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PubMed2000

Peroxide processing in photosynthesis: antioxidant coupling and redox signalling.

Photosynthesis has a high capacity for production of hydrogen peroxide (H2O2), but the intracellular levels of this relatively weak oxidant are controlled by the antioxidant system, comprising a network of enzymatic and non-enzymatic components that notably includes reactions linked to the intracellular ascorbate and glutathione pools. Mutants and transformed plants with specific decreases in key components offer the opp ortunity to dissect the complex system that maintains redox homeostasis. Since H2O2 is a signal-transducing molecule relaying information on intracellular redox state, the pool size must be rigorously controlled within each compartment of the cell. This review focuses on compartment-specific differences in the stringency of redox coupling between ascorbate and glutathione, and the significance this may have for the flexibility of the control of gene expression that is linked to photosynthetic H2O2 production.

Antioxidants↗
PubMed2002

Sensitivity of photosynthesis in a C4 plant, maize, to heat stress.

Our objective was to determine the sensitivity of components of the photosynthetic apparatus of maize (Zea mays), a C4 plant, to high temperature stress. Net photosynthesis (Pn) was inhibited at leaf temperatures above 38 degrees C, and the inhibition was much more severe when the temperature was increased rapidly rather than gradually. Transpiration rate increased progressively with leaf temperature, indicating that inhibition was not associated with stomatal closure. Nonphotochemical fluorescence quenching (qN) increased at leaf temperatures above 30 degrees C, indicating increased thylakoid energization even at temperatures that did not inhibit Pn. Compared with CO(2) assimilation, the maximum quantum yield of photosystem II (F(v)/F(m)) was relatively insensitive to leaf temperatures up to 45 degrees C. The activation state of phosphoenolpyruvate carboxylase decreased marginally at leaf temperatures above 40 degrees C, and the activity of pyruvate phosphate dikinase was insensitive to temperature up to 45 degrees C. The activation state of Rubisco decreased at temperatures exceeding 32.5 degrees C, with nearly complete inactivation at 45 degrees C. Levels of 3-phosphoglyceric acid and ribulose-1,5-bisphosphate decreased and increased, respectively, as leaf temperature increased, consistent with the decrease in Rubisco activation. When leaf temperature was increased gradually, Rubisco activation acclimated in a similar manner as Pn, and acclimation was associated with the expression of a new activase polypeptide. Rates of Pn calculated solely from the kinetics of Rubisco were remarkably similar to measured rates if the calculation included adjustment for temperature effects on Rubisco activation. We conclude that inactivation of Rubisco was the primary constraint on the rate of Pn of maize leaves as leaf temperature increased above 30 degrees C.

Acclimatization↗
PubMed2002

Acclimation of rice photosynthesis to irradiance under field conditions.

Acclimation to irradiance was measured in terms of light-saturated photosynthetic carbon assimilation rates (P(max)), Rubisco, and pigment content in mature field-grown rice (Oryza sativa) plants in tropical conditions. Measurements were made at different positions within the canopy alongside irradiance and daylight spectra. These data were compared with a second experiment in which acclimation to irradiance was assessed in uppermost leaves within whole-plant shading regimes (10% low light [LL], 40% medium light [ML], and 100% high light [HL] of full natural sunlight). Two varieties, japonica (tropical; new plant type [NPT]) and indica (IR72) were compared. Values for Rubisco amount, chlorophyll a/b, and P(max) all declined from the top to the base of the canopy. In the artificial shading experiment, acclimation of P(max) (measured at 350 microL L(-1) CO(2)) occurred between LL and ML for IR72 with no difference observed between ML and HL. The Rubisco amount increased between ML and HL in IR72. A different pattern was seen for NPT with higher P(max) (measured at 350 microL L(-1) CO(2)) at LL than IR72 and some acclimation of this parameter between ML and HL. Rubisco levels were higher in NPT than IR72 contrasting with P(max). Comparison of data from both experiments suggests a leaf aging effect between the uppermost two leaf positions, which was not a result of irradiance acclimation. Results are discussed in terms of: (a) acclimation of photosynthesis and radiation use efficiency at high irradiance in rice, and (b) factors controlling photosynthetic rates of leaves within the canopy.

