Abstract: Ultrastructural studies of leaves of seven Panicum species in or closely related to the Laxa group and classified as C3, C4 or C3-C4 intermediate were undertaken to examine features associated with C3 and C4 photosynthesis. The C3 species Panicum rivulare Trin. had few organelles in bundle sheath cell profiles (2 chloroplasts, 1.1 mitochondria, and 0.3 peroxisomes per cell section) compared to an average of 10.6 chloroplasts, 17.7 mitochondria, and 3.2 peroxisomes per bundle sheath cell profile for three C3-C4 species, Panicum milioides Nees ex Trin., Panicum decipiens Nees ex Trin. and Panicum schenckii Hack. However, two other C3 species, Panicum laxum Sw. and Panicum hylaeicum Mez, contained about 0.7, 0.5, and 0.3 as many chloroplasts, mitochondria, and peroxisomes, respectively, as in bundle sheath cell profiles of the C3-C4 species. Chloroplasts and mitochondria in bundle sheath cells were larger than those in mesophyll cells for the C4 species Panicum prionitis Griseb. and the C3-C4 species, but in C3 species the organelles were similar in size or were smaller in the bundle sheath cells. The C3-C4 species and P. laxum and P. hylaeicum exhibited an unusually close association of organelles in bundle sheath cells with mitochondria frequently surrounded in profile by chloroplasts. The high concentrations in bundle sheath cells of somewhat larger organelles than in mesophyll cells correlates with the reduced photorespiration of the C3-C4 species.

Abstract: Mitochondria were isolated from mesophyll protoplasts and bundle sheath protoplasts or strands which were obtained by enzymatic digestion of six C4 species: Zea mays, Sorghum bicolor, Panicum miliaceum, Panicum capillare, Panicum maximum and Chloris gayana, representative of three C4 types. Photorespiratory glycine oxidation and related enzyme activities of mesophyll and bundle sheath mitochondria were compared. Mesophyll mitochondria showed good P/O ratios with malate and succinate as substrate but lacked the ability to oxidize glycine. On the other hand, mitochondria isolated from bundle sheath protoplasts of P. miliaceum and bundle sheath strands of Z. mays possessed glycine oxidation activity similar to that of mitochondria from C3 plant leaves. The two enzymes involved in glycine metabolism in mitochondria, serine hydroxymethyltransferase and glycine decarboxylase, were also assayed in the mitochondria of the two cell types. The activities of the two enzymes in bundle sheath mitochondria were in the range found in C3 mitochondria. In contrast, the activities in mesophyll mitochondria were either not detectable or far lower than those in bundle sheath mitochondria and ascribed to contaminating bundle sheath mitochondria. The present results indicate the deficiency of a complete glycine oxidation system in mesophyll mitochondria and also a differentiation between mesophyll and bundle sheath cells of C4 plants with respect to the photorespiratory activities of the mitochondria.

Abstract: The paraveinal mesophyll (PVM) is a unique and specialized, one-cell-thick tissue spanning the vascular bundles at the level of the phloem in soybean (Glycine max) (L.) Merr.) leaves. Its position within the leaf dictates that all photosynthate produced in the palisade and spongy mesophyll must pass through this specialized layer enroute to the phloem. Symplastic continuity, via plasmodesmata, exists between the PVM and bundle sheath, palisade parenchyma and spongy mesophyll. During leaf ontogeny the PVM is the first tissue to differentiate and at maturity these cells are six to eight times larger than other mesophyll cells, are highly vacuolate, and are interconnected by tubular arms. The PVM undergoes several unique structural and metabolic modifications during leaf development. The PVM cytoplasm, in vegetative plants, is dense, enriched in rough endoplasmic reticulum and dictyosomes, but contains few, small starch-free chloroplasts and few microbodies. Unlike the tonoplast of mesophyll cells, the tonoplast of the PVM is unusually thick and dense-staining. During leaf development the vacuoles of PVM cells accumulate a glycoprotein derived from the dictyosomes which reacts with the protein staining reagents, mercuric bromophenol blue and sulfaflavine, and is degraded by Pronase. Both the vacuolar material and tonoplast are also stained by phosphotungstic acid, which at low pH is relatively selective for glycoprotein. A unique role of the PVM in the transport and compartmentation of nitrogen reserves in soybeans is discussed.

