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Yusuke Matsuda - One of the best experts on this subject based on the ideXlab platform.

  • acclimation of wild type cells and co2 insensitive mutants of the green alga Chlorella ellipsoidea to elevated co2
    Plant Cell and Environment, 2007
    Co-Authors: Teruhiko Ochiai, Brian Colman, Yusuke Matsuda
    Abstract:

    CO(2)-insensitive mutants of the green alga Chlorella ellipsoidea were previously shown to be unable to repress an inorganic carbon-concentrating mechanism (CCM) when grown under 5% CO(2). When air-grown, wild-type (WT) cells were transferred to 5% CO(2), an abrupt drop of P(max) to 43% the original level of air-grown cells was observed within the initial 12 h. Photosynthetic affinities of WT cells to dissolved inorganic carbon (DIC) were maintained at high levels for the initial 4 d of acclimation, and then decreased gradually to lower levels over the next 6 d. In contrast to WT cells, the CO(2)-insensitive mutant, ENU16, exhibited a constant P(max) at maximum levels and a low K(1/2)[DIC] throughout the acclimation period. The rapid P(max) drop within 12 h of acclimation in WT cells was significantly reduced by treatment with 0.5 mm of 6-ethoxybenzothiazole-2-sulphonamide (EZA), a specific membrane-permeable inhibitor of carbonic anhydrase (CA), suggesting the participation of internal CAs in the temporary drop in P(max) in WT cells. WT and ENU16 cells were grown in controlled equilibrium [CO(2)], and the photosynthetic rate of each acclimated cell type was measured under equilibrated growth [DIC] conditions. In WT cells acclimated to 0.14-0.4% [CO(2)], K(1/2)[DIC] values increased as [CO(2)] increased, and the photosynthetic rates at growth DIC conditions were shown to decrease to about 70% the P(max) level in this intermediate [CO(2)] range. Such decreases in the net photosynthetic rates were not observed in ENU16. These results suggest that algal primary production could be depressed significantly under moderately enriched CO(2) conditions as a result of acquiring intermediate affinities for DIC because of their sensitive responses to changes in the ambient [CO(2)].

  • Acclimation of wild‐type cells and CO2‐insensitive mutants of the green alga Chlorella ellipsoidea to elevated [CO2]
    Plant cell & environment, 2007
    Co-Authors: Teruhiko Ochiai, Brian Colman, Yusuke Matsuda
    Abstract:

    CO(2)-insensitive mutants of the green alga Chlorella ellipsoidea were previously shown to be unable to repress an inorganic carbon-concentrating mechanism (CCM) when grown under 5% CO(2). When air-grown, wild-type (WT) cells were transferred to 5% CO(2), an abrupt drop of P(max) to 43% the original level of air-grown cells was observed within the initial 12 h. Photosynthetic affinities of WT cells to dissolved inorganic carbon (DIC) were maintained at high levels for the initial 4 d of acclimation, and then decreased gradually to lower levels over the next 6 d. In contrast to WT cells, the CO(2)-insensitive mutant, ENU16, exhibited a constant P(max) at maximum levels and a low K(1/2)[DIC] throughout the acclimation period. The rapid P(max) drop within 12 h of acclimation in WT cells was significantly reduced by treatment with 0.5 mm of 6-ethoxybenzothiazole-2-sulphonamide (EZA), a specific membrane-permeable inhibitor of carbonic anhydrase (CA), suggesting the participation of internal CAs in the temporary drop in P(max) in WT cells. WT and ENU16 cells were grown in controlled equilibrium [CO(2)], and the photosynthetic rate of each acclimated cell type was measured under equilibrated growth [DIC] conditions. In WT cells acclimated to 0.14-0.4% [CO(2)], K(1/2)[DIC] values increased as [CO(2)] increased, and the photosynthetic rates at growth DIC conditions were shown to decrease to about 70% the P(max) level in this intermediate [CO(2)] range. Such decreases in the net photosynthetic rates were not observed in ENU16. These results suggest that algal primary production could be depressed significantly under moderately enriched CO(2) conditions as a result of acquiring intermediate affinities for DIC because of their sensitive responses to changes in the ambient [CO(2)].