Acclimatization↗
PubMed2003

Energy status and its control on embryogenesis of legumes. Embryo photosynthesis contributes to oxygen supply and is coupled to biosynthetic fluxes.

Legume seeds are heterotrophic and dependent on mitochondrial respiration. Due to the limited diffusional gas exchange, embryos grow in an environment of low oxygen. O(2) levels within embryo tissues were measured using microsensors and are lowest in early stages and during night, up to 0.4% of atmospheric O(2) concentration (1.1 micro M). Embryo respiration was more strongly inhibited by low O(2) during earlier than later stages. ATP content and adenylate energy charge were lowest in young embryos, whereas ethanol emission and alcohol dehydrogenase activity were high, indicating restricted ATP synthesis and fermentative metabolism. In vitro and in vivo experiments further revealed that embryo metabolism is O(2) limited. During maturation, ATP levels increased and fermentative metabolism disappeared. This indicates that embryos become adapted to the low O(2) and can adjust its energy state on a higher level. Embryos become green and photosynthetically active during differentiation. Photosynthetic O(2) production elevated the internal level up to approximately 50% of atmospheric O(2) concentration (135 micro M). Upon light conditions, embryos partitioned approximately 3-fold more [(14)C]sucrose into starch. The light-dependent increase of starch synthesis was developmentally regulated. However, steady-state levels of nucleotides, free amino acids, sugars, and glycolytic intermediates did not change upon light or dark conditions. Maturing embryos responded to low O(2) supply by adjusting metabolic fluxes rather than the steady-state levels of metabolites. We conclude that embryogenic photosynthesis increases biosynthetic fluxes probably by providing O(2) and energy that is readily used for biosynthesis and respiration.

Adenosine Triphosphate↗
PubMed2003

Knock-out of the genes coding for the Rieske protein and the ATP-synthase delta-subunit of Arabidopsis. Effects on photosynthesis, thylakoid protein composition, and nuclear chloroplast gene expression.

In Arabidopsis, the nuclear genes PetC and AtpD code for the Rieske protein of the cytochrome b(6)/f (cyt b(6)/f) complex and the delta-subunit of the chloroplast ATP synthase (cpATPase), respectively. Knock-out alleles for each of these loci have been identified. Greenhouse-grown petc-2 and atpd-1 mutants are seedling lethal, whereas heterotrophically propagated plants display a high-chlorophyll (Chl)-fluorescence phenotype, indicating that the products of PetC and AtpD are essential for photosynthesis. Additional effects of the mutations in axenic culture include altered leaf coloration and increased photosensitivity. Lack of the Rieske protein affects the stability of cyt b(6)/f and influences the level of other thylakoid proteins, particularly those of photosystem II. In petc-2, linear electron flow is blocked, leading to an altered redox state of both the primary quinone acceptor Q(A) in photosystem II and the reaction center Chl P700 in photosystem I. Absence of cpATPase-delta destabilizes the entire cpATPase complex, whereas residual accumulation of cyt b(6)/f and of the photosystems still allows linear electron flow. In atpd-1, the increase in non-photochemical quenching of Chl fluorescence and a higher de-epoxidation state of xanthophyll cycle pigments under low light is compatible with a slower dissipation of the transthylakoid proton gradient. Further and clear differences between the two mutations are evident when mRNA expression profiles of nucleus-encoded chloroplast proteins are considered, suggesting that the physiological states conditioned by the two mutations trigger different modes of plastid signaling and nuclear response.

Arabidopsis↗
PubMed2003

Photosynthesis and growth of tobacco with a substituted bacterial Rubisco mirror the properties of the introduced enzyme.

