Effects of irradiance and temperature on photosynthesis in C3, C4 and C3/C4 Panicum species

Species in the Laxa and Grandia groups of the genus Panicum are adapted to low, wet areas of tropical and subtropical America. Panicum milioides is a species with C₃ photosynthesis and low apparent photorespiration and has been classified as a C₃/C₄ intermediate. Other species in the Laxa group are C₃ with normal photorespiration. Panicum prionitis is a C₄ species in the Grandia group. Since P. milioides has some leaf characteristics intermediate to C₃ and C₄ species, its photosynthetic response to irradiance and temperature was compared to the closely related C₃ species, P. laxum and P. boliviense and to P. prionitis. The response of apparent photosynthesis to irradiance and temperature was similar to that of P. laxum and P. boliviense, with saturation at a photosynthetic photo flux density of about 1 mmol m^-2 s^-1 at 30°C and temperature optimum near 30°C. In contrast, P. prionitis showed no light saturation up to 2 mmol m^-2 s^-1 and an optimum temperature near 40°C. P. milioides exhibited low CO₂ loss into CO₂-free air in the light and this loss was nearly insensitive to temperature. Loss of CO₂ in the light in the C₃ species, P. laxum and P. boliviense, was several-fold higher than in P. milioides and increased 2- to 5-fold with increases in temperature from 10 to 40°C. The level of dark respiration and its response to temperature were similar in all four Panicum species examined. It is concluded that the low apparent photorespiration in P. milioides does not influence its response of apparent photosynthesis to irradiance and temperature in comparison to closely related C₃ Panicum species.Abbreviations AP apparent photosynthesis - I CO₂ compensation point - gl leaf conductance; gm, mesophyll conductance - PPFD photosynthetic photon flux density - PR apparent photorespiration rate - RuBPC sibulose bisphosphate carboxylase     

Carbon-isotope discrimination by leaves of Flaveria species exhibiting different amounts of C3-and C4-cycle co-function

Carbon-isotope ratios were examined as ¹³C values in several C₃, C₄, and C₃–C₄ Flaveria species, and compared to predicted ¹³C, values generated from theoretical models. The measured ¹³C values were within 4 of those predicted from the models. The models were used to identify factors that contribute to C₃-like ¹³C values in C₃–C₄ species that exhibit considerable C₄-cycle activity. Two of the factors contributing to C₃-like ¹³C values are high CO₂ leakiness from the C₄ pathway and pi/pa values that were higher than C₄ congeners. A marked break occurred in the relationship between the percentage of atmospheric CO₂ assimilated through the C₄ cycle and the ¹³C value. Below 50% C₄-cycle assimialtion there was no significant relationship between the variables, but above 50% the ¹³C values became less negative. These results demonstrate that the level of C₄-cycle expression can increase from, 0 to 50% with little integration of carbon transfer from the C₄ to the C₃ cycle. As expression increaces above 50%, however, increased integration of C₃- and C₄-cycle co-function occurs.Abbreviations and symbols RuBP carboxylase ribulose-1,5-bisphosphate carboxylase (EC 4.1.1.39) - PEP carboxylase phosphoenolpyruvate carboxylase (EC 4.1.1.31) - pa atmospheric CO₂ partial pressure - pi intercellular CO₂ partial pressure - isotope ratio - quantum yield for CO₂ uptake     

Co-function of C3-and C4-photosynthetic pathways in C3, C4 and C3-C4 intermediate Flaveria species