  • EPR study on the photo-reduction of spin probe in the green alga Chlorella ellipsoidea
    Science Access, 2001
    Co-Authors: Shuji Yamada, Yusuke Matsuda, Asako Kawamori
    Abstract:

    CO2 sensing system is believed to exist on the surface of plasmalemma of the green alga Chlorella ellipsoidea. It is well known that C. ellipsoidea can adjust the activity of carbon concentrating mechanism (CCM) in response to growth CO2 concentrations. We prepared two wild-type cells of C. ellipsoidea, i.e., High CO2 (5%)-grown cells and Low CO2-grown cells that have low and high CCM activities, respectively. CO2 insensitive mutants of C. ellipsoidea, which express the maximum level of CCM even grown in 5% CO2, were also cultured under low and high CO2-conditions. Cells were treated with 2 mM of 4-oxo-TEMPO(2,2,6,6-tetramethyl-4-oxopiperidinonxy) (TEMPONE), a spin probe, and 40 mM of K3Fe(CN)6, an oxidant. EPR signals from intracellular TEMPONE were observed under illumination or the dark in the presence of 150 mM of Na2MnEDTA, a signal quencher for external TEMPONE. The signal intensity of intracellular TEMPONE decreased gradually in a time dependent manner even under the dark, but the rate of reduction was stimulated as light intensity was increased. Two mM of DCMU, a specific inhibitor for the electron transport from QA to QB, halted the reduction of EPR signal. These data indicate that intracellular TEMPONE is reduced by photosynthetic electron flow primarily at PS II. Furthermore, the difference in effects of wavelength of light on the photoreduction of TEMPONE were also investigated.

  • A New Screening Method for Algal Photosynthetic Mutants (CO2-Insensitive Mutants of the Green Alga Chlorella ellipsoidea).
    Plant physiology, 1996
    Co-Authors: Yusuke Matsuda, B. Colman
    Abstract:

    A new method has been developed for screening algal photosynthetic mutants. This method uses autoradiography to assess gross photosynthetic 14C fixation by green algal colonies on agar plates and allows the identification of clones that differ in photosynthetic characteristics from wild-type cells. Three wild-type cells, high-CO2-grown Chlorella ellipsoidea, air-grown C. ellipsoidea, and air-grown Chlorella saccharophila, had K0.5 values for dissolved inorganic carbon (DIC) of 1083, 250, and 50 [mu]M, respectively, and as plaques on agar plates at Chl densities greater than 25 [mu]g cm-2 exhibited relative amounts of 14C fixation of 15, 55, and 100%, respectively. Cells of C. ellipsoidea were mutagenized with x-rays and screened by this method. Growth of C. ellipsoidea in high CO2 represses DIC transport and thus lowers its affinity for DIC. Five of the mutants detected by this method showed high-affinity photosynthesis similar to air-grown wild-type cells even when grown in high CO2. Seven other mutants when grown in high CO2 showed affinities for DIC intermediate between air-grown and high-CO2-grown wild-type cells. The affinities of high-CO2-grown mutants were reflected precisely in their capacities to accumulate DIC intracellularly. These results indicate that the mutants are fully or partially insensitive to the repressive effect of ambient CO2 concentration on DIC transport.

  • Induction of CO2 and Bicarbonate Transport in the Green Alga Chlorella ellipsoidea (II. Evidence for Induction in Response to External CO2 Concentration).
    Plant physiology, 1995
    Co-Authors: Yusuke Matsuda, Brian Colman
    Abstract:

    The critical species and concentrations of dissolved inorganic carbon (DIC) required for the induction of DIC transport during adaptation to low CO2 were determined for the green alga Chlorella ellipsoidea. The concentration of dissolved CO2 needed for the induction of both CO2 and HCO3- transport was independent of pH during adaptation, whereas the total DIC concentration required increased at alkaline pH. At pH 7.5, the minimum equilibrium DIC concentration at which high CO2 characteristics were maintained, i.e. transport was repressed, was 2100 [mu]M, whereas the maximum equilibrium DIC concentration below which DIC transport was fully induced (DICIND) was 500 [mu]M. Intracellular DIC concentration during adaptation to DICIND decreased temporarily after 2 h to 60% of the maximum level but recovered after 3 h of adaptation. After 3 h of adaptation to DICIND, cells exhibited maximum O2 evolution rate at DICIND. When cells partially adapted to DICIND were returned to high CO2, there was an immediate halt to the induction of transport and a gradual decrease in transport capacity over 23 h. The capacity for the induction of transport was unaffected by the absence of light. These results indicate that changes in the internal DIC pool during adaptation to low CO2 do not trigger the induction of DIC transport and that the induction is not light dependent. Induction of DIC transport in C. ellipsoidea appears to occur in response to the continuous exposure of cells to a critical CO2 concentration in the external medium.

Brian Colman - One of the best experts on this subject based on the ideXlab platform.