Complete replacement, by biolistic plastid transformation, of the hexadecameric ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) of tobacco (Nicotiana tabacum) with the dimeric version from the bacterium, Rhodospirillum rubrum, resulted in fully autotrophic and reproductive tobacco plants that required high CO(2) concentrations to grow (Whitney SM, Andrews TJ [2001] Proc Natl Acad Sci USA 98: 14738-14743). Growth and photosynthesis of these plants was compared with that of nontransformed tobacco and other controls where the rbcL gene for the large subunit of tobacco Rubisco was linked to the aadA selectable-marker gene, simulating the gene arrangement of the transformants with R. rubrum Rubisco. An arrangement of the rbcL and aadA genes that gave rise to an abundant monocistronic rbcL transcript and a one-fifth as abundant bicistronic rbcL-aadA transcript had Rubisco levels and photosynthetic properties similar to those of nontransformed tobacco. Direct linkage of the rbcL and aadA genes, resulting in exclusive production of a bicistronic mRNA transcript analogous to that of the transformants with R. rubrum Rubisco, reduced transcript abundance and tobacco Rubisco content. The analogous transcript with the R. rubrum rbcM gene substituted for rbcL was not only reduced in abundance, but was also translated less efficiently. The photosynthetic rates of the transformants and controls were measured at high CO(2) concentrations, using a mass spectrometric method. The rates and their responses to atmospheric CO(2) concentration mirrored the amounts and the kinetic properties of the Rubiscos present. The contents of total nitrogen, carbohydrates, and photosynthetic metabolites of the leaves were also consistent with the content and type of Rubisco.

Carbohydrate Metabolism↗
PubMed2003

Rubisco activase is required for optimal photosynthesis in the green alga Chlamydomonas reinhardtii in a low-CO(2) atmosphere.

This report describes a Chlamydomonas reinhardtii mutant that lacks Rubisco activase (Rca). Using the BleR (bleomycin resistance) gene as a positive selectable marker for nuclear transformation, an insertional mutagenesis screen was performed to select for cells that required a high-CO2 atmosphere for optimal growth. The DNA flanking the BleR insert of one of the high-CO2-requiring strains was cloned using thermal asymmetric interlaced-polymerase chain reaction and inverse polymerase chain reaction and sequenced. The flanking sequence matched the C. reinhardtii Rca cDNA sequence previously deposited in the National Center for Biotechnology Information database. The loss of a functional Rca in the strain was confirmed by the absence of Rca mRNA and protein. The open reading frame for Rca was cloned and expressed in pSL18, a C. reinhardtii expression vector conferring paromomycin resistance. This construct partially complemented the mutant phenotype, supporting the hypothesis that the loss of Rca was the reason the mutant grew poorly in a low-CO2 atmosphere. Sequencing of the C. reinhardtii Rca gene revealed that it contains 10 exons ranging in size from 18 to 470 bp. Low-CO2-grown rca1 cultures had a growth rate and maximum rate of photosynthesis 60% of wild-type cells. Results obtained from experiments on a cia5 rca1 double mutant also suggest that the CO2-concentrating mechanism partially compensates for the absence of an active Rca in the green alga C. reinhardtii.

Animals↗
PubMed2004

Impact of altered gibberellin metabolism on biomass accumulation, lignin biosynthesis, and photosynthesis in transgenic tobacco plants.

Gibberellins (GAs) are involved in regulation of many aspects during plant development. To investigate the impact of altered GA levels on plant growth and metabolism, transgenic tobacco (Nicotiana tabacum) plants have been engineered to express either a GA20-oxidase (AtGA20-ox) or a GA2-oxidase (AtGA2-ox) gene from Arabidopsis under control of the cauliflower mosaic virus 35S promoter. Resulting plants were characterized by elongated or stunted shoot growth, respectively, indicating changes in the content of bioactive GAs. In accordance with the effect on plant growth, biomass production was increased or decreased in AtGA20-ox or AtGA2-ox plants, respectively, and was found to be positively correlated with the rate of photosynthesis as determined at the whole plant level. Differences in dry matter accumulation were most likely due to changes in lignin deposition as indicated by histochemical staining and quantitative measurements. Altered lignification of transgenic plants was paralleled by up- or down-regulation of the expression of lignin biosynthetic genes. Short-term GA3 feeding of excised petioles induced lignin formation in the absence of a transcriptional activation of pathway-specific genes. Thus, short-term GA treatment mediates lignin deposition most likely by polymerization of preformed monomers, whereas long-term effects on lignification involve elevated production of precursors by transcriptional stimulation of the biosynthetic pathway. Interestingly, analysis of stem cross sections revealed a differential effect of GA on the formation of xylem and pith cells. The number of lignified vessels was increased in AtGA20-ox plants pointing to a stimulation of xylem formation while the number of pith cells declined indicating a negative regulation.