The potential for C₄ photosynthesis was investigated in five C₃-C₄ intermediate species, one C₃ species, and one C₄ species in the genus Flaveria, using ¹⁴CO₂ pulse-¹²CO₂ chase techniques and quantum-yield measurements. All five intermediate species were capable of incorporating ¹⁴CO₂ into the C₄ acids malate and aspartate, following an 8-s pulse. The proportion of ¹⁴C label in these C₄ products ranged from 50–55% to 20–26% in the C₃-C₄ intermediates F. floridana Johnston and F. linearis Lag. respectively. All of the intermediate species incorporated as much, or more, ¹⁴CO₂ into aspartate as into malate. Generally, about 5–15% of the initial label in these species appeared as other organic acids. There was variation in the capacity for C₄ photosynthesis among the intermediate species based on the apparent rate of conversion of ¹⁴C label from the C₄ cycle to the C₃ cycle. In intermediate species such as F. pubescens Rydb., F. ramosissima Klatt., and F. floridana we observed a substantial decrease in label of C₄-cycle products and an increase in percentage label in C₃-cycle products during chase periods with ¹²CO₂, although the rate of change was slower than in the C₄ species, F. palmeri. In these C₃-C₄ intermediates both sucrose and fumarate were predominant products after a 20-min chase period. In the C₃-C₄ intermediates, F. anomala Robinson and f. linearis we observed no significant decrease in the label of C₄-cycle products during a 3-min chase period and a slow turnover during a 20-min chase, indicating a lower level of functional integration between the C₄ and C₃ cycles in these species, relative to the other intermediates. Although F. cronquistii Powell was previously identified as a C₃ species, 7–18% of the initial label was in malate+aspartate. However, only 40–50% of this label was in the C-4 position, indicating C₄-acid formation as secondary products of photosynthesis in F. cronquistii. In 21% O₂, the absorbed quantum yields for CO₂ uptake (in mol CO₂·[mol quanta]^-1) averaged 0.053 in F. cronquistii (C₃), 0.051 in F. trinervia (Spreng.) Mohr (C₄), 0.052 in F. ramosissima (C₃-C₄), 0.051 in F. anomala (C₃-C₄), 0.050 in F. linearis (C₃-C₄), 0.046 in F. floridana (C₃-C₄), and 0.044 in F. pubescens (C₃-C₄). In 2% O₂ an enhancement of the quantum yield was observed in all of the C₃-C₄ intermediate species, ranging from 21% in F. ramosissima to 43% in F. pubescens. In all intermediates the quantum yields in 2% O₂ were intermediate in value to the C₃ and C₄ species, indicating a co-function of the C₃ and C₄ cycles in CO₂ assimilation. The low quantum-yield values for F. pubescens and F. floridana in 21% O₂ presumably reflect an ineffcient transfer of carbon from the C₄ to the C₃ cycle. The response of the quantum yield to four increasing O₂ concentrations (2–35%) showed lower levels of O₂ inhibition in the C₃-C₄ intermediate F. ramosissima, relative to the C₃ species. This indicates that the co-function of the C₃ and C₄ cycles in this intermediate species leads to an increased CO₂ concentration at the site of ribulose-1,5-bisphosphate carboxylase/oxygenase and a concomitant decrease in the competitive inhibition by O₂.Abbreviations PEP phosphoenolpyruvate - PGA 3-phosphoglycerate - RuBP ribulose-1,5-bisphosphate     

Molecular properties of phosphoenolpyruvate carboxylase from C3, C3−C4 intermediate,and C4 Flaveria species

Phosphoenolpyruvate carboxylase (PEPCase; EC 4.1.1.31) from Flaveria trinervia Mohr (C₄), F. floridana Johnston (C₃–C₄), and F. cronquistii Powell (C₃) leaves were compared by electrotransfer blotting/enzyme-linked immunoassay (Western-blot analysis), mobility of the native enzyme in polyacrylamide gels and in isoelectric focusing (IEF) gels, peptide mapping, and in-vitro translation of RNA isolated from each plant. The PEPCases from the C₃ and C₃–C₄ plants were very similar to each other in terms of electrophoretic mobilities on gels and isoenzyme patterns on IEF gels, and identical in peptide mapping. Quantitative differences were noted, however, in that the C₃–C₄ intermediate plant contained more PEPCase overall and that the relative activity of individual isoenzymes shifted between the C₃ and C₃–C₄ intermediate PEPCases. The PEPCase from the C₄ plant had a different isoenzyme pattern, a different peptide map, and was far more abundant than the other two enzymes. Western blot analysis demonstrated the cross-reactivity of PEPCases from all three Flaveria species with antibody raised against maize PEPCase. The results provide evidence, at the molecular level, that supports the view of C₃–C₄ intermediate species as C₃-like plants with some C₄-like photosynthetic characteristics, but there are differences from the C₃ plant in the quantity and properties of the PEPCase from the C₃–C₄ intermediate plant.Abbreviations IEF isoelectric focusing - kDa kilodalton - PEPCase phosphoenolpyruvate carboxylase - Rubisco Ribulose-1,5-bisphosphate carboxylase/oxygenase     

Mechanism of the gas phase reactions of the C5H5Ni and C5H5Co ions with substituted pyridines. A combined experimental and theoretical study

Different products have been observed in the reactions of C₅H₅Co⁺ and C₅H₅Ni⁺ ions with halogen-substituted pyridines (XPy) that have been studied by ion trap mass spectrometry (ITMS) techniques. In particular, an addition product C₅H₅M(XPy)⁺ and a product ion C₅H₄M(Py)⁺ corresponding to a loss of a HX molecule (X = F, Cl, Br) have been detected. The relative yield of these products is determined by the nature of the metal and by the nature and position of the halogen on the pyridine ring. A computational study at the DFT level on model-systems formed by 2-fluoro and 2-bromopyridine reacting either with the C₅H₅Ni⁺ or the C₅H₅Co⁺ ion has been carried out. This study shows the existence of a general mechanistic pattern. The rate-determining step of this mechanism is the migration of the halogen from the pyridine ring to the metal. A final hydrogen abstraction step carried out by the halogen leads to the expulsion of a HX molecule. The existence of avoided crossings between surfaces of different multiplicities (ground and first excited state) allows the system to follow lower energy reaction pathways. The barrier determined for the reactions involving 2-bromopyridine is significantly lower than that found for 2-fluoropyridine. This is mainly due to the poor migrating/leaving character and low polarizability of fluorine compared to that of bromine.     

