C4-dicarboxylic Acid Pathway Research Articles

Metabolic pathways of CO2 fixing microorganisms determined C-fixation rates in grassland soils along the precipitation gradient

CO2 fixing microorganisms (CFMs) play a crucial role in carbon (C) sequestration in vegetation restricted areas, e.g., under semiarid and arid conditions. The factors controlling the underlying pathways of the CO2 fixation by microorganisms living in soils remain unclear. Here, almost all genes responsible for the eight CO2 fixation pathways in semiarid soil CFMs communities were identified using metagenomic analysis: including the reductive citrate cycle (rTCA), dicarboxylate-hydroxybutyrate cycle (DC/4-HB), reductive pentose phosphate cycle (Calvin), 3-hydroxypropionate bicycle (3-HP), 3-hydroxypropionate/4-hydroxybutyrate (3-HP/4-HB), C4-dicarboxylic acid, CAM cycle, and reductive acetyl-CoA pathway (Wood-Ljungdahl pathway). By tracing the CO2 fixation flux via 13C labeling, it was shown that the CO2 fixation rates increased along the precipitation gradient. The rTCA and 3-HP pathways for CO2 fixing microorganisms were closely associated with 13C incorporation into the soil organic matter under high mean annual precipitation (MAP) (400–600 mm), whereas the Calvin cycle played a vital role in soils under low MAP (<400 mm) conditions. The abundance of the key genes within the C fixing pathways showed that the microbial C accumulation in soils was mainly influenced by the MAP. In semi-arid to semi-humid grassland soils, where CO2 fixation by CFMs provided about 8.1–27 mg C m−2 day−1 input into the ecosystem, we demonstrated that the rTCA, Calvin, and 3-HP cycle were vital to this essential pathway of C sequestration.

Purification, crystallization and preliminary X-ray diffraction studies on pyruvate phosphate dikinase from maize.

Pyruvate phosphate dikinase (PPDK) from maize catalyzes the reversible conversion of ATP, orthophosphate and pyruvate to AMP, pyrophosphate and PEP. In higher plants, this enzyme is believed to be involved in the C(4) dicarboxylic acid pathway. PPDK was crystallized by the vapour-diffusion method using polyethylene glycol as a precipitant. The crystals belong to the orthorhombic space group C2, with unit-cell parameters a = 108.2, b = 100.2, c = 108.4 A, beta = 96.5 degrees, and diffract to 2.3 A using SPring-8 synchrotron radiation.

Diet selection by steers using microhistological and stable carbon isotope ratio analyses.

Two methods of determining diet botanical composition, microhistological (MH), and stable carbon isotope ratio (CR) analyses were used to determine botanical composition of ingesta and fecal grab samples in steers grazing rhizoma peanut-mixed tropical grass pastures. Three pastures were used over two grazing seasons, 1992 and 1993, in Brooksville, FL. A weighted-disc double-sampling technique was used to determine forage mass and botanical composition, percentage of rhizoma peanut (Arachis glabrata), grass (Paspalum notatum and Cynodon dactlyon), and forb (primarily Chenopodium ambrosioides) on offer every 28 d throughout the grazing seasons. There was an effect of sampling date (P<.001), sampling date x pasture (P<.001), and sampling date x year (P<.001) on forage mass on offer. There was a pasture x year x sampling date interaction (P<.001) for all botanical components. In 1992 and 1993, using cannulated steers sampled every 56 d, there were interactions with year for rhizoma peanut and forb (P<.05), but not for grass with MH analysis (components: rhizoma peanut, grass, and forb). Ingesta and fecal rhizoma peanut (r = .73 and .92 for 1992 and 1993, respectively) and ingesta and fecal forb (r = .86 and .98 for 1992 and 1993, respectively) were positively correlated (P<.001). Ingesta and fecal grass were positively correlated (r = .52, P<.001), but the correlation was not as high. With the CR analysis (components: Calvin cycle [C3] plants and C4-dicarboxylic acid pathway [C4] plants), ingesta and corrected fecal (corrected for in vitro organic matter digestibility [IVOMD]) C3 plants were positively correlated (r = .62; P<.001). Diet composition of fecal grab samples from noncannulated steers, collected on the same sampling schedule as for hand-clipped pasture samples, differed at times due to the complexity of the sward (both rhizoma peanut and forb constituted a single component, C3, in the CR analysis). Based on these results, if there is a substantial contribution of forb to the diet, fecal microhistological analysis may be more informative than fecal carbon ratio analysis for estimating diet selection by cattle grazing tropical pastures.