Abstract: A mechanical isolation procedure was developed to study the respiratory properties of mitochondria from the mesophyll and bundle sheath tissue of Panicum miliaceum, a NAD-malic enzyme C(4) plant. A mesophyll fraction and a bundle sheath fraction were obtained from young leaves by differential mechanical treatment. The purity of both fractions was about 80%, based on analysis of the cross-contamination of ribulose bisphosphate carboxylase activity and phosphoenolpyruvate carboxylase activity.Mitochondria were isolated from the two fractions by differential centrifugation and Percoll density gradient centrifugation. The enrichment of mitochondria relative to chloroplast material was about 75-fold in both preparations.Both types of mitochondria oxidized NADH and succinate with respiratory control. Malate oxidation in mesophyll mitochondria was sensitive to KCN and showed good respiratory control. In bundle sheath mitochondria, malate oxidation was largely insensitive to KCN and showed no respiratory control. The oxidation was strongly inhibited by salicylhydroxamic acid, showing that the alternative oxidase was involved. The bundle sheath mitochondria of this type of C(4) species contribute to C(4) photosynthesis through decarboxylation of malate. Malate oxidation linked to an uncoupled, alternative pathway may allow decarboxylation to proceed without the restraints which might occur via coupled electron flow through the cytochrome chain.

Abstract: Lipoxygenases 1 and 2 were localized in etiolated germinating soybean seeds ( Glycine max [L.]. Merr. var. Williams) by an indirect immunofluorescence staining technique. Sections of paraffin-embedded seedlings were stained with affinity-purified antibodies directed against lipoxygenase 1 or 2. The specificity of the immunofluorescence technique was examined by use of nonimmune serum or immunoglobulin G preparations after total adsorption with the appropriate lipoxygenase coupled to Sepharose 4B. After immunofluorescence staining with antilipoxygenase 1 or 2 IgG storage tissues of cotyledons fluoresce strongly the first days of germination. After 3 days, the abaxial hypodermis, the epidermis, and the vascular bundle sheaths show fluorescence, especially after incubation with antilipoxygenase 2 IgG. Fluorescence in cortex and pith of the hypocotyl migrates to the vascular cylinder during germination. In primary leaves, all tissues show fluorescence after 1 day of germination. In storage tissues of cotyledons, cytoplasm around the protein bodies fluoresces, whereas in other tissues protein bodies or other large cell organelles fluoresce. It is reasonable to suggest that lipoxygenase exerts its function in cells at the time that rigorous changes in metabolism take place, namely at the start of mobilization of reserves in storage tissues and start of biosynthesis of chloroplastids in several tissues.

Abstract: The purpose of this study was to assess the relationship of oven-dry density, initial moisture content, vascular bundles and shrinkage to stem height and diameter of Cocos nucifera L. High correlations were obtained between initial moisture content and stem height, initial moisture content and oven-dry density, oven-dry density and stem height, vascular bundles per cm2 and stem height, vascular bundles and oven-dry density for cross-sections and shrinkage and vascular bundles per cm2. The high correlations between vascular bundles per cm2 and oven-dry density on the one hand and to shrinkage on the other can be used for visual grading of coconut timber.

Abstract: Photosynthesis in green plants or algae may be represented by an overall equation: The energy necessary to promote this overall reaction is derived from light through absorption by pigment molecules — chiefly the chlorophylls.Photosynthesis occurs in chloroplasts - subcellular organelles in which all the chlorophyll pigments are located. The chloroplasts comprise membranous, structures, and can be classified into two types. To the first type belong chloroplasts with appressed stacks of lamellar membranes, termed grana. These chloroplasts occur in mesophyll cells (C3 plants). The second type of chloroplasts are those with lamellar membranes that do not form the grana structures; they occur in bundle sheath cells of maize and other monocotyledons (C4plants, Hatch & Slack, 1970). In algae a greater diversity of structure occurs (Kirk & Tilney-Basset, 1978). Fluorescence microscopy indicates that chlorophyll molecules are localized mainly in the grana membrane regions of mesophyll-type chloroplasts and uniformly throughout the bundle sheath cells (Spencer & Wildman, 1962). Mesophyll chloroplasts are flattened saucer-shaped organelles (20 or more in each cell) of between 5000 and 10000 nm in diameter, and of thickness 1000–2000 nm, whilst the individual grana are each of the order of 300–500 nm in diameter. The available evidence suggests that individual lamellar membranes are arranged to form vesicles, or sacks where the internal space is completely delimited from the external space. These individual closed membrane structures were termed thylakoids (Menke, 1962).