Performance of a generalist grasshopper on a C<Subscript>3</Subscript> and a C<Subscript>4</Subscript> grass: compensation for the effects of elevated CO<Subscript>2</Subscript> on plant nutritional quality

The increasing CO₂ concentration in Earths atmosphere is expected to cause a greater decline in the nutritional quality of C₃ than C₄ plants. As a compensatory response, herbivorous insects may increase their feeding disproportionately on C₃ plants. These hypotheses were tested by growing the grasses Lolium multiflorum C₃) and Bouteloua curtipendula C₄) at ambient (370 ppm) and elevated (740 ppm) CO₂ levels in open top chambers in the field, and comparing the growth and digestive efficiencies of the generalist grasshopper Melanoplus sanguinipes on each of the four plant × CO₂ treatment combinations. As expected, the nutritional quality of the C₃ grass declined to a greater extent than did that of the C₄ grass at elevated CO₂; protein levels declined in the C₃ grass, while levels of carbohydrates (sugar, fructan and starch) increased. However, M. sanguinipes did not significantly increase its consumption rate to compensate for the lower nutritional quality of the C₃ grass grown under elevated CO₂. Instead, these grasshoppers appear to use post-ingestive mechanisms to maintain their growth rates on the C₃ grass under elevated CO₂. Consumption rates of the C₃ and C₄ grasses were also similar, demonstrating a lack of compensatory feeding on the C₄ grass. We also examined the relative efficiencies of nutrient utilization from a C₃ and C₄ grass by M. sanguinipes to test the basis for the C₄ plant avoidance hypothesis. Contrary to this hypothesis, neither protein nor sugar was digested with a lower efficiency from the C₄ grass than from the C₃ grass. A novel finding of this study is that fructan, a potentially large carbohydrate source in C₃ grasses, is utilized by grasshoppers. Based on the higher nutrient levels in the C₃ grass and the better growth performance of M. sanguinipes on this grass at both CO₂ levels, we conclude that C₃ grasses are likely to remain better host plants than C₄ grasses in future CO₂ conditions.     

Incorporation of 14CO2 into C4 acids by leaves of C3-C4 intermediate and C3 species of Moricandia and Panicum at the CO2 compensation concentration

Comparative ¹⁴CO₂ pulse-¹²CO₂ chase studies performed at CO₂ compensation ()-versus air-concentrations of CO₂ demonstrated a four-to eightfold increase in assimilation of ¹⁴CO₂ into the C₄ acids malate and aspartate by leaves of the C₃-C₄ intermediate species Panicum milioides Nees ex Trin., P. decipiens Nees ex Trin., Moricandia arvensis (L.) DC., and M. spinosa Pomel at . Specifically, the distribution of ¹⁴C in malate and aspartate following a 10-s pulse with ¹⁴CO₂ increases from 2% to 17% (P. milioides) and 4% to 16% (M. arvensis) when leaves are illuminated at the CO₂ compensation concentration (20 l CO₂/l, 21% O₂) versus air (340 l CO₂/l, 21% O₂). Chasing recently incorporated ¹⁴C for up to 5 min with ¹²CO₂ failed to show any substantial turnover of label in the C₄ acids or in carbon-4 of malate. The C₄-acid labeling patterns of leaves of the closely related C₃ species, P. laxum Sw. and M. moricandioides (Boiss.) Heywood, were found to be relatively unresponsive to changes in pCO₂ from air to . These data demonstrate that the C₃-C₄ intermediate species of Panicum and Moricandia possess an inherently greater capacity for CO₂ assimilation via phosphoenolpyruvate (PEP) carboxylase (EC 4.1.1.31) at the CO₂ compensation concentration than closely related C₃ species. However, even at , CO₂ fixation by PEP carboxylase is minor compared to that via ribulosebisphosphate carboxylase (EC 4.1.1.39) and the C₃ cycle, and it is, therefore, unlikely to contribute in a major way to the mechanism(s) facilitating reduced photorespiration in the C₃-C₄ intermediate species of Panicum and Moricandia.Abbreviations Rubisco ribulose-1,5-bisphosphate carboxylase/oxygenase - PEP phosphoenolpyruvate - CO₂ compensation concentration - 3PGA 3-phosphoglycerate - SuP sugar monophosphates - SuP₂ sugar bisphosphates Published as Paper No. 8249, Journal Series, Nebraska Agricultural Research Division     

