Disturbance of chlorophyll biosynthesis at Mg branch affects the chloroplast ROS homeostasis and Ca<Superscript>2+</Superscript> signaling in <Emphasis Type="Italic">Pisum sativum</Emphasis>

Reactive oxygen species (ROS) and calcium (Ca²⁺), two crucial intracellular signaling molecules, have been reported to play important roles in chlorophyll biosynthesis. In this study, we aimed to investigate whether disturbance of chlorophyll synthesis affects chloroplast ROS and Ca²⁺ homeostases. Chlorophyll biosynthesis was inhibited at the Mg branch by virus-induced gene silencing (VIGS) of CHLI gene encoding the Mg chelatase CHLI subunit in pea (Pisum sativum). Subsequently, ROS and intracellular free Ca²⁺ concentration ([Ca²⁺]_i) in these chlorophyll-deficient pea plants were evaluated by histochemical and fluorescent staining assays. The results showed that the superoxide anion and hydrogen peroxide were predominantly generated in chloroplasts of the yellow leaves of pea VIGS-CHLI plants. The expression of genes encoding chloroplast antioxidant enzymes (CuZn-superoxide dismutase, ascorbate peroxidase, glutathione reductase, phospholipid glutathione peroxidase, peroxiredoxin and thioredoxins) were also decreased in the leaves of VIGS-CHLI plants compared with the control plants. Additionally, the [Ca²⁺]_i were significantly reduced in the yellow leaves of VIGS-CHLI plants compared with the green leaves of VIGS-GFP control plants. The expression of genes encoding Ca²⁺ signaling related proteins (thylakoid Ca²⁺ transporter, calmodulins and calcineurin B-like protein) was down-regulated in yellow VIGS-CHLI leaves. These results indicate that inhibition of chlorophyll biosynthesis at the Mg branch by silencing CHLI affects chloroplast ROS homeostasis and Ca²⁺ signaling and down-regulates the expression of ROS scavenging genes and Ca²⁺ signaling related genes.     

Identification and levels of 2'-carboxyarabinitol in leaves

2′-Carboxyarabinitol 1-phosphate (CA1P) is a naturally occurring inhibitor of ribulose-1,5 bisphosphate carboxylase/oxygenase activity. A chloroplast phosphatase has previously been identified that degrades CA1P in vitro to carboxyarabinitol (CA) plus phosphate, but CA has not yet been detected in plants. Here, we detail procedures to isolate and assay CA from leaves and utilize mass spectrometry to demonstrate for the first time that CA is present in plants. CA was present in leaves of all 13 species examined, including those of C₃, C₄, and Crassulacean acid metabolism photosynthetic subgroups. CA was present both in species with high levels of CA1P (e.g. Phaseolus vulgaris, Lycopersicon esculentum, Beta vulgaris) as well as in species with low levels of CA1P (e.g. Spinacea oleracea, Triticum aestivum). CA levels in the light were sometimes greater than those in the dark. Bean leaves had the most CA of any species tested, with levels in the light approaching 1 micromole per milligram of chlorophyll. In illuminated bean leaves, about 63% of the CA is located outside the chloroplast. CA is one of only a few branched chain sugar acids to be identified from plants.     

Influence of Nitrate and Ammonia on Photosynthetic Characteristics and Leaf Anatomy of Moricandia arvensis

The leaf anatomy and certain photosynthetic properties of nitrate- and ammonia-grown plants of Moricandia arvensis (L.) DC., a species previously reported to be a C₃-C₄ intermediate, were investigated. Nitrate-grown plants had a high level of malate in the leaves while ammonia-grown plants had low levels of malate. In young leaves of nitrate-grown plants, there was a diurnal fluctuation of malate content, increasing during the day and decreasing during the night. Titratable acidity remained low in leaves of both nitrate- and ammonia-grown plants.