Abstract: In Panicum species of the Laxa group, some of which have characteristics intermediate to C3 and C4 photosynthesis species, some mitochondria in leaf bundle sheath cells are surrounded by chloroplasts when viewed in profile. Serial sectioning of leaves of one Laxa species, Panicum schenckii Hack, shows that these mitochondria are enclosed by chloroplasts. Complete enclosure rather than invagination also is indicated by absence of two concentric chloroplast membranes surrounding the mitochondrial profiles.

Abstract: Quantitative cytochemical studies of cortical parenchyma cells of roots of Pisum sativum in which the central vascular bundle is severed, showed esterase activity to be an early marker of the determination of cells to form a vascular bridge. Explantation, onto a basal culture medium, of wound segments taken from roots at different times after severing the stele showed the irreversibility of the esterase activity on removal from the inducing environment, so confirming this as a marker of cell determination. A general determination for the stele was shown to occur by 8-10 h after wounding, but information relating to tracheid secondary-cell-wall formation was not apparently available until 18-20 h after wounding. Determination appeared to occur well before mitosis. The timings of the differentiation steps indicate a simple diffusion model to explain the mechanism of arrival of the initiating molecules.

Abstract: The aim of this work was to investigate the fate of phosphoenolpyruvate (PEP) produced by decarboxylation of oxaloacetate during photosynthesis in the bundle sheaths of leaves of the PEP-carboxykinase C4 grass Spartina anglica Hubb. Mesophyll protoplasts and bundle sheath cells were separated enzymically and used to investigate activities and distributions of putative enzymes of the C4 cycle and the photosynthetic carbon metabolism of bundle sheath cells. The results indicate that neither conversion of PEP to pyruvate nor its conversion to 3-phosphoglycerate can account for all of the carbon flux through the C4 cycle during photosynthesis. It is likely, therefore, either that PEP moves directly from bundle sheath to mesophyll or that more than one pathway of regeneration of PEP is involved in the C4 cycle in this plant.

Abstract: The oxidation of carbohydrate by the pentose-phosphate pathway in the shoot apical meristem and developing leaf primordia of Dianthus chinensis was assessed by measuring the activity of glucose-6-phosphate dehydrogenase (EC 1.1.1.49). On a kg(-1) dry weight h(-1) basis, activity rose from 250 mmol in the apical meristem to 550 mmol in the first two leaf primordia and then declined to 350 mmol in the sixth pair of leaf primordia, and finally to 200 mmol in leaves just emerged from the shoot bud. Measurements of activity in the sixth leaf pair from the apex showed differential distribution in leaf tissues. Epidermal and mesophyll tissue had about the same activity as whole-leaf tissue, but vascular bundles had 70% greater activity. Within the vascular tissue, activity in the phloem was twice as high as in the xylem. When activity was expressed on a per-cell basis, there was a continuous increase from 20 fmol in the apex to 2 pmol in the sixth leaf pair. Activity on a per unit cell volume basis showed that cells of the apical meristem and the epidermis, mesophyll and xylem of the sixth leaf pair had similar values, about 30 amol; only the two youngest pairs of primordia and the phloem had values two or three times this amount.