Analyzing the Light Energy Distribution in the Photosynthetic Apparatus of C4 Plants Using Highly Purified Mesophyll and Bundle-Sheath Thylakoids.

The chlorophyll fluorescence characteristics of mesophyll and bundle-sheath thylakoids from plant species with the C4 dicarboxylic acid pathway of photosynthesis were investigated using flow cytometry. Ten species with the NADP-malic enzyme (NADP-ME) biochemical type of C4 photosynthesis were tested: Digitaria sanguinalis (L.) Scop., Euphorbia maculata L., Portulaca grandiflora Hooker, Saccharum officinarum L., Setaria viridis (L.) Beauv., Zea mays L., and four species of the genus Flaveria. This study also included three species with NAD-ME biochemistry (Atriplex rosea L., Atriplex spongiosa F. Muell., and Portulaca oleracea L.). Two C4 species of unknown biochemical type were investigated: Cyperus papyrus L. and Atriplex tatarica L. Pure mesophyll and bundle-sheath thylakoids were prepared by flow cytometry and characterized by low-temperature fluorescence spectroscopy. In pure bundle-sheath thylakoids from many species with C4 photosynthesis of the NADP-ME type, significant amounts of photosystem II (PSII) emission can be detected by fluorescence spectroscopy. Simulation of fluorescence excitation spectra of these thylakoids showed that PSII light absorption contributes significantly to the apparent excitation spectrum of photosystem I. Model calculations indicated that the excitation energy of PSII is efficiently transferred to photosystem I in bundle-sheath thylakoids of many NADP-ME species.

Ecological evidence concerning the adaptive significance of the C4 dicarboxylic acid pathway of photosynthesis.

Ecological data were analyzed to determine the environmental conditions associated with the occurrence of plants possessing C3 and C4 photosynthetic pathways. Non-parametric analysis of variance, multiple regression analysis and discriminant analysis techniques were applied to information derived from separate studies on plant species in California and central Europe. All the analyses revealed significant differences in ecology between the C3 and C4 groups. The occurrence of C4 species was greater where summer or winter temperatures were relatively high and moisture availability relatively low. Using discriminant analysis, more than 80% of the species considered were classified into the correct photosynthetic category on the basis of their distribution with respect to temperature. No significant differences between the C3 and C4 groups were found for other environmental factors, including light, soil nitrogen, soil salinity and continentality of climate. These results support the suggestion from previous physiological studies that C4 species may possess competitive advantages under conditions of high temperature and intermittent water stress. The C4 species were relatively restricted in the range of environmental conditions where they occurred, and as a group C4 species may be ecologically more specialized than C3 species.

13C Content of human collagen as a measure of prehistoric diet in woodland North America

PLANTS with the C4 dicarboxylic acid pathway of photosynthesis have markedly higher 13C/12C ratios than plants with the C3 or Calvin pathway1–4. Cumulative assessments of relative 13C concentration (δ13C) values for C3 plant foliage yield an average of −26.5‰, relative to the PDB standard whereas C4 plants average −12.5‰ (ref. 5). These differences in carbon isotope ratios are passed along the food chain to animals and man, so that the nature of the plant food base can be determined through measurements on different body tissues of the consumers. In this study, the time of introduction of maize, a C4 plant, to the prehistoric inhabitants of several regions in woodland North America, almost exclusively a C3 plant regime, was determined through isotopic measurements on human bone collagen. The subsequent elevation of maize to the level of a diet staple is also documented.