  • acclimation of wild type cells and co2 insensitive mutants of the green alga Chlorella ellipsoidea to elevated co2
    Plant Cell and Environment, 2007
    Co-Authors: Teruhiko Ochiai, Brian Colman, Yusuke Matsuda
    Abstract:

    CO(2)-insensitive mutants of the green alga Chlorella ellipsoidea were previously shown to be unable to repress an inorganic carbon-concentrating mechanism (CCM) when grown under 5% CO(2). When air-grown, wild-type (WT) cells were transferred to 5% CO(2), an abrupt drop of P(max) to 43% the original level of air-grown cells was observed within the initial 12 h. Photosynthetic affinities of WT cells to dissolved inorganic carbon (DIC) were maintained at high levels for the initial 4 d of acclimation, and then decreased gradually to lower levels over the next 6 d. In contrast to WT cells, the CO(2)-insensitive mutant, ENU16, exhibited a constant P(max) at maximum levels and a low K(1/2)[DIC] throughout the acclimation period. The rapid P(max) drop within 12 h of acclimation in WT cells was significantly reduced by treatment with 0.5 mm of 6-ethoxybenzothiazole-2-sulphonamide (EZA), a specific membrane-permeable inhibitor of carbonic anhydrase (CA), suggesting the participation of internal CAs in the temporary drop in P(max) in WT cells. WT and ENU16 cells were grown in controlled equilibrium [CO(2)], and the photosynthetic rate of each acclimated cell type was measured under equilibrated growth [DIC] conditions. In WT cells acclimated to 0.14-0.4% [CO(2)], K(1/2)[DIC] values increased as [CO(2)] increased, and the photosynthetic rates at growth DIC conditions were shown to decrease to about 70% the P(max) level in this intermediate [CO(2)] range. Such decreases in the net photosynthetic rates were not observed in ENU16. These results suggest that algal primary production could be depressed significantly under moderately enriched CO(2) conditions as a result of acquiring intermediate affinities for DIC because of their sensitive responses to changes in the ambient [CO(2)].

  • Acclimation of wild‐type cells and CO2‐insensitive mutants of the green alga Chlorella ellipsoidea to elevated [CO2]
    Plant cell & environment, 2007
    Co-Authors: Teruhiko Ochiai, Brian Colman, Yusuke Matsuda
    Abstract:

    CO(2)-insensitive mutants of the green alga Chlorella ellipsoidea were previously shown to be unable to repress an inorganic carbon-concentrating mechanism (CCM) when grown under 5% CO(2). When air-grown, wild-type (WT) cells were transferred to 5% CO(2), an abrupt drop of P(max) to 43% the original level of air-grown cells was observed within the initial 12 h. Photosynthetic affinities of WT cells to dissolved inorganic carbon (DIC) were maintained at high levels for the initial 4 d of acclimation, and then decreased gradually to lower levels over the next 6 d. In contrast to WT cells, the CO(2)-insensitive mutant, ENU16, exhibited a constant P(max) at maximum levels and a low K(1/2)[DIC] throughout the acclimation period. The rapid P(max) drop within 12 h of acclimation in WT cells was significantly reduced by treatment with 0.5 mm of 6-ethoxybenzothiazole-2-sulphonamide (EZA), a specific membrane-permeable inhibitor of carbonic anhydrase (CA), suggesting the participation of internal CAs in the temporary drop in P(max) in WT cells. WT and ENU16 cells were grown in controlled equilibrium [CO(2)], and the photosynthetic rate of each acclimated cell type was measured under equilibrated growth [DIC] conditions. In WT cells acclimated to 0.14-0.4% [CO(2)], K(1/2)[DIC] values increased as [CO(2)] increased, and the photosynthetic rates at growth DIC conditions were shown to decrease to about 70% the P(max) level in this intermediate [CO(2)] range. Such decreases in the net photosynthetic rates were not observed in ENU16. These results suggest that algal primary production could be depressed significantly under moderately enriched CO(2) conditions as a result of acquiring intermediate affinities for DIC because of their sensitive responses to changes in the ambient [CO(2)].