Interspecific variation in assimilation of 14CO2 into C4 acids by leaves of C3, C4 and C3−C4 intermediate Flaveria species near the CO2 compensation concentration

The assimilation of ¹⁴CO₂ into the C₄ acids malate and aspartate by leaves of C₃, C₄ and C₃–C₄ intermediate Flaveria species was investigated near the CO₂ compensation concentration ^* in order to determine the potential role of phosphoenolpyruvate (PEP) carboxylase (EC 4.1.1.31) in reducing photorespiration in the intermediates. Relative to air concentrations of CO₂, the proportion of CO₂ fixed by PEP carboxylase at ^* increased in all six C₃–C₄ intermediate species examined. However, F. floridana J.R. Johnston and F. ramosissima Klatt were shown to be markedly less responsive to reduced external CO₂, with only about a 1.6-fold enhancement of CO₂ assimilation by PEP carboxylase, as compared to a 3.0- to 3.7-fold increase for the other C₃–C₄ species examined, namely, F. linearis Lag., F. anomala B.L. Robinson, F. chloraefolia A. Gray and F. pubescens Rydb. The C₃ species F. pringlei Gandoger and F. cronquistii A.M. Powell exhibited a 1.5- and 2.9-fold increase in labeled malate and aspartate, respectively, at ^*. Assimilation of CO₂ by PEP carboxylase in the C₄ species F. trinervia (Spreng.) C. Mohr, F. australasica Hook., and the C₄-like species F. brownii A.M. Powell was relatively insensitive to subatmospheric levels of CO₂. The interspecific variation among the intermediate Flaverias may signify that F. floridana and F. ramosissima possess a more C₄-like compartmentation of PEP carboxylase and ribulose-1,5-bisphosphate carboxylase/oxygenase (EC 4.1.1.39) between the mesophyll and bundle-sheath cells. Chasing recently labeled malate and aspartate with ¹²CO₂ for 5 min at ^* resulted in an apparent turnover of 25% and 30% of the radiocarbon in these C₄ acids for F. ramosissima and F. floridana, respectively. No substantial turnover was detected for F. linearis, F. anomala, F. chloraefolia or F. pubescens. With the exception of F. floridana and F. ramosissima, it is unlikely that enhanced CO₂ fixation by PEP carboxylase at the CO₂ compensation concentration is a major mechanism for reducing photorespiration in the intermediate Flaveria species. Moreover, these findings support previous related ¹⁴CO₂-labeling studies at air-levels of CO₂ which indicated that F. floridana and F. ramosissima were more C₄-like intermediate species. This is further substantiated by the demonstration that F. floridana PEP carboxylase, like the enzyme in C₄ plants, undergoes a substantial activation (2.2-fold) upon illuminating dark-adapted green leaves. In contrast, light activation was not observed for the enzyme in F. linearis or F. chloraefolia.Abbreviations and symbols PEP phosphoenolpyruvate - Rubisco ribulose-1,5-bisphosphate carboxylase/oxygenase - CO₂ compensation concentration - ^* a subatmospheric level of CO₂ approximating Published as Paper No. 8832, Journal Series, Nebraska Agricultural Research Division     

Purification and properties of glycolate oxidase from plants with different photosynthetic pathways: Distinctness of C4 enzyme from that of a C3 species and a C3–C4 intermediate

Glycolate oxidase (GO; EC 1.1.3.1) was purified from the leaves of three plant species:Amaranthus hypochondriacus L.(NAD-ME type C₄ dicot),Pisum sativum L. (C₃ species) andParthenium hysterophorus L. (C₃–C₄. intermediate). A flavin moiety was present in the enzyme from all the three species. The enzyme from the C₄ plant had a low specific activity, exhibited lower K_M for glycolate, and required a lower pH for maximal activity, compared to the C₃ enzyme. The enzyme from the C₄ species oxidized glyoxylate at <10% of the rate with glycolate, while the GO from the C₃ plant oxidized glyoxylate at a rate of about 35 to 40% of that with glycolate. The sensitivity of GO from C₄ plant to -hydroxypyridinemethane sulfonate, 2-hydroxy-3-butynoate and other inhibitors was less than that of the enzyme from C₃ source. The properties of GO fromParthenium hysterophorus, were similar to those of the enzyme fromPisum sativum. The characteristics of glycolate oxidase from leaves of a C₄ plant,Amaranthus hypochondriacus are different from those of the C₃ species or the C₃–C₄ intermediate.     