Studies of Host Plant Preference and Suitability Exhibited by Early-instar Range Caterpillar Larvae 12

The preferences of instar II range caterpillar, Hemileuca olivae Cockerell, were tested relative to blue grama grass. Buffalograss, sorghum, foxtail millet, and sudan grass were highly preferred. Larvae fed upon, but did not prefer, Kentucky bluegrass, Russian wildrye, and side-oats grama. Non-preferred species were Japanese millet, western wheatgrass, corn, proso millet, wheat, barley, and oats. The grasses most suitable for instar I and II larval development were sorghum, sudan grass, blue grama, and com. Foxtail millet, western wheatgrass and wheat were intermediate, Japanese millet, oats, barley, Russian wildrye, and Kentucky bluegrass least suitable. In general, plants possessing the C4-dicarboxylic acid pathway of carbon fixation (C4 plants) were preferred and more suitable for larval growth than species possessing only the C3-Calvin cycle pathway (C3 plants).

Endogenous Photophosphorylation by Mesophyll and Bundle Sheath Chloroplasts from Setaria italica and Amaranthus paniculatus1

The basic features of the C4 dicarboxylic acid pathway of photosynthesis are the fixation of carbon dioxide into C4 acids (namely oxalacetate, malate and aspartate) in the mesophyll cells and their transport into the bundle sheath cells where they are decarboxylated. Thus released carbon dioxide is refixed and further metabolized via the Calvin cycle in the sheath cells (Hatch and Slack, 1970; Black, 1973). Such two-step carboxylation is calculated to result in an energy cost of 5 ATP and 2 NADPH2 per molecule of C02 fixed, compared to that of 3 ATP and 2 NADPH2 per each C02 molecule reduced through Calvin cycle alone (Chen, Brown and Black, 1969; Hatch and Slack, 1970). Chen et al. (1969) suggested that the extra requirement of ATP in C4 plants is met with the capacities of C4 plant-chloroplasts to catalyze higher rates of cyclic photophosphorylation. Bundle sheath chloroplasts of C4 plants, producing malate as the principal C4 acid, e.g., maize and sorghum, are deficient in photosystem (PS) II compared to PS I (Anderson, Woo and Boardman, 19716; Downton, 1971 ; Mayne, Edwards and Black, 1971). Such PS II deficiency has important consequences for Calvin cycle metabolism in the bundle sheath cells of these plants, since all the carbon fixed during photosynthesis eventually has to be reduced through the Calvin cycle. In such instances, it is proposed that for normal C02 reduction, NADPH2 is provided by malic enzyme (Slack, Hatch and Goodchild, 1969; Downton, 1971) and ATP is supplied through cyclic photophosphorylation (Chen et al ., 1969 ; Slack et al ., 1969). These complications do not arise with other C4 plants which use aspartate as the major C4 acid (e.g. Amaranthus and Atriplex ) where bundle sheath chloroplasts possess an active PS II as well as PS I. Though the studies on mesophyll and bundle sheath chloroplasts showed high cyclic photophosphorylation (mediated through phenazine methosulphate) capacities of bundle sheath chloroplast fragments (Anderson, Boardman and Spencer, 1971a; Polya and Osmond, 1972), convincing evidence for endogenous cyclic photophosphorylation is still not available. Here we report the presence of an active endogenous photophosphorylation of bundle sheath chloroplasts insensitive to 3(3,4-dichlorophenyl)-l,l-dimethyl urea (DCMU), evidently mediated through PS I. Plants of Setaria italica Beauv. var. H-l and Amaranthus paniculatus L. were raised in seed pans under an approximate 12-h photoperiod (35 °C day and 20 °C night). The second to fourth leaves starting from the oldest leaf were picked from 3 to 4 week old seedlings. The leaves were chilled to 0 °C and cut into pieces of c. 1 cm2. Mesophyll and bundle sheath chloroplasts were prepared by differential grinding of leaf tissue (Raghavendra, 1975;