Arabidopsis Proteins↗
PubMed2004

Traffic lights in trichodesmium. Regulation of photosynthesis for nitrogen fixation studied by chlorophyll fluorescence kinetic microscopy.

We investigated interactions between photosynthesis and nitrogen fixation in the non-heterocystous marine cyanobacterium Trichodesmium IMS101 at the single-cell level by two-dimensional (imaging) microscopic measurements of chlorophyll fluorescence kinetics. Nitrogen fixation was closely associated with the appearance of cells with high basic fluorescence yield (F(0)), termed bright cells. In cultures aerated with normal air, both nitrogen fixation and bright cells appeared in the middle of the light phase. In cultures aerated with 5% oxygen, both processes occurred at a low level throughout most of the day. Under 50% oxygen, nitrogen fixation commenced at the beginning of the light phase but declined soon afterwards. Rapid reversible switches between fluorescence levels were observed, which indicated that the elevated F(0) of the bright cells originates from reversible uncoupling of the photosystem II (PSII) antenna from the PSII reaction center. Two physiologically distinct types of bright cells were observed. Type I had about double F(0) compared to the normal F(0) in the dark phase and a PSII activity, measured as variable fluorescence (F(v) = F(m) - F(0)), similar to normal non-diazotrophic cells. Correlation of type I cells with nitrogen fixation, oxygen concentration, and light suggests that this physiological state is connected to an up-regulation of the Mehler reaction, resulting in oxygen consumption despite functional PSII. Type II cells had more than three times the normal F(0) and hardly any PSII activity measurable by variable fluorescence. They did not occur under low-oxygen concentrations, but appeared under high-oxygen levels outside the diazotrophic period, suggesting that this state represents a reaction to oxidative stress not necessarily connected to nitrogen fixation. In addition to the two high-fluorescence states, cells were observed to reversibly enter a low-fluorescence state. This occurred mainly after a cell went through its bright phase and may represent a fluorescence-quenching recovery phase.

Chlorophyll↗
PubMed2005

Reductions of Rubisco activase by antisense RNA in the C4 plant Flaveria bidentis reduces Rubisco carbamylation and leaf photosynthesis.

To function, the catalytic sites of Rubisco (EC 4.1.1.39) need to be activated by the reversible carbamylation of a lysine residue within the sites followed by rapid binding of magnesium. The activation of Rubisco in vivo requires the presence of the regulatory protein Rubisco activase. This enzyme is thought to aid the release of sugar phosphate inhibitors from Rubisco's catalytic sites, thereby influencing carbamylation. In C3 species, Rubisco operates in a low CO2 environment, which is suboptimal for both catalysis and carbamylation. In C4 plants, Rubisco is located in the bundle sheath cells and operates in a high CO2 atmosphere close to saturation. To explore the role of Rubisco activase in C4 photosynthesis, activase levels were reduced in Flaveria bidentis, a C4 dicot, by transformation with an antisense gene directed against the mRNA for Rubisco activase. Four primary transformants with very low activase levels were recovered. These plants and several of their segregating T1 progeny required high CO2 (>1 kPa) for growth. They had very low CO2 assimilation rates at high light and ambient CO2, and only 10% to 15% of Rubisco sites were carbamylated at both ambient and very high CO2. The amount of Rubisco was similar to that of wild-type plants. Experiments with the T1 progeny of these four primary transformants showed that CO2 assimilation rate and Rubisco carbamylation were severely reduced in plants with less than 30% of wild-type levels of activase. We conclude that activase activity is essential for the operation of the C4 photosynthetic pathway.

Carbon Dioxide↗
PubMed2005

Lateral diffusion of CO2 in leaves is not sufficient to support photosynthesis.