Metabolic regulation of carbon flux during C4 photosynthesis

The potential for glycolate and glycine metabolism and the mechanism of refixation of photorespiratory CO₂ in leaves of C₄ plants were studied by parallel inhibitor experiments with thin leaf slices, different leaf cell types and isolated mitochondria of C₃ and C₄ Panicum species. CO₂ evolution by leaf slices of P. bisulcatum, a C₃ species, fed glycolate or glycine was light-independent and O₂-sensitive. The C₄ P. maximum and P. miliaceum leaf slices fed glycolate or glycine evolved CO₂ in the dark but not in the light. In C₄ species, dark CO₂ evolution was abolished by the addition of phosphoenolpyruvate (PEP)⁴. The addition of maleate, a PEP carboxylase inhibitor, resulted in photorespiratory CO₂ efflux by C₄ leaf slices in the light also. However, PEP and maleate had no effect on either glycolate-dependent O₂ uptake by the C₄ leaf slices or on glycolate and glycine metabolism in C₃ leaf slices. The rate of photorespiratory CO₂ evolution in the C₃ Panicum species was 3 times higher than that observed with the C₄ species. The ratio of glycolate-dependent CO₂ evolution to O₂ uptake in both groups was 1:2. Isolated C₄ mesophyll protoplasts or their mitochondria did not metabolize glycolate or glycine. However, both C₃ mesophyll protoplasts and C₄ bundle sheath strands readily metabolized glycolate and glycine in a light-independent, O₂-sensitive manner, and the addition of PEP or maleate had no effect. C₄ bundle sheath- and C₃-mitochondria were capable of oxidizing glycine. This oxidation was linked to the mitochondrial electron transport chain, was coupled to three phosphorylation sites and was sensitive to electron transport inhibitors. C₄ bundle sheath- and C₃-mitochondrial glycine decarboxylation was stimulated by oxaloacetate and NAD had no effect. In marked contrast, mitochondria isolated from C₄ mesophyll cells were incapable of oxidizing or decarboxylating added glycine. The results suggest that in leaves of C₄ plants bundle sheath cells are the primary site of O₂-sensitive photorespiratory CO₂ evolution and the PEP carboxylase present in the mesophyll cells has the Potential for efficiently refixing CO₂ before it escapes out of the leaf. The relative role of the PEP carboxylase mediated CO₂ pump and reassimilation of photorespiratory CO₂ are discussed in relation to the apparent lack of photorespiration in leaves of C₄ species.Abbreviations BSA bovine serum albumin - Chl chlorophyll - PEP phosphoenolpyruvate - Rbu-P ₂ ribulose 1,5-bisphosphate - Rib-5-P ribose-5-phosphate - Ru-5-P ribuluse-5-phosphate - FCCP carbonyl cyanide p-trifluoromethoxyphenylhydrazone Journal Series Paper, New Jersey Agricultural Experiment Station     

Intraspecific variation for CO2 compensation point and differential growth among variants in a C3−C4 intermediate plant

CO₂ compenstation point (), the concentration of CO₂ at which photosynthesis and respiration are at equilibrium, is a commonly used diagnostic for the C₄ photosynthetic pathway, since it reflects the reduced photorespiration that is a property of C₄ photosynthesis. Geographic variation for was examined within Flaveria linearis, a C₃–C₄ intermediate species. Collections from four widely separated Floridian populations were propagated in a greenhouse and measured for . Little differentiation among populations was found, but significant within-population variation was present. Temperature is a hypothesized selective agent for the C₄ photosynthetic pathway. To test this hypothesis, plants exhibiting a range of were cloned and placed in growth chambers at 25°C and 40°C. After 7 weeks, valves were remeasured and plants were harvested and weighed. There was a poor correlation between initial and final measures of for a given genotype (r=0.38, P>0.1). Broad sense heritability for was computed to be 0.10. At 25°C, there was no relationship between final size and . At 40°C, more C₄-like plants, as indicated by their low , had grown larger. Differences in relative growth rate were attributable more to differences in net assimilation rate than in leaf area ratio. Taken together, these results demonstrate that although significant plasticity exists in the amount of photorespiration in this C₃–C₄ species, high temperature appears to be an effective selective agent for the reduction of photorespiration and the enhancement of C₄-like traits.     