  • Induction of CO2 and Bicarbonate Transport in the Green Alga Chlorella ellipsoidea (II. Evidence for Induction in Response to External CO2 Concentration).
    Plant physiology, 1995
    Co-Authors: Yusuke Matsuda, Brian Colman
    Abstract:

    The critical species and concentrations of dissolved inorganic carbon (DIC) required for the induction of DIC transport during adaptation to low CO2 were determined for the green alga Chlorella ellipsoidea. The concentration of dissolved CO2 needed for the induction of both CO2 and HCO3- transport was independent of pH during adaptation, whereas the total DIC concentration required increased at alkaline pH. At pH 7.5, the minimum equilibrium DIC concentration at which high CO2 characteristics were maintained, i.e. transport was repressed, was 2100 [mu]M, whereas the maximum equilibrium DIC concentration below which DIC transport was fully induced (DICIND) was 500 [mu]M. Intracellular DIC concentration during adaptation to DICIND decreased temporarily after 2 h to 60% of the maximum level but recovered after 3 h of adaptation. After 3 h of adaptation to DICIND, cells exhibited maximum O2 evolution rate at DICIND. When cells partially adapted to DICIND were returned to high CO2, there was an immediate halt to the induction of transport and a gradual decrease in transport capacity over 23 h. The capacity for the induction of transport was unaffected by the absence of light. These results indicate that changes in the internal DIC pool during adaptation to low CO2 do not trigger the induction of DIC transport and that the induction is not light dependent. Induction of DIC transport in C. ellipsoidea appears to occur in response to the continuous exposure of cells to a critical CO2 concentration in the external medium.

  • Induction of CO2 and Bicarbonate Transport in the Green Alga Chlorella ellipsoidea (I. Time Course of Induction of the Two Systems)
    Plant physiology, 1995
    Co-Authors: Yusuke Matsuda, Brian Colman
    Abstract:

    Changes in the physiological properties of the green alga Chlorella ellipsoidea (UTEX 20) were determined during adaptation from high CO2 to air. Cells of C. ellipsoidea, grown in high CO2, had an extremely low affinity for dissolved inorganic carbon (DIC). However, high-affinity DIC transport was induced rapidly after switching to air, which caused a massive decrease in the DIC concentration in the medium. Rates of O2 evolution without added carbonic anhydrase (CA) were compared with calculated rates of uncatalyzed CO2 formation in the medium as a measure of active HCO3-uptake. Cells were found to be able to use HCO3- after 5 h of adaptation and this capacity increased during the next 17 h. The stimulation of O2 evolution upon CA addition was used as a measurement of active CO2 transport: such stimulation occurred 2 h after transfer and increased during the next 5 h. Increases in O2 evolution rates were correlated closely with an increasing capacity to accumulate intracellular pools of acid-labile DIC and with decreases in K1/2(CO2) and CO2-compensation point of the cells. Treatment of cells with cycloheximide (5 [mu]g mL-1) during adaptation completely inhibited DIC transport induction, whereas treatment with chloramphenicol (400 [mu]g mL-1) had no effect, indicating the requirement for cytoplasmic protein synthesis in the induction. These results suggest that both CO2 and HCO3- transport are induced upon transfer of cells from high CO2 to air and that there is a temporal separation between the induction of the two systems.

  • The localization of active inorganic carbon transport at the plasma membrane in Chlorella ellipsoidea
    Canadian Journal of Botany, 1991
    Co-Authors: Caterina Rotatore, Brian Colman
    Abstract:

    A study has been made of the capacity of whole cells and intact, photosynthetically active, isolated chloroplasts of Chlorella ellipsoidea (UTEX 20) to take up CO2 by active transport. Assays for carbonic anhydrase activity and the monitoring of CO2 uptake by cells and chloroplasts was carried out by mass spectrometry. No external carbonic anhydrase was detected in whole cells or on the outer surface of the isolated chloroplasts. Upon illumination, whole cells rapidly depleted CO2 from the medium to a level below the equilibrium CO2 concentration before maximum photosynthetic O2 evolution rates were established. Addition of bovine carbonic anhydrase resulted in the reestablishment of equilibrium CO2 concentrations, indicating that the cells were actively and selectively depleting the medium of CO2. This CO2 uptake was inhibited by 10 μM diethylstilbestrol. No such rapid depletion of CO2 was observed with isolated intact chloroplasts, although the organelles demonstrated rates of photosynthetic O2 evolutio...

Katsuhiko Fujii - One of the best experts on this subject based on the ideXlab platform.

Yongru Sun - One of the best experts on this subject based on the ideXlab platform.

  • Expression of rabbit neutrophile peptide-1 in nitrate reductase-deficient mutant of Chlorella ellipsoidea
    Yi chuan = Hereditas, 2006
    Co-Authors: Xiao-yu Zhang, Shimin Zhao, Yongru Sun, Peng Wang, Xin Shen, Chengcai Chu, Yiqin Wang
    Abstract:

    Application of transgenic Cholrella as bioreactor to express rabbit neutrophile pepetide-1 (NP-1) shows great practical value. In this paper, an NP-1 expression vector containing two selective marker genes NPTII and nitrate reductase gene was constructed. The NP-1 gene was transformed into the nitrate reductase-deficient mutant nrm-4 of Chlorella ellipsoidea via electroporation, and the transgenic alga expressed the active NP-1 were obtained.