Lateral diffusion of CO(2) was investigated in photosynthesizing leaves with different anatomy by gas exchange and chlorophyll a fluorescence imaging using grease to block stomata. When one-half of the leaf surface of the heterobaric species Helianthus annuus was covered by 4-mm-diameter patches of grease, the response of net CO(2) assimilation rate (A) to intercellular CO(2) concentration (C(i)) indicated that higher ambient CO(2) concentrations (C(a)) caused only limited lateral diffusion into the greased areas. When single 4-mm patches were applied to leaves of heterobaric Phaseolus vulgaris and homobaric Commelina communis, chlorophyll a fluorescence images showed dramatic declines in the quantum efficiency of photosystem II electron transport (measured as F(q)'/F(m)') across the patch, demonstrating that lateral CO(2) diffusion could not support A. The F(q)'/F(m)' values were used to compute images of C(i) across patches, and their dependence on C(a) was assessed. At high C(a), the patch effect was less in C. communis than P. vulgaris. A finite-volume porous-medium model for assimilation rate and lateral CO(2) diffusion was developed to analyze the patch images. The model estimated that the effective lateral CO(2) diffusion coefficients inside C. communis and P. vulgaris leaves were 22% and 12% of that for free air, respectively. We conclude that, in the light, lateral CO(2) diffusion cannot support appreciable photosynthesis over distances of more than approximately 0.3 mm in normal leaves, irrespective of the presence or absence of bundle sheath extensions, because of the CO(2) assimilation by cells along the diffusion pathway.

Calibration↗
PubMed2005

Genetic engineering of the biosynthesis of glycinebetaine enhances photosynthesis against high temperature stress in transgenic tobacco plants.

Genetically engineered tobacco (Nicotiana tabacum) with the ability to synthesis glycinebetaine was established by introducing the BADH gene for betaine aldehyde dehydrogenase from spinach (Spinacia oleracea). The genetic engineering enabled the plants to accumulate glycinebetaine mainly in chloroplasts and resulted in enhanced tolerance to high temperature stress during growth of young seedlings. Moreover, CO2 assimilation of transgenic plants was significantly more tolerant to high temperatures than that of wild-type plants. The analyses of chlorophyll fluorescence and the activation of Rubisco indicated that the enhancement of photosynthesis to high temperatures was not related to the function of photosystem II but to the Rubisco activase-mediated activation of Rubisco. Western-blotting analyses showed that high temperature stress led to the association of Rubisco activase with the thylakoid membranes from the stroma fractions. However, such an association was much more pronounced in wild-type plants than in transgenic plants. The results in this study suggest that under high temperature stress, glycinebetaine maintains the activation of Rubisco by preventing the sequestration of Rubisco activase to the thylakoid membranes from the soluble stroma fractions and thus enhances the tolerance of CO2 assimilation to high temperature stress. The results seem to suggest that engineering of the biosynthesis of glycinebetaine by transformation with the BADH gene might be an effective method for enhancing high temperature tolerance of plants.

Betaine↗
PubMed2005

Characteristics of electrical signals in poplar and responses in photosynthesis.

To gain an understanding of the role of electrical signaling in trees, poplar (Populus trichocarpa, Populus tremula x P. tremuloides) shoots were stimulated by chilling as well as flaming. Two kinds of signal propagation were detected by microelectrode measurements (aphid technique) in the phloem of leaf veins: (1) basipetal, short-distance signaling that led to rapid membrane hyperpolarization caused by K+-efflux within the leaf lamina; and (2) acropetal, long-distance signaling that triggered depolarization of the membrane potential in the leaf phloem. In the latter, the depolarizing signals travel across the stem from the manipulated leaves to adjacent leaves where the net CO2 uptake rate is temporarily depressed toward compensation. With regard to photosystem II, both heat-induced long-distance and short-distance signaling were investigated using two-dimensional "imaging" analysis of chlorophyll fluorescence. Both types of signaling significantly reduced the quantum yield of electron transport through photosystem II. Imaging analysis revealed that the signal that causes yield reduction spreads through the leaf lamina. Coldblocking of the stem proved that the electrical signal transmission via the phloem becomes disrupted, causing the leaf gas exchange to remain unaffected. Calcium-deficient trees showed a marked contrast inasmuch as the amplitude of the electrical signal was distinctly reduced, concomitant with the absence of a significant response in leaf gas exchange upon flame wounding. In summary, the above results led us to conclude that calcium as well as potassium is involved in the propagation of phloem-transmitted electrical signals that evoke specific responses in the photosynthesis of leaves.