The relative contributions of reduced photorespiration,and improved water-and nitrogen-use efficiencies,to the advantages of C3−C4 intermediate photosynthesis in Flaveria

Summary Analyses of carbon-assimilation patterns in response to intercellular CO₂ concentrations, and the photosynthetic water-and nitrogen-use efficiencies, were conducted for a C₃, a C₄, and three C₃–C₄ species in the genus Flaveria in order to determine some of the advantages and disadvantages of C₃–C₄ intermediate photosynthesis. Operational intercellular CO₂ partial pressures (pi), determined when the atmospheric CO₂ partial pressure (pa) was approximately 330 bar, in the C₃–C₄ species were generally equal to, or greater than, those observed in the C₃ species under well-watered or water-stressed conditions. This reflects equal, or lower, water-use efficiencies (WUEs) in the C₃–C₄ species. The only case in which higher WUEs were observed in the C₃–C₄ species, compared to the C₃ species, was when photosynthesis rates were limited by available nitrogen and were less than 12.5 mol CO₂ m^-2s^-1. At higher photosynthesis rates, the C₃–C₄ species exhibited lower values of photosynthesis rate for equal values of stomatal conductance (lower WUE), compared to the C₃ species. Comparing slopes for the linear regions of the relationship between leaf nitrogen content and net photosynthesis rate (taken as an index of photosynthetic nitrogen-use efficiency, NUE), the C₄ species exhibited the highest NUE, followed by the C₃–C₄ species, F. ramosissima, with the other two C₃–C₄ species and the C₃ species being equal and exhibiting the lowest NUEs. The lack of consistent advantages in NUE and WUE in the C₃–C₄ species F. pubescens and F. floridana suggest that in some C₃–C₄ Flaveria species C₄-like anatomy and biochemistry do not provide the same gas exchange advantages that we typically attribute to the CO₂-concentrating mechanism of fully-expressed C₄ plants.     

Occurrence of the suberized lamella in leaves of grasses of different photosynthetic types. I. In parenchymatous bundle sheaths and PCR (“Kranz”) sheaths

Summary Leaf blades of 42 grasses (Poaceae) have been examined ultrastructurally for the occurrence of a suberized lamella in walls of parenchymatous bundle sheaths and PCR (= Kranz) sheaths in both large and small vascular bundles. The sample includes species from a range of major grass taxa, and represents all photosynthetic types found in the grasses. Three grasses with unusual C₄ leaf anatomy were also included:Alloteropsis semialata, Aristida biglandulosa, Arundinella nepalensis. The presence of a suberized lamella in PCR cell walls was perfectly correlated with photosynthetic type. All PEP-carboxykinase type and NADP-malic enzyme type C₄ species examined possessed a suberized lamella in outer tangential and radial walls, but with variable presence in inner tangential walls. PCR cells of bothAlloteropsis semialata andArundinella nepalensis also possessed a suberized lamella. A lamella was totally absent from parenchymatous bundle sheath cells of the C₃ species examined (5 spp.) and ofPanicum milioides, a C₃-C₄ intermediate. It was also absent from PCR cells of NAD-malic enzyme type C₄ species (14 spp.) andAristida biglandulosa. The results are discussed in relation to the leakage of CO₂ from PCR cells, and to differences between C₄ types in ¹³C values, chloroplast position in PCR cells, and other anatomical characteristics.     

Glycine decarboxylase is confined to the bundle-sheath cells of leaves of C3−C4 intermediate species

Immunogold labelling has been used to determine the cellular distribution of glycine decarboxylase in leaves of C₃, C₃–C₄ intermediate and C₄ species in the genera Moricandia, Panicum, Flaveria and Mollugo. In the C₃ species Moricandia foleyi and Panicum laxum, glycine decarboxylase was present in the mitochondria of both mesophyll and bundle-sheath cells. However, in all the C₃–C₄ intermediate (M. arvensis var. garamatum, M. nitens, M. sinaica, M. spinosa, M. suffruticosa, P. milioides, Flaveria floridana, F. linearis, Mollugo verticillata) and C₄ (P. prionitis, F. trinervia) species studied glycine decarboxylase was present in the mitochondria of only the bundle-sheath cells. The bundle-sheath cells of all the C₃–C₄ intermediate species have on their centripetal faces numerous mitochondria which are larger in profile area than those in mesophyll cells and are in close association with chloroplasts and peroxisomes. Confinement of glycine decarboxylase to the bundle-sheath cells is likely to improve the potential for recapture of photorespired CO₂ via the Calvin cycle and could account for the low rate of photorespiration in all C₃–C₄ intermediate species.Abbreviation and symbol kDa kilodaltons - CO₂ compensation point     

Reassimilation of carbon dioxide by Flaveria (Asteraceae) species representing different types of photosynthesis