C4 pathway of photosynthesis in a member of the polygonaceae: Calligonum persicum(Boiss. &amp; Buhse) boiss

Summary Initial products of 14CO2 fixation, CO2 compensation point and carbon isotope discrimination indicate the occurrence of the C4 dicarboxylic acid pathway of photosynthesis in Calligonum persicum. In this species, the green centric branches do not have the typical form of Kranz-anatomy with a concentric arrangement of mesophyll and bundle sheath layers around the vascular bundles. Instead, they possess two distinct, peripheral rings of green cells. Small vascular bundles adjoin the internal face of the inner layer of green cells, but only at intervals.

Light-Enhanced Dark 14CO2 Fixation by Leaves in Relation to the C4 Dicarboxylic Acid Pathway of Photosynthesis

The technique of light-enhanced dark carbon dioxide fixation has been employed to critically examine the primary route of CO2 incorporation in leaves of two C4 plants (Setaria italica and Amaranthus paniculatus) in comparison with that of a C3 plant (Rumex vesicarius). Prior illumination of detached leaves enhanced their dark 14CO2 fixation five- to tenfold over that of unilluminated leaves. No variation was observed among C3 and C4 plants, in the conditions required for their light-enhanced dark fixation. Fifty to sixty percent of initially incorporated radioactivity by the leaves of R. vesicarius was in 3-phosphoglycerate. The leaves of S. italica and A. paniculatus fixed most of the radioactivity into the C4 acids malate and aspartate. Sugar phosphates, including 3-phosphoglycerate, were not labelled at all. Our data confirm that, in C4 plants, CO2 is initially fixed through phosphoenolpyruvate carboxylation and thus disprove earlier claims that ribulose bisphosphate can be the primary acceptor of CO2 in a C4 plant. The observations further suggested that aspartate formation during carbon fixation might be favoured by illumination.

A technique for determining the proportion of C3 and C4 species in plant samples using stable natural isotope's of carbon

SummaryA technique is described which uses the ratio of the natural 12C and 13C isotopes of carbon to calculate the proportions of Calvin pathway (or C3) species and C4 dicarboxylic acid pathway (or C4) species in mixed samples of shoots, roots or seeds. Mean percentage of a particular component can be predicted to within ± 2% of actual values with average standard errors of less than 1% with two component mixtures under good conditions using threefold replication. The technique is particularly useful for estimating the proportion of C3 and C4 species in samples of visually indistinguishable, intermingled root systems from mixed field communities or from competition experiments. The proportion of C3 and C4 species in both oesophageal fistula and feed samples can be obtained using this technique, and it is proposed that data from fistula and pasture samples could be used to determine the extent to which animals selectively eat C3 or C4 species. The advantages and disadvantages of the technique are discussed.

Carbon Reduction Pathways and Standing Crop in Three Chihuahuan Desert Plant Communities

The complete floras of three Chihuahuan desert plant communities were surveyed to determine the relative numbers of species that possess either the C3, C4 or CAM carbon reduction pathway. There were more C3, species in all three communities. Spring-flowering species were predominantly C3 and CAM; C4 species most commonly flowered in the summer. Plant standing crop data were used as an indication of the contribution of each pathway to the primary production of the communities. In the most geographically widespread of the three communities, an alluvial plain, CAM plants contributed 48% of the total biomass while only representing 10% of the possible species present. Less than 1% of the biomass was contributed by C4 species although they comprised 24% of the possible species present. The C4 pathway did make a substantial standing crop contribution in a very localized community, an ephemeral desert lake bottom, where one C4 grass species accounted for 90% of the biomass present. Early investigations by Bennett-Clark (1933) and more recent work by Kortschak et al. (1965) and Hatch et al. (1967) have shown that many plant species possess biochemical pathways whereby CO, can be supplied to the pentose or Calvin-cycle (C,) pathway for carbon reduction much more efficiently than species which have only the CS pathway. Plants which possess Crassulacean acid metabolism (CAM) pathway are able to fix CO, initially into organic acids in the mesophyll chlorenchyma during the dark and then subsequently utilize the CO, for the formation of carbohydrate via the C, pathway during the light period. Plants which possess the C4-dicarboxylic acid (C4) pathway fix CO, into organic acids in the mesophyll chlorenchyma in the light. This carbon is then reduced to carbohydrates by the C, pathway in the bundle sheath chlorencyhma. Since these supplemental pathways (CAM, C4) have been elucidated there has been much study and speculation concerning the adaptive significance and competitive advantage these pathways may add to the C, pathway (Black et al. 1969; Bjorkman and Berry 1973).