In nitrate-grown plants, the activity of phosphoenolpyruvate (PEP) carboxylase was about 2-fold higher than in ammonia-grown plants, the latter having activity typical of C₃ species. Also, in nitrate-grown plants, the ratio of activities of ribulose 1,5-bisphosphate (RuBP) carboxylase/PEP carboxylase was lower than in ammonia-grown plants. Nitrate reductase activities were higher in nitrate- than in ammonia-grown plants and the greatest activity was found in younger leaves.

With nitrate-grown plants, during a pulse-chase experiment the label in malate, as a percentage of the total labeled products, increased from about 7% after a 10-second pulse with ¹⁴CO₂ up to 17% during a 5-minute chase with ¹²CO₂. The pattern of ¹⁴C labeling in various metabolites suggests the primary carboxylation is through RuBP carboxylase with a secondary carboxylation through PEP carboxylase. In similar experiments, with ammonia-grown plants, the percentage label in malate was only 0% to 4% with no increase in malate labeling during the chase period. The CO₂ compensation point was lower in nitrate-grown than ammonia-grown plants.

There was no evidence of Kranz-like anatomy in either the nitrate or ammonia-grown plants. Mitochondria of bundle-sheath cells were strikingly positioned along the inner tangential wall. This might allow the chloroplasts of these cells to fix the mitochondrial photorespired CO₂ more effectively and contribute to the low CO₂ compensation point in the species. Chloroplasts of bundle-sheath cells and contiguous mesophyll cells were similar in size and structure in plants grown on different media, although chloroplast thylakoids and stromata of the ammonia-grown plants stained more intensely than those of nitrate-grown plants. In addition, irregular clusters of phytoferritin particles occurred in the chloroplasts of the ammonia-grown plants.

The results indicate that the substantial activity of PEP carboxylase, incorporation of CO₂ into malate, the high malate content, and in part the relatively low CO₂ compensation point in Moricandia arvensis may be accounted for by metabolism of nitrate rather than by a state of C₃-C₄ intermediacy.

     

Different Growth and Physiological Responses to Cadmium of the Three Miscanthus Species

Miscanthus has been proposed as a promising crop for phytoremediation due to its high biomass yield and remarkable adaptability to different environments. However, little is known about the resistance of Miscanthus spp. to cadmium (Cd). To determine any differences in resistance of Miscanthus to Cd, we examined plant growth, net photosynthetic rate (Pn), activities of anti-oxidant and C₄ photosynthetic enzymes, concentrations of Cd in leaves and roots, and observed the chloroplast structure in three Miscanthus species treated with 0, 10, 50, 100 or 200 μM Cd in solutions. Miscanthus sinensis showed more sensitivity to Cd, including sharp decreases in growth, Pn, PEPC activity and damage to chloroplast structure, and the highest H₂O₂ and Cd concentrations in leaves and roots after Cd treatments. Miscanthus sacchariflorus showed higher resistance to Cd and better growth, had the highest Pn and phosphoenolpyruvate carboxylase (PEPC) activities and integrative chloroplast structure and the lowest hydrogen peroxide (H₂O₂) and leaf and root Cd concentrations. The results could play an important role in understanding the mechanisms of Cd tolerance in plants and in application of phytoremediation.     

High-Level Transient Expression of ER-Targeted Human Interleukin 6 in Nicotiana benthamiana

Tobacco plants can be used to express recombinant proteins that cannot be produced in a soluble and active form using traditional platforms such as Escherichia coli. We therefore expressed the human glycoprotein interleukin 6 (IL6) in two commercial tobacco cultivars (Nicotiana tabacum cv. Virginia and cv. Geudertheimer) as well as the model host N. benthamiana to compare different transformation strategies (stable vs. transient expression) and subcellular targeting (apoplast, endoplasmic reticulum (ER) and vacuole). In T₀ transgenic plants, the highest expression levels were achieved by ER targeting but the overall yields of IL6 were still low in the leaves (0.005% TSP in the ER, 0.0008% in the vacuole and 0.0005% in the apoplast). The apoplast variant accumulated to similar levels in leaves and seeds, whereas the ER-targeted variant was 1.2-fold more abundant in seeds and the vacuolar variant was 6-fold more abundant in seeds. The yields improved in subsequent generations, with the best-performing T₂ plants producing the ER-targeted IL6 at 0.14% TSP in both leaves and seeds. Transient expression of ER-targeted IL6 in leaves using the MagnICON system resulted in yields of up to 7% TSP in N. benthamiana, but only 1% in N. tabacum cv. Virginia and 0.5% in cv. Geudertheimer. Although the commercial tobacco cultivars produced up to threefold more biomass than N. benthamiana, this was not enough to compensate for the lower overall yields. The recombinant IL6 produced by transient and stable expression in plants was biologically active and presented as two alternative bands matching the corresponding native protein.     