  • Isolation and characterization of a nitrate reductase deficient mutant of Chlorella ellipsoidea (Chlorophyta)
    Journal of Applied Phycology, 2005
    Co-Authors: Yiqin Wang, Shimin Zhao, Xiao-yu Zhang, Peng Wang, Lei Zhang, Degui Geng, Yan Chen, Yongru Sun
    Abstract:

    Using X-ray mutagenesis and a chlorate-resistance selection method, a nitrate reductase (NR)-deficient mutant of Chlorella ellipsoidea nrm-4 was isolated, which is incapable of using NO3− as a nitrogen source. NR activity was not detected in nrm-4, suggesting this is a null mutation. Molecular analysis revealed that nrm-4 has a two base deletion at position 2348–49 of its DNA sequence, which produced a frame-shift mutation. The nrm-4 mutation is stable and nrm-4 algae could only use NH4+ and NO2− as nitrogen sources. Expression of a wild-type NR gene could complement this NR-deficient mutant. Furthermore, this transgenic strain was able to grow in NO3− medium, when the growth rate of the nrm-4 strain was equal to that of the wide type alga. The nrm-4 strain could potentially be used as a bioreactor that uses nitrate as a selectable marker instead of an antibiotic or herbicide.

  • Rapid isolation and functional analysis of promoter sequences of the nitrate reductase gene from Chlorella ellipsoidea
    Journal of Applied Phycology, 2004
    Co-Authors: Peng Wang, Yongru Sun, Lei Zhang, Yiqin Wang
    Abstract:

    The thermal asymmetric interlaced (TAIL) PCR technique was used to isolate the 5'-flanking regions of the nitrate reductase (NR) gene of Chlorella ellipsoidea. Isolated 5'-flanking regions were fused to the GUS gene, and tested for transient expression in the alga. The isolated 5'-flanking regions were able to drive GUS reporter gene expression with a strong activity, showing that the isolated 5'-flanking regions contained the promoter elements necessary for the transcription of the NR gene. The expression pattern of the GUS gene and the transcriptional pattern of the NR gene are similar, which demonstrates that NR-specific regulation is mainly under transcriptional control. The nucleotide sequence data reported have been deposited in the DDBJ/EMBL/GenBank databases under the accession number AY307383.

  • Molecular cloning and characterization of a cDNA encoding nitrate reductase from Chlorella ellipsoidea (Chlorophyta)
    Journal of Applied Phycology, 2003
    Co-Authors: Peng Wang, Yiqin Wang, Degui Geng, Yongru Sun
    Abstract:

    Nitrate reductase (NR) genes have beencloned from higher plants, fungi and algae.Based on seven of the amino acid residuesmost strongly conserved between Chlorella vulgaries and Chlamydomonasreinhardtii NR gene, a degenerate primerwas designed. This degenerate primer wasused to amplify the corresponding homologyin Chlorella ellipsoidea. A 3304 bpfull-length cDNA was cloned by rapidamplification of cDNA ends (RACE). Thededuced amino acid sequence of this cDNAhas a high degree of similarity withpreviously identified members of the NRgene. This suggests that the amplified cDNAencodes a functional NR. Northern blotexpression analysis suggests that this geneis strongly induced by nitrate, but isrepressed by ammonium. The nucleotidesequence data reported in this paper willappear in the DDBJ/EMBL/GeneBank databasesunder accession number AY275834.

  • Highly efficient expression of rabbit neutrophil peptide-1 gene in Chlorella ellipsoidea cells
    Current Genetics, 2001
    Co-Authors: Ying Chen, Yiqin Wang, Yongru Sun, Lei Zhang
    Abstract:

    A highly efficient system was developed for the expression of foreign genes in Chlorella ellipsoidea cells. The effect of five promoters on the expression efficiency of β-glucuronidase (GUS) gene was evaluated by transient expression of the UidA gene. Among these promoters, Ubiquitin-Ω was found to be the most efficient and was selected to drive the expression of foreign genes in Chlorella cells. A gene encoding the mature rabbit neutrophil peptide-1 (NP-1) was introduced into the cells. Integration of the gene for NP-1 into the Chlorella genome was confirmed by PCR and Southern blot analysis. In vitro anti-microbial testes demonstrated the expression of biologically active NP-1 by the transgenic Chlorella cells.

Naoto Urano - One of the best experts on this subject based on the ideXlab platform.