Photosynthetic Carbon Metabolism of a Marine Grass

The delta(13)C value of a tropical marine grass Thalassia testudinum is -9.04 per thousand. This value is similar to the delta(13)C value of terrestrial tropical grasses. The delta(13)C values of the organic acid fraction, the amino acid fraction, the sugar fraction, malic acid, and glucose are: -11.2 per thousand, -13.1 per thousand, -10.1 per thousand, -11.1 per thousand, and -11.5 per thousand, respectively. The delta(13)C values of malic acid and glucose of Thalassia are similar to the delta(13)C values of these intermediates in sorghum leaves and attest to the presence of the photosynthetic C(4)-dicarboxylic acid pathway in this marine grass. The inorganic HCO(3) (-) for the growth of the grass fluctuates between -6.7 to -2.7 per thousand during the day. If CO(2) fixation in Thalassia is catalyzed by phosphoenolpyruvate carboxylase (which would result in a -3 per thousand fractionation between HCO(3) (-) and malic acid), the predicted delta(13)C value for Thalassia would be -9.7 to -5.7 per thousand. This range is close to the observed range of -12.6 to -7.8 per thousand for Thalassia and agree with the operation of the C(4)-dicarboxylic acid pathway in this plant. The early products of the fixation of HCO(3) (-) in the leaf sections are malic acid and aspartic acid which are similar to the early products of CO(2) fixation in C(4) terrestrial plants.Electron microscopy of the leaves of Thalassia reveal thick walled epidermal cells exceedingly rich in mitochondria and C(3)-type chloroplasts. The mesophyll cells have many different shapes and surround air lacunae which contain O(2) and CO(2). The mesophyll cells are highly vacuolated and the parietal cytoplasm contains an occasional chloroplast. This chloroplast contains grana but the lamellar system is not as developed as the system in epidermal chloroplasts. Extensive phloem tissue is present but the xylem elements are reduced in this aquatic grass. The vascular tissue is not surrounded by bundle sheath cells.This work does not establish the exact relation between structure and function in Thalassia, but it does show the C(4)-type photosynthetic carbon metabolism in this grass involves epidermal and mesophyll cells and internally produced O(2) and CO(2) in the air lacunae.

Plant-herbivore interactions : The indigestibility of C4 bundle sheath cells by grasshoppers.

Plants with the C4 dicarboxylic acid pathway of photosynthetic CO2 fixation are generally nutritionally inferior to C3 (Calvin cycle) plants as foodstuff for herbivores. A possible contributing factor to this nutritional inferiority is the concentration, in C4 plants, of large quantities of nutritional material in very tough, thick-walled vascular bundle sheath cells which herbivores may not be able to break down. Experiments with 10 species of grass-hoppers from different areas in the United States revealed large numbers of unbroken bundle sheath cells, contents intact, in fecal pellets produced when the grasshoppers were fed C4 vegetation. We conclude that the material stored in C4 bundle sheath cells is at least partially unavailable to herbivores, and that this may contribute to the observed nutritional inferiority of C4 vegetation.