Transient production of artemisinin in Nicotiana benthamiana is boosted by a specific lipid transfer protein from A. annua

Our lack of full understanding of transport and sequestration of the heterologous products currently limit metabolic engineering in plants for the production of high value terpenes. For instance, although all genes of the artemisinin/arteannuin B (AN/AB) biosynthesis pathway (AN-PW) from Artemisia annua have been identified, ectopic expression of these genes in Nicotiana benthamiana yielded mostly glycosylated pathway intermediates and only very little free (dihydro)artemisinic acid [(DH)AA]. Here we demonstrate that Lipid Transfer Protein 3 (AaLTP3) and the transporter Pleiotropic Drug Resistance 2 (AaPDR2) from A. annua enhance accumulation of (DH)AA in the apoplast of N. benthamiana leaves. Analysis of apoplast and cell content and apoplast exclusion assays show that AaLTP3 and AaPDR2 prevent reflux of (DH)AA from the apoplast back into the cells and enhances overall flux through the pathway. Moreover, AaLTP3 is stabilized in the presence of AN-PW activity and co-expression of AN-PW+AaLTP3+AaPDR2 genes yielded AN and AB in necrotic N. benthamiana leaves at 13 days post-agroinfiltration. This newly discovered function of LTPs opens up new possibilities for the engineering of biosynthesis pathways of high value terpenes in heterologous expression systems.     

Carbonic anhydrase in relation to higher plants

The review incorporates recent information on carbonic anhydrase (CA, EC: 4.2.1.1) pertaining to types, homology, regulation, purification, in vitro stability, and biological functions with special reference to higher plants. CA, a ubiquitous enzyme in prokaryotes and higher organisms represented by four distinct families, is involved in diverse biological processes, including pH regulation, CO₂ transfer, ion exchange, respiration, and photosynthetic CO₂ fixation. CA from higher plants traces its origin with prokaryotes and exhibits compartmentalization among their organs, tissues, and cellular organelles commensurate with specific functions. In leaves, CA represents 1–20 % of total soluble protein and abundance next only to ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBPCO) in chloroplast, facilitating CO₂ supply to phosphoenol pyruvate carboxylase in C₄ and CAM plants and RuBPCO in C₃ plants. It confers special significance to CA as an efficient biochemical marker for carbon sequestration and environmental amelioration in the current global warming scenario linked with elevated CO₂ concentrations.     

Trigalactosyldiacylglycerol 3 protein orthologs are required for basal disease resistance in Nicotiana benthamiana

Phosphatidic acid plays an important role in Nicotiana benthamiana immune responses against phytopathogenic bacteria. We analyzed the contributions of endoplasmic reticulum-derived chloroplast phospholipids, including phosphatidic acid, to the resistance of N. benthamiana against Ralstonia solanacearum. Here, we focused on trigalactosyldiacylglycerol 3 (TGD3) protein as a candidate required for phosphatidic acid signaling. On the basis of Arabidopsis thaliana TGD3 sequences, we identified two putative TGD3 orthologs in the N. benthamiana genome, NbTGD3-1 and NbTGD3-2. To address the role of TGD3s in plant defense responses, we created double NbTGD3-silenced plants using virus-induced gene silencing. The NbTGD3-silenced plants showed a moderately reduced growth phenotype. Bacterial growth and the appearance of bacterial wilt disease were accelerated in NbTGD3-silenced plants, compared with control plants, challenged with R. solanacearum. The NbTGD3-silenced plants showed reduced both expression of allene oxide synthase that encoded jasmonic acid biosynthetic enzyme and NbPR-4, a marker gene for jasmonic acid signaling, after inoculation with R. solanacearum. Thus, NbTGD3-mediated endoplasmic reticulum—chloroplast lipid transport might be required for jasmonic acid signaling-mediated basal disease resistance in N. benthamiana.     