Chlorophyll↗
PubMed2006

alpha-Tocopherol plays a role in photosynthesis and macronutrient homeostasis of the cyanobacterium Synechocystis sp. PCC 6803 that is independent of its antioxidant function.

alpha-Tocopherol is synthesized exclusively in oxygenic phototrophs and is known to function as a lipid-soluble antioxidant. Here, we report that alpha-tocopherol also has a novel function independent of its antioxidant properties in the cyanobacterium Synechocystis sp. PCC 6803. The photoautotrophic growth rates of wild type and mutants impaired in alpha-tocopherol biosynthesis are identical, but the mutants exhibit elevated photosynthetic activities and glycogen levels. When grown photomixotrophically with glucose (Glc), however, these mutants cease growth within 24 h and exhibit a global macronutrient starvation response associated with nitrogen, sulfur, and carbon, as shown by decreased phycobiliprotein content (35% of the wild-type level) and accumulation of the nblA1-nblA2, sbpA, sigB, sigE, and sigH transcripts. Photosystem II activity and carboxysome synthesis are lost in the tocopherol mutants within 24 h of photomixotrophic growth, and the abundance of carboxysome gene (rbcL, ccmK1, ccmL) and ndhF4 transcripts decreases to undetectable levels. These results suggest that alpha-tocopherol plays an important role in optimizing photosynthetic activity and macronutrient homeostasis in Synechocystis sp. PCC 6803. Several lines of evidence indicate that increased oxidative stress in the tocopherol mutants is unlikely to be the underlying cause of photosystem II inactivation and Glc-induced lethality. Interestingly, insertional inactivation of the pmgA gene, which encodes a putative serine-threonine kinase similar to RsbW and RsbT in Bacillus subtilis, results in a similar increase in glycogen and Glc-induced lethality. Based on these results, we propose that alpha-tocopherol plays a nonantioxidant regulatory role in photosynthesis and macronutrient homeostasis through a signal transduction pathway that also involves PmgA.

Antioxidants↗
PubMed2006

Impaired photosynthesis in phosphatidylglycerol-deficient mutant of cyanobacterium Anabaena sp. PCC7120 with a disrupted gene encoding a putative phosphatidylglycerophosphatase.

Phosphatidylglycerol (PG) is a ubiquitous phospholipid in thylakoid membranes of cyanobacteria and chloroplasts and plays an important role in the structure and function of photosynthetic membranes. The last step of the PG biosynthesis is dephosphorylation of phosphatidylglycerophosphate (PGP) catalyzed by PGP phosphatase. However, the gene-encoding PGP phosphatase has not been identified and cloned from cyanobacteria or higher plants. In this study, we constructed a PG-deficient mutant from cyanobacterium Anabaena sp. PCC7120 with a disrupted gene (alr1715, a gene for Alr1715 protein, GenBank accession no. BAB78081) encoding a putative PGP phosphatase. The obtained mutant showed an approximately 30% reduction in the cellular content of PG. Following the reduction in the PG content, the photoautotrophical growth of the mutant was restrained, and the cellular content of chlorophyll was decreased. The decreases in net photosynthetic and photosystem II (PSII) activities on a cell basis also occurred in this mutant. Simultaneously, the photochemical efficiency of PSII was considerably declined, and less excitation energy was transferred toward PSII. These findings demonstrate that the alr1715 gene of Anabaena sp. PCC7120 is involved in the biosynthesis of PG and essential for photosynthesis.

Amino Acid Sequence↗
PubMed2006

Transgenic tobacco plants overexpressing chloroplastic ferredoxin-NADP(H) reductase display normal rates of photosynthesis and increased tolerance to oxidative stress.