Differences in natural carbon isotope ratios of milk and hair from cattle grazing tropical and temperate pastures

ABOUT 99% of all carbon is the 12C isotope while 1% is 13C. The precise ratio of the isotopes will vary depending on the material analysed. In plants, fractionation of carbon is brought about primarily by carbon dioxide assimilation in photosynthesis and is due to preferential utilisation of 12C and exclusion of 13C. Curiously enough, it has been found recently1,2 that higher plants which fix carbon dioxide by way of the Calvin C3 cycle pathway differ in 13C/12C ratios from plants which fix carbon dioxide through the C4-dicarboxylic acid pathway. Temperate pasture species fix carbon by way of the Calvin pathway and have 13C/12C ratios of approximately −28‰ (ref. 3 and M.M.L., J.H.T., and R. J. Jones, unpublished), whereas tropical pasture grasses fix carbon through the C4-dicarboxylic acid pathway4 and have 13C/12C ratios of approximately −12‰ (ref. 2 and M.M.L., J.H.T., and R.J.J., unpublished). 13C/12C ratios are expressed relative to a carbonate standard5.

Photosynthesis in mesophyll protoplasts and bundle sheath cells of various types of C4 plants I. Carboxylases and CO2 fixation studies

Summary Mesophyll protoplasts and bundle sheath cells have been isolated from 3 groups of C4 plants by enzymatic digestion of leaves with cellulase and pectinase. In all species studied the phosphopyruvate carboxylase was localized almost exclusively in mesophyll cells, and ribulose-1, 5-diphosphate carboxylase in bundle sheath cells. Isolated bundle sheath cells from species having high NADP-malic enzyme activity showed varying degrees of light dependent fixation of CO2. Bundle sheath cells of sorghum and sugarcane showed little capacity for CO2 fixation while bundle sheath cells of Digitaria sanguinalis and Setaria lutescens showed substantial activity of light induced CO2 fixation. Similar or higher rates of CO2 fixation in the light were obtained with the isolated bundle sheath cells of C4 plants having low NADP-malic enzyme activity. The highest rate of CO2 fixation (103 μmoles/mg chl/hr) was obtained with bundle sheath cells of Panicum capillare. In all cases, there was a light induced fixation of CO2 by mesophyll protoplasts of C4 plants although the rates were low in comparison to the rates with isolated mesophyll protoplasts of C3 plants. The addition of phosphopyruvate to mesophyll protoplasts of C4 plants induced high rates of CO2 fixation. The results are discussed in relation to the proposed function of the C4-dicarboxylic acid pathway in C4 plants.

Translocation of Photosynthetic Assimilate From Grass Leaves, as Influenced by Environment and Species

The export of 14C-labelled photosynthate from leaves of two grasses with Calvin cycle (C*3) photosynthesis, and five grasses with the C4 dicarboxylic acid pathway, was followed by continuous monitoring through the hours in light immediately after assimilation of 14CO2 and also through the subsequent dark period. The proportion of activity exported in the first few hours was much greater in all C4 grasses than in the C3 species, being greatest in the C4 grasses best adapted to growth at cool temperatures. Large differences between species were also apparent in their rates of assimilate export per unit leaf area, but there was no consistent relation between these rates and the cross-sectional area of phloem serving a given catchment leaf area. The rate of assimilate transfer per unit phloem area in Lolium temulentum was comparable to rates previously recorded for other C3 plants, but the rates for all C4 grasses except Digitaria sanguinalis were considerably faster. The rate of export in Paspalum dilatatum increased progressively up to the highest light intensity used, giving no evidence of saturation of translocation capacity. It was greater in plants grown under higher light intensities, and increased with increasing sink demand, but was not increased by darkening or removing the leaf blade distal to the area exposed to 14CO2 in Lolium temulentum. Most of the photosynthate not exported within 6 h was stored as polysaccharide in Panicum maximum, and was subsequently mobilized over a brief period during the following night. 14C assimilated at the end of the light period was mobilized first, while that assimilated at the beginning of the day was mobilized only at the end of the following night. However, the rate of mobilization could be increased by partial defoliation or by other treatments which increased the demand for assimilates from the leaf. The time to mobilization was unaffected by night temperature in Lolium temulentum, but increased markedly at temperatures below 15°C in the C4 grasses Panicum maximum and Paspalum dilatatum. The adaptive value and some possible explanations of the differences between C3 and C4 grasses in their translocation of assimilates are considered.