Ectopic Expression of Sweet Potato Cysteine Protease SPCP3 Alters Phenotypic Traits and Enhances Drought Stress Sensitivity in Transgenic Arabidopsis Plants

In this report sweet potato cysteine protease SPCP3 cDNAs, with or without the corresponding granulin-like domain, were overexpressed in transgenic Arabidopsis plants. Transgenic Arabidopsis plants with ectopic expression of full-length SPCP3 exhibited slight promotion of earlier floral transition from vegetative to reproductive growth and a higher percentage of yellowing siliques per plant. Transgenic progeny seeds showed similar patterns of germination rates and germination curves but lower germination percentages compared to those of wild-type control seeds. During drought treatment, photochemical F _v/F _m values and relative water content of transgenic plants were significantly reduced compared to those of wild-type controls. Transgenic Arabidopsis plants with ectopic expression of sweet potato SPCP3 with or without the corresponding C-terminal granulin-like domain exhibited similar drought-stress sensitivity patterns. Drought stress also enhanced SPCP3 gene expression, photochemical F _v/F _m reduction, and wilting in sweet potato detached leaves. Based on these data, we conclude that sweet potato granulin-containing cysteine protease SPCP3 is a functional gene, and its ectopic expression alters phenotypic traits and enhances drought-stress sensitivity in transgenic Arabidopsis plants. The presence of the C-terminal granulin-like domain has no significant influence on SPCP3-mediated drought-stress sensitivity in transgenic Arabidopsis plants.     

Whole Leaf Carbon Exchange Characteristics of Phosphate Deficient Soybeans (Glycine max L.)

Low phosphate nutrition results in increased chlorophyll fluorescence, reduced photosynthetic rate, accumulation of starch and sucrose in leaves, and low crop yields. This study investigated physiological responses of soybean (Glycine max [L.] Merr.) leaves to low inorganic phosphate (Pi) conditions. Responses of photosynthesis to light and CO₂ were examined for leaves of soybean grown at high (0.50 millimolar) or low (0.05 millimolar) Pi. Leaves of low Pi plants exhibited paraheliotropic orientation on bright sunny days rather than the normal diaheliotropic orientation exhibited by leaves of high Pi soybeans. Leaves of plants grown at high Pi had significantly higher light saturation points (1000 versus 630 micromole photons [400-700 nanometers] per square meter per second) and higher apparent quantum efficiency (0.062 versus 0.044 mole CO₂ per mole photons) at ambient (34 pascals) CO₂ than did low Pi leaves, yet stomatal conductances were similar. High Pi leaves also had significantly higher carboxylation efficiency (2.90 versus 0.49 micromole CO₂ per square meter per second per pascal), a lower CO₂ compensation point (6.9 versus 11.9 pascals), and a higher photosynthetic rate at 34 pascals CO₂ (19.5 versus 6.7 micromoles CO₂ per square meter per second) than did low Pi leaves. Soluble protein (0.94 versus 0.73 milligram per square centimeter), ribulose-1,5-bisphosphate carboxylase/oxygenase content (0.33 versus 0.25 milligram per square centimeter), and ribulose-1,5-bisphosphate carboxylase/oxygenase specific activity (25.0 versus 16.7 micromoles per square meter per second) were significantly greater in leaves of plants in the high Pi treatment. The data indicate that Pi stress alters the plant's CO₂ reduction characteristics, which may in turn affect the plant's capacity to accommodate normal radiation loads.     

