions of SAC1 and SNRK2.2.

  12. Multiple-endpoint assay provides a detailed mechanistic view of responses to herbicide exposure in Chlamydomonas reinhardtii

    Energy Technology Data Exchange (ETDEWEB)

    Nestler, Holger [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Ueberlandstrasse 133, 8600 Duebendorf (Switzerland); ETH Zurich, Swiss Federal Institute of Technology, Institute of Biogeochemistry and Pollutant Dynamics, Universitaetstrasse 16, 8092 Zurich (Switzerland); Groh, Ksenia J.; Schoenenberger, Rene; Behra, Renata [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Ueberlandstrasse 133, 8600 Duebendorf (Switzerland); Schirmer, Kristin [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Ueberlandstrasse 133, 8600 Duebendorf (Switzerland); ETH Zurich, Swiss Federal Institute of Technology, Institute of Biogeochemistry and Pollutant Dynamics, Universitaetstrasse 16, 8092 Zurich (Switzerland); EPF Lausanne, School of Architecture, Civil and Environmental Engineering, 1015 Lausanne (Switzerland); Eggen, Rik I.L. [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Ueberlandstrasse 133, 8600 Duebendorf (Switzerland); ETH Zurich, Swiss Federal Institute of Technology, Institute of Biogeochemistry and Pollutant Dynamics, Universitaetstrasse 16, 8092 Zurich (Switzerland); Suter, Marc J.-F., E-mail: suter@eawag.ch [Eawag, Swiss Federal Institute of Aquatic Science and Technology, Ueberlandstrasse 133, 8600 Duebendorf (Switzerland); ETH Zurich, Swiss Federal Institute of Technology, Institute of Biogeochemistry and Pollutant Dynamics, Universitaetstrasse 16, 8092 Zurich (Switzerland)

    2012-04-15

    The release of herbicides into the aquatic environment raises concerns about potential detrimental effects on ecologically important non-target species, such as unicellular algae, necessitating ecotoxicological risk assessment. Algal toxicity tests based on growth, a commonly assessed endpoint, are integrative, and hence do not provide information about underlying toxic mechanisms and effects. This limitation may be overcome by measuring more specific biochemical and physiological endpoints. In the present work, we developed and applied a novel multiple-endpoint assay, and analyzed the effects of the herbicides paraquat, diuron and norflurazon, each representing a specific mechanism of toxic action, on the single celled green alga Chlamydomonas reinhardtii. The endpoints added to assessment of growth were pigment content, maximum and effective photosystem II quantum yield, ATP content, esterase and oxidative activity. All parameters were measured at 2, 6 and 24 h of exposure, except for growth and pigment content, which were determined after 6 and 24 h only. Effective concentrations causing 50% of response (EC50s) and lowest observable effect concentrations (LOECs) were determined for all endpoints and exposure durations where possible. The assay provided a detailed picture of the concentration- and time-dependent development of effects elicited by the analyzed herbicides, thus improving the understanding of the underlying toxic mechanisms. Furthermore, the response patterns were unique to the respective herbicide and reflected the different mechanisms of toxicity. The comparison of the endpoint responses and sensitivities revealed that several physiological and biochemical parameters reacted earlier or stronger to disturbances than growth. Overall, the presented multiple-endpoint assay constitutes a promising basis for investigating stressor and toxicant effects in green algae.

  13. Chlamydomonas reinhardtii Chloroplasts Contain a Homodimeric Pyruvate:Ferredoxin Oxidoreductase That Functions with FDX11[W][OA

    Science.gov (United States)

    van Lis, Robert; Baffert, Carole; Couté, Yohann; Nitschke, Wolfgang; Atteia, Ariane

    2013-01-01

    Eukaryotic algae have long been known to live in anoxic environments, but interest in their anaerobic energy metabolism has only recently gained momentum, largely due to their utility in biofuel production. Chlamydomonas reinhardtii figures remarkably in this respect, because it efficiently produces hydrogen and its genome harbors many genes for anaerobic metabolic routes. Central to anaerobic energy metabolism in many unicellular eukaryotes (protists) is pyruvate:ferredoxin oxidoreductase (PFO), which decarboxylates pyruvate and forms acetyl-coenzyme A with concomitant reduction of l