Ferredoxin-NADP(H) reductase (FNR) catalyzes the last step of photosynthetic electron transport in chloroplasts, driving electrons from reduced ferredoxin to NADP+. This reaction is rate limiting for photosynthesis under a wide range of illumination conditions, as revealed by analysis of plants transformed with an antisense version of the FNR gene. To investigate whether accumulation of this flavoprotein over wild-type levels could improve photosynthetic efficiency and growth, we generated transgenic tobacco (Nicotiana tabacum) plants expressing a pea (Pisum sativum) FNR targeted to chloroplasts. The alien product distributed between the thylakoid membranes and the chloroplast stroma. Transformants grown at 150 or 700 micromol quanta m(-2) s(-1) displayed wild-type phenotypes regardless of FNR content. Thylakoids isolated from plants with a 5-fold FNR increase over the wild type displayed only moderate stimulation (approximately 20%) in the rates of electron transport from water to NADP+. In contrast, when donors of photosystem I were used to drive NADP+ photoreduction, the activity was 3- to 4-fold higher than the wild-type controls. Plants expressing various levels of FNR (from 1- to 3.6-fold over the wild type) failed to show significant differences in CO2 assimilation rates when assayed over a range of light intensities and CO2 concentrations. Transgenic lines exhibited enhanced tolerance to photooxidative damage and redox-cycling herbicides that propagate reactive oxygen species. The results suggest that photosynthetic electron transport has several rate-limiting steps, with FNR catalyzing just one of them.

Carbon Dioxide↗
PubMed1997

Intracellular carbonic anhydrase is essential to photosynthesis in Chlamydomonas reinhardtii at atmospheric levels of CO2. Demonstration via genomic complementation of the high-CO2-requiring mutant ca-1.

Genomic complementation of the high-CO2-requiring mutant ca-1-12-1C of Chlamydomonas reinhardtii was achieved by transformation with DNA pools from an indexed cosmid library of wild-type genomic DNA. Transformation of mutant cells with cosmid DNA from two microtiter plates in the library produced colonies that grew phototrophically at atmospheric CO2 levels. Transformations with cosmid DNA from each of the rows and files of the two plates pinpointed one well in each plate with a cosmid bearing the targeted gene. Sequencing of cosmid subclones revealed a gene encoding a recently identified C. reinhardtii chloroplast carbonic anhydrase (CAH3). Transformations with chimeric constructs combining different portions of the wild-type and mutant genes indicated the presence of a mutation in the 5'-half of the gene. Comparison of mutant and wild-type gene sequences in this region revealed a G-to-A substitution in the mutant gene, which produced a nonsense codon. The data presented demonstrate that the carbonic anhydrase produced from the CAH3 gene is essential to the inorganic carbon-concentrating mechanism in C. reinhardtii and that genomic complementation can be a facile and efficient means for isolating genes associated with defects affecting photosynthesis and other physiological processes in this eukaryotic green alga.

Amino Acid Sequence↗
PubMed1997

Evidence for transcriptional regulation of plastid photosynthesis genes in Arabidopsis thaliana roots.

Mechanisms underlying suppressed levels of transcripts for plastid photosynthesis genes in nongreen tissues such as roots and calli were analyzed in Arabidopsis thaliana, a plant suitable for further genetic dissection. A region encoding promoters of rbcL, the gene encoding the large subunit of ribulose-1,5-biphosphate carboxylase/oxygenase, and the atpB/E operon for the beta and epsilon subunits of coupling factor one were cloned and sequenced. Transcripts for rbcL, atpB/E, and psbA, the gene for the D1 protein in the photosystem II reaction center, were barely detectable in roots of A. thaliana, whereas 16S rRNA was detected at a low level. The run-on transcription experiment revealed that expression of rbcL, atpB/E, and psbA was regulated at transcription. The copy number of plastid DNA in roots was one-fifth that in green leaves on the basis of total cellular DNA, suggesting that in the latter the DNA copy-number regulation also exists in plastid gene expression. Digestion of DNA with methyl-sensitive and -insensitive isoschizomeric endonucleases and subsequent polymerase chain reaction, as well as in vitro transcription of plastid DNAs with Escherichia coli RNA polymerase, resulted in no evidence of regulation by DNA modification. In spite of predominant suppression of expression of rbcL, atpB/E, and psbA at transcription in roots and calli, 16S rRNA levels were decreased because of low RNA stability.

Arabidopsis↗