Biochemical studies on the iojap mutant of maize

The white leaf tissue of seedlings of Zea mays L. affected by the recessive nuclear gene iojap shows no photosynthetic activity; it contains about 1.4% of carotenoid and less than 0.1% of chlorophyll a content of normal green tissue. Neither fraction I protein nor chloroplast adenosine triphosphatase (EC 3.6.1.4) (CF₁) is detectable. This confirms earlier observations that plastids of white sectors of iojap maize do not contain ribosomes. About 40% of the activity of phosphoenolpyruvate carboxylase (EC 4.1.1.31) in green leaves could be found in white leaves indicating that the phosphoenolpyruvate carboxylase EC 4.1.1.31 is made on cytoplasmic ribosomes. The oxygen consumption of iojap-affected leaves is decreased.     

Mutations in the Capsid Protein of Brome Mosaic Virus Affecting Encapsidation Eliminate Vesicle Induction In Planta: Implications for Virus Cell-to-Cell Spread

Positive-strand RNA viruses are known to rearrange the endomembrane network to make it more conducive for replication, maturation, or egress. Our previous transmission electron microscopic (TEM) analysis showed that ectopic expression of wild-type (wt) capsid protein (CP) of Brome mosaic virus (BMV) has an intrinsic property of modifying the endoplasmic reticulum (ER) to induce vesicles similar to those present in wt BMV infection. In this study, we evaluated the functional significance of CP-mediated vesicle induction to the BMV infection cycle in planta. Consequently, the cytopathologic changes induced by wt CP or its mutants defective in virion assembly due to mutations engineered in either N- or C-proximal domains were comparatively analyzed by TEM in two susceptible (Nicotiana benthamiana and Chenopodium quinoa) and one nonhost (N. clevelandii) plant species. The results showed that in susceptible hosts, CP-mediated ER-derived vesicle induction is contingent on the expression of encapsidation-competent CP. In contrast, unlike in N. benthamiana and C. quinoa, transient expression of wt CP in nonhost N. clevelandii plants eliminated vesicle induction. Additionally, comparative source-to-sink analysis of virus spread in leaves of N. benthamiana and N. clevelandii coexpressing wt BMV and Cucumber mosaic virus (CMV) showed that despite trans-encapsidation, CMV failed to complement the defective cell-to-cell movement of BMV. The significance and relation of CP-mediated vesicle induction to virus cell-to-cell movement are discussed.     

Photosynthate metabolism in the source leaves of n(2)-fixing soybean plants

Soybean plants (Glycine max [L.] Merr. cv Williams), which were symbiotic with Bradyrhizobium japonicum, and which grew well upon reduced nitrogen supplied solely through N₂ fixation processes, often exhibited excess accumulation of starch and sucrose and diminished soluble protein in their source leaves. Nitrate and ammonia, when supplied to the nodulated roots of N₂-fixing plants, mediated a reduction of foliar starch accumulation and a corresponding increase in soluble protein in the source leaves. This provided an opportunity to examine the potential metabolic adjustments by which NO₃⁻ and NH₄⁺ (N) sufficiency or deficiency exerted an influence upon soybean leaf starch synthesis. When compared with soybean plants supplied with N, elevated starch accumulation was focused in leaf palisade parenchyma tissue of N₂-fixing plants. Foliar activities of starch synthesis pathway enzymes including fructose-1,6-bisphosphate phosphatase, phosphohexoisomerase, phosphoglucomutase (PGM), as well as adenosine diphosphate glucose pyrophosphorylase (in some leaves) exhibited highest activities in leaf extracts of N₂-fixing plants when expressed on a leaf protein basis. This was interpreted to mean that there was an adaptation of these enzyme activities in the leaves of N₂-fixing plants, and this contributed to an increase in starch accumulation. Another major causal factor associated with increased starch accumulation was the elevation in foliar levels of fructose-6-phosphate, glucose-6-phosphate, and glucose-1-phosphate (G1P), which had risen to chloroplast concentrations considerably in excess of the K_m values for their respective target enzymes associated with starch synthesis, e.g. elevated G1P with respect to adenosine diphosphate glucose pyrophosphorylase (ADPG-PPiase) binding sites. The cofactor glucose-1,6-bisphosphate (G1,6BP) was found to be obligate for maximal PGM activity in soybean leaf extracts of N₂-fixing as well as N-supplemented plants, and G1,6BP levels in N₂-fixing plant leaves was twice that of levels in N-supplied treatments. However the concentration of chloroplastic G1,6BP in illuminated leaves was computed to be saturating with respect to PGM in both N₂-fixing and N-supplemented plants. This suggested that the higher level of this cofactor in N₂-fixing plant leaves did not confer any higher PGM activation and was not a factor in higher starch synthesis rates. Relative to plants supplied with NO₃⁻ and NH₄⁺, the source leaf glycerate-3-phosphate (3-PGA) and orthophosphate (Pi) concentrations in leaves of N₂-fixing plants were two to four times higher. Although Pi is a physiological competitive inhibitor of leaf chloroplast ADPG-PPiase, and hence, starch synthesis, elevated chloroplast 3-PGA levels in N₂-fixing plant leaves apparently prevented interference of Pi with ADPG-PPiase catalysis and starch synthesis.     