Metabolic regulation in C4 photosynthesis: PEP-carboxylase and energy charge

The effects of ATP, ADP and AMP on the activity of PEP carboxylase from Pennisetum purpureum (a C4 plant) have been investigated. AMP caused slight changes in activity. Both ATP and ADP were inhibitory. The extent of inhibition was related to the concentration of Mg(2+). Kinetics of inhibition (with PEP as variable substrate) were investigated at limiting concentrations of Mg(2+) (2 mM); with excess Mg(2+) (5 mM); or with the concentration of Mg(2+) buffered at about 8 mM. At low Mg(2+) parabolic competitive-kinetics were observed. With higher Mg(2+) concentration linear competitive kinetics of inhibition were observed with a K i (slope) of 1.25 mM ATP. This was increased to 2.03 mM on addition of the allosteric activator glucose-6-P (5 mM). With Mg(2+) buffered ATP was an activator at low concentrations of PEP.Response to enegery charge was investigated using either calculated concentrations of all adenylates or mixtures of AMP and ATP pre-incubated with adenylate kinase. Inhibition at high charge could be reduced by increasing the concentration of Mg(2+) or by addition of the activator glucose-6-P. With enzyme-generated charge series and Mg(2+) buffered at 8 mM an increase in activity was observed at high charge values.These results are discussed in terms of a possible regulatory role of adenylates in the C4 dicarboxylic acid pathway of photosynthesis.

Oxaloacetate as the Hill Oxidant in Mesophyll Cells of Plants Possessing the C 4 -Dicarboxylic Acid Cycle of Leaf Photosynthesis

Isolated mesophyll cells from leaves of plants that use the C(4) dicarboxylic acid pathway of CO(2) fixation have been used to demonstrate that oxaloacetic acid reduction to malic acid is coupled to the photochemical evolution of oxygen through the presumed production of NADPH. The major acid-stable product of light-dependent CO(2) fixation is shown to be malic acid. In the presence of phosphoenolpyruvate and bicarbonate the stoichiometry of CO(2) fixation into acid-stable products to O(2) evolution is shown to be near 1.0. Thus oxaloacetic acid acts directly as the Hill oxidant in mesophyll cell chloroplasts. The experiments are taken as a firm demonstration that the C(4) dicarboxylic acid cycle of photosynthesis is the major pathway for the fixation of CO(2) in mesophyll cells of plants having this pathway.

Photosynthetic Pathways and Selective Herbivory: A Hypothesis

We propose that plants possessing the C4-dicarboxylic acid pathway of photosynthetic carbon fixation are generally inferior food sources for herbivores, and are often avoided by them, relative to plants possessing only the C3-Calvin cycle pathway. As initial support of this hypothesis, we present data from the literature, dealing primarily with insects, that suggest that C4 species are ingested in proportions lower than their availability in a number of natural situations, that they tend to be avoided in laboratory preference tests, and that they result in lower survival and fecundity in herbivores feeding on them. We suggest a number of physiological, anatomical, and nutritional differences between C3 and C4 species that may be involved in generating the observed pattern of herbivore preference. The ecological relevance of the differences between C3 and C4 species has been discussed to date only in relation to hypothetical effects on interspecific competition. If our hypothesis is true, selective herbivory may be another ecologically important consequence of the differences. There is enough evidence pointing toward the validity of our hypothesis that it should be further tested, and the distribution of plants as C3 and C4 species should be taken into account in community and ecosystem studies.