The capability to reassimilate CO₂ originating from intracellular decarboxylating processes connected with the photorespiratory glycolate pathway and-or decarboxylation of C₄ acids during C₄ photosynthesis has been investigated with four species of the genus Flaveria (Asteraceae). The C₃-C₄ intermediate species F. pubescens and F. anomala reassimilated CO₂ much more efficiently than the C₃ species F. cronquistii and, with respect to this feature, behaved similarly to the C₄ species F. trinervia. Therefore, under atmospheric conditions the intermediate species photorespired with rates only between 10–20% of that measured with F. cronquistii. At low oxygen concentrations (1,5%) the reassimilation potential of F. anomala approached that of F. trinervia and was distinct from that found with F. pubescens. The data are discussed with respect to a possible sequence of events during evolution of C₄ photosynthesis. If compared with related data for C₃-C₄ intermediate species from other genera they support the hypothesis that, during evolution of C₄ photosynthesis, an efficient capacity for CO₂ reassimilation evolved prior to a CO₂-concentrating mechanism.Abbreviations C₃, C₄ assimilated CO₂ initially found in 3-phosphoglycerate (C₃) or malate and aspartate (C₄) - D reassimilation coefficient - R _n , R _t net, total CO₂ evolution as measured with 0.03 and 3% CO₂, respectively - RuBP ribulose-1,5-bisphosphate - TPS true photosynthesis     

Short-term carbon-isotope discrimination in C3−C4 intermediate species

Short-term discrimination in assimilation of stable isotopes of carbon was measured for leaves of the C₃ speciesPhaseolus vulgaris L. cv. Hawkesbury Wonder andFlaveria pringlei Gandoger, the C₄ speciesAmaranthus edulis Speg., and the C₃–C₄ intermediate speciesPanicum milioides Nees ex. Trin,Flaveria floridana Johnson, andFlaveria anomala B.L. Robinson. Discriminations in the C₃ and C₄ species were similar to those expected from theoretical considerations. When ambient CO₂ pressure was 330 bar the mean discriminations in the C₃ species andPanicum milioides were similar, whereas the mean discriminations inF. floridana andF. anomala were less than discrimination in C₃ species andPanicum milioides. When ambient CO₂ pressure was 100 bar the mean discriminations inPanicum milioides andF. anomala were greater, and that inF. floridana was less, than that inPhaseolus vulgaris. We conclude that the pattern of discrimination inPanicum milioides is consistent with the presence of a glycine shuttle; inF. floridana andF. anomala, discrimination is consistent with the presence of a C₄ pathway coupled with the operation of a glycine shuttle.Abbreviations and symbols PEP phosphoenolpyruvate - Rubisco ribulose, 1,5-bisphosphate carboxylase-oxygenase (EC 4.1.1.39) - p _a ambient CO₂ pressure - p _i intercellular CO₂ pressure - carbon-isotope discrimination - carbonisotope composition relative to PeeDee Belemnite     

The use of carbon isotope ratios to evaluate legume contribution to soil enhancement in tropical pastures

Soil carbon distribution with depth, stable carbon isotope ratios in soil organic matter and their changes as a consequence of the presence of legume were studied in three 12-year-old tropical pastures (grass alone —Brachiaria decumbens (C₄), legume alone —Pueraria phaseoloides (C₃) and grass + legume) on an Oxisol in Colombia. The objective of this study was to determine the changes that occurred in the¹³C isotope composition of soil from a grass + legume pasture that was established by cultivation of a native savanna dominated by C₄ vegetation. The¹³C natural abundance technique was used to estimate the amount of soil organic carbon originating from the legume. Up to 29% of the organic carbon in soil of the grass + legume pasture was estimated to be derived from legume residues in the top 0–2-cm soil depth, which decreased to 7% at 8–10 cm depth. Improvements in soil fertility resulting from the soil organic carbon originated from legume residues were measured as increased potential rates of nitrogen mineralization and increased yields of rice in a subsequent crop after the grass + legume pasture compared with the grass-only pasture. We conclude that the¹³C natural abundance technique may help to predict the improvements in soil quality in terms of fertility resulting from the presence of a forage legume (C₃) in a predominantly C₄ grass pasture.     