Effects of light quality on the chloroplastic ultrastructure and photosynthetic characteristics of cucumber seedlings

To understand how light quality influences plant photosynthesis, we investigated chloroplastic ultrastructure, chlorophyll fluorescence and photosynthetic parameters, Rubisco and chlorophyll content and photosynthesis-related genes expression in cucumber seedlings exposed to different light qualities: white, red, blue, yellow and green lights with the same photosynthetic photon flux density of 100 μmol m^?2 s^?1. The results revealed that plant growth, CO₂ assimilation rate and chlorophyll content were significantly reduced in the seedlings grown under red, blue, yellow and green lights as compared with those grown under white light, but each monochromatic light played its special role in regulating plant morphogenesis and photosynthesis. Seedling leaves were thickened and slightly curled; Rubisco biosynthesis, expression of the rca, rbcS and rbcL, the maximal photochemical efficiency of PSII (Fv/Fm) and quantum yield of PSII electron transport (Ф_PSII) were all increased in seedlings grown under blue light as compared with those grown under white light. Furthermore, the photosynthetic rate of seedlings grown under blue light was significantly increased, and leaf number and chlorophyll content of seedlings grown under red light were increased as compared with those exposed to other monochromatic lights. On the contrary, the seedlings grown under yellow and green lights were dwarf with the new leaves etiolated. Moreover, photosynthesis, Rubisco biosynthesis and relative gene expression were greatly decreased in seedlings grown under yellow and green light, but chloroplast structural features were less influenced. Interestingly, the Fv/Fm, Ф_PSII value and chlorophyll content of the seedlings grown under green light were much higher than those grown under yellow light.     

An Examination of the Plastid DNA of Hypohaploid Nicotiana plumbaginifolia Plants

DNA was extracted from different morphological types of hypohaploid Nicotiana plumbaginifolia plants. The cellular levels of chloroplast DNA (expressed as percent of total DNA) were found to be approximately two- to threefold higher in two albino hypohaploids than in a green hypohaploid. The level of chloroplast DNA in the green hypohaploid was not significantly different from either in vitro or in vivo grown haploid N. plumbaginifolia plants. Molecular hybridization with DNA probes for the large subunit of ribulose bisphosphate carboxylase from spinach and with Pvull fragments representing the entire Nicotiana tabacum chloroplast genome revealed no gross qualitative differences in the chloroplast DNAs of hypohaploid plants. Based on these observations we have concluded that the lack of chloroplast function observed in the albino forms of hypohaploid N. plumbaginifolia plants is not due to changes in the chloroplast genome.     