Relative nutritional quality of C3 and C4 grasses for a graminivorous lepidopteran,Paratrytone melane (Hesperiidae)

Summary We tested the hypothesis that C₄ grasses are inferior to C₃ grasses as host plants for herbivorous insects by measuring the relative performance of larvae of a graminivorous lepidopteran, Paratrytone melane (Hesperiidae), fed C₃ and C₄ grasses. Relative growth rates and final weights were higher in larvae fed a C₃ grass in Experiment I. However, in two additional experiments, relative growth rates and final weights were not significantly different in larvae fed C₃ and C₄ grasses. We examined two factors which are believed to cause C₄ grasses to be of lower nutritional value than C₃ grasses: foliar nutrient levels and nutrient digestibility. In general, foliar nutrient levels were higher in C₃ grasses. In Experiment I, protein and soluble carbohydrates were digested from a C₃ and a C₄ grass with equivalent efficiencies. Therefore, differences in larval performance are best explained by higher nutrient levels in the C₃ grass in this experiment. In Experiment II, soluble carbohydrates were digested with similar efficiencies from C₃ and C₄ grasses but protein was digested with greater efficiency from the C₃ grasses. We conclude (1) that the bundle sheath anatomy of C₄ grasses is not a barrier to soluble carbohydrate digestion and does not have a nutritionally significant effect on protein digestion and (2) that P. melane may consume C₄ grasses at compensatory rates.     

The roles of malate and aspartate in C4 photosynthetic metabolism of Flaveria bidentis (L.)

In C₄ grasses belonging to the NADP-malic enzyme-type subgroup, malate is considered to be the predominant C₄ acid metabolized during C₄ photosynthesis, and the bundle sheath cell chloroplasts contain very little photosystem-II (PSII) activity. The present studies showed that Flaveria bidentis (L.), an NADP-malic enzyme-type C₄ dicotyledon, had substantial PSII activity in bundle sheath cells and that malate and aspartate apparently contributed about equally to the transfer of CO₂ to bundle sheath cells. Preparations of bundle sheath cells and chloroplasts isolated from these cells evolved O₂ at rates between 1.5 and 2 mol · min^–1 · mg^–1 chlorophyll (Chl) in the light in response to adding either 3-phosphoglycerate plus HCO ₃ ^– or aspartate plus 2-oxoglutarate. Rates of more than 2 mol O₂ · min^–1 · mg^–1 Chl were recorded for cells provided with both sets of these substrates. With bundle sheath cell preparations the maximum rates of light-dependent CO₂ fixation and malate decarboxylation to pyruvate recorded were about 1.7 mol · min^–1 · mg^–1 Chl. Compared with NADP-malic enzyme-type grass species, F. bidentis bundle sheath cells contained much higher activities of NADP-malate dehydrogenase and of aspartate and alanine aminotransferases. Time-course and pulse-chase studies following the kinetics of radiolabelling of the C-4 carboxyl of C₄ acids from ¹⁴CO₂ indicated that the photosynthetically active pool of malate was about twice the size of the aspartate pool. However, there was strong evidence for a rapid flux of carbon through both these pools. Possible routes of aspartate metabolism and the relationship between this metabolism and PSII activity in bundle sheath cells are considered.Abbreviations DHAP dihydroxyacetone phosphate - NADP-ME(-type) NADP-malic enzyme (type) - NADP-MDH NADP-malate dehydrogenase - OAA oxaloacetic acid - 2-OG 2-oxoglutarate - PEP phosphoenolpyruvate - PGA 3-phosphoglycerate - Pi orthophosphate - Ru5P ribulose 5-phosphate     

Stable isotope characteristics across narrow savanna/woodland ecotones in Wolfe Creek Meteorite Crater,Western Australia

The stable isotopic composition (δ¹³C) of sediments from lakes are frequently analyzed to reconstruct the proportion of the regional vegetation that used either the C₃ or C₄ photosynthetic pathways, often without conducting a detailed survey of the current local vegetation. We performed a study on the modern vegetation composition within the Wolfe Creek Meteorite Crater to complement our future paleoecological investigation of the crater. A bull’s-eye pattern exists where C₄ grasses dominate an outer ring and salt tolerant species, including shrubs, herbs, chenopods, and halophytic algae, dominate the inner pan of the crater. The ecotone between the inner and outer zones is narrow and occupied by tall (>7 m) Acacia ampliceps, with some C₄ grasses in the understory. Along with the highest water table and most saline soils the center of the crater has C₃ plants present with the highest δ¹³C and δ¹⁵N values. The range of δ¹³C and δ¹⁵N values from the analysis of surface soil organic matter (OM) was much smaller compared with the range of values from plant materials implying that either: (1) the current plant OM has not yet been integrated into the soils, or (2) processes within the soil have acted to homogenize isotopic variability within the crater. The application of a two end member mixing model to calculate %C₄ and %C₃ biomass from the δ¹³C of surface soil OM was complicated by: (1) the crater containing both a dry habitat with C₄ grasses and a central pan with C₄ halophytic plants and, (2) the large variation in the δ¹³C of the plants and soil OM.