A Temperature-Sensitive Chlorophyll b-Deficient Mutant of Sweetclover (Melilotus alba)

The ch4 mutant of sweetclover (Melilotus alba) has previously been demonstrated to be partially deficient in chlorophyll and to have a higher ratio of chlorophyll a to b than normal plants. We were able to substantiate these findings when plants were grown at 23°C and lower (permissive temperatures). However, when grown at 26°C (nonpermissive temperature) the plants produced small yellow leaves which exhibited one-twentieth the chlorophyll content of normal plants. Affected leaves did not increase their chlorophyll content when plants were incubated at permissive temperatures, but leaves which developed at the lower temperature contained increased amounts of chlorophyll. Similarly, only new leaves, not previously grown leaves, exhibited the yellow phenotype when the mutant plant was shifted from the permissive temperature to the nonpermissive temperature. Ribulose 1,5-bisphosphate carboxylase activity was decreased by half, relative to normal plants, in the mutant plants grown at the nonpermissive temperature, indicating that general protein synthesis was not greatly impaired and that the effect of the mutation was perhaps specific for chlorophyll content. HPLC analysis indicated that carotenoid content was not diminished to the same extent as chlorophyll and we have determined that the thylakoid protein kinase is not altered, as is the case for other chlorophyll b-deficient mutants. Experiments suggest that changes in photoperiod may be able to modulate the effect of temperature.     

Photosynthetic Plasticity in Flaveria brownii: Growth Irradiance and the Expression of C(4) Photosynthesis

Photosynthesis was examined in leaves of Flaveria brownii A. M. Powell, grown under either 14% or 100% full sunlight. In leaves of high light grown plants, the CO₂ compensation point and the inhibition of photosynthesis by 21% O₂ were significantly lower, while activities of ribulose 1,5-bisphosphate carboxylase/oxygenase and various C₄ cycle enzymes were considerably higher than those in leaves grown in low light. Both the CO₂ compensation point and the degree of O₂ inhibition of apparent photosynthesis were relatively insensitive to the light intensity used during measurements with plants from either growth conditions. Partitioning of atmospheric CO₂ between Rubisco of the C₃ pathway and phosphoenolpyruvate carboxylase of the C₄ cycle was determined by exposing leaves to ¹⁴CO₂ for 3 to 16 seconds, and extrapolating the labeling curves of initial products to zero time. Results indicated that ~94% of the CO₂ was fixed by the C₄ cycle in high light grown plants, versus ~78% in low light grown plants. Thus, growth of F. brownii in high light increased the expressed level of C₄ photosynthesis. Consistent with the carbon partitioning patterns, photosynthetic enzyme activities (on a chlorophyll basis) in protoplasts from leaves of high light grown plants showed a more C₄-like pattern of compartmentation. Pyruvate, Pi dikinase and phosphoenolpyruvate carboxylase were more enriched in the mesophyll cells, while NADP-malic enzyme and ribulose 1,5-bisphosphate carboxylase/oxygenase were relatively more abundant in the bundle sheath cells of high light than of low light grown plants. Thus, these results indicate that F. brownii has plasticity in its utilization of different pathways of carbon assimilation, depending on the light conditions during growth.     

Photosynthesis in submersed macrophytes of a temperate lake

The photosynthetic carbon fixation pathways and levels of carbon-fixing enzymes of four dominant submersed macrophytes of Lawrence Lake, southern Michigan, were investigated during the main growth season (May to November). All four species (Scirpus subterminalis Torr., Najas flexilis (Willd.) Rostk. and Schmidt, Potamogeton praelongus Wulf., and Myriophyllum heterophyllum Michx.) were C₃ plants based on their patterns of ¹⁴C pulse-chase incorporation. High levels of phosphoenolpyruvate carboxylase were also found in these species. These levels, as well as the ribulose 1,5-biphosphate carboxylase/phosphoenolpyruvate carboxylase ratio of the leaves, varied throughout the growing season and exhibited highest values in July. No shift in carbon fixation pathways, however, could be detected from July to October. The possible functions of phosphoenolypyruvate carboxylase in these plants, as well as the significance of C₃ metabolism in submersed plants of temperate lakes, are delineated.