Abstract: Ultrastructural studies of leaves of seven Panicum species in or closely related to the Laxa group and classified as C3, C4 or C3-C4 intermediate were undertaken to examine features associated with C3 and C4 photosynthesis. The C3 species Panicum rivulare Trin. had few organelles in bundle sheath cell profiles (2 chloroplasts, 1.1 mitochondria, and 0.3 peroxisomes per cell section) compared to an average of 10.6 chloroplasts, 17.7 mitochondria, and 3.2 peroxisomes per bundle sheath cell profile for three C3C4 species, Panicum milioides Nees ex Trin., Panicum decipiens Nees ex Trin. and Panicum schenckii Hack. However, two other C3 species, Panicum laxum Sw. and Panicum hylaeicum Mez, contained about 0.7, 0.5, and 0.3 as many chloroplasts, mitochondria, and peroxisomes, respectively, as in bundle sheath cell profiles of the C3-C4 species. Chloroplasts and mitochondria in bundle sheath cells were larger than those in mesophyll cells for the C4 species Panicum prionitis Griseb. and the C3-C4 species, but in C3 species the organelles were similar in size or were smaller in the bundle sheath cells. The C3-C4 species and P. laxum and P. hylaeicum exhibited an unusually close association of organelles in bundle sheath cells with mitochondria frequently surrounded in profile by chloroplasts. The high concentrations in bundle sheath cells of somewhat larger organelles than in mesophyll cells correlates with the reduced photorespiration of the C3C4 species.

Abstract: The occurrence and level of nuclear DNA endoreplication (Feulgen cytophotometry) were compared in mesophyll and bundle sheath cells during leaf differentiation ofMuscari comosum (2 C DNA-12.7 pg),Allium cepa (2C DNA-33.5 pg),Allium porrum (2 C DNA -65.3 pg), fromLiliaceae family, andAmaryllis belladonna (2 C DNA-30.1 pg) andClivia miniata (2C DNA-46.9 pg) fromAmaryllidaceae family. In all the studied species mesophyll nuclei contain 2 C DNA, whereas bundle sheath cells inLiliaceae family replicate nuclear DNA up to 4 C-16 C level, inAmaryllidaceae family they remain at 2 C level. Although the number of chloroplasts per cell is species- and cell-specific, some relationship exists between the plastid index, plastid growth and the occurrence of nuclear DNA endoreplication. No correlation has been found between the level of nuclear endopolyploidy and the number of chloroplasts per cell.

Abstract: Bundle sheath strands (free of mesophyll tissue) were isolated from leaves of Zea mays L., Panicum miliaceum L. and Chloris gayana L. The response of oxygen exchange to CO2 and O2 concentration in these strands was examined using 18O2 and mass spectrometry. Bundle sheath strands from all these species showed considerable O2 uptake at low CO2, 40-80 µmol (mg Chl)-1 h-1. This O2 uptake was stimulated by O2 in the range 60-300 µM and was suppressed 60-70% by 5 mM NaHCO3. The remaining light-dependent O2 uptake was similarly sensitive to increasing O2 concentration. These results are interpreted in terns of the CO2 concentrating function of C4 photosynthesis and the low apparent rates of photorespiration observed in C4 plants in vivo.

Abstract: Young rolled leaves of sugarcane (Saccharum sp., variety Co 1007) are cultivated in vitro on a synthetic medium. The first anatomical modifications appear in nonlignified, subepidermal fibers, on the external side of the leaves, by the 3rd day. Cellular growth precedes proliferation. Thereafter the process of dedifferentiation affects all of the tissues of the leaves. Proliferations of epidermal tissue, subepidermal fibers, bundle sheath cells, and vascular tissue give rise, respectively, to the epidermal tissue, peripheral cambial zone, parenchyma, and vascular tissues of the calluses. Thus, organogenic specificity of each foliar tissue is preserved.

Abstract: Developmental study of the stem-node-leaf vascular continuum of Austrobaileya scandens White reveals that the vasculature within each leaf originates from a single procambial strand, that becomes separated into two strands only at the junction of leaf and stem At lower levels in the stem the two strands become incorporated into independent portions of the stele At later stages of development the solitary vascular bundle within the young leaf undergoes considerable lateral growth, resulting in an essentially continuous arc of vascular tissue Ontogenetic evidence indicates that the vascular bundle in the midrib of the lamina should be regarded as a fundamentally single bundle and not interpreted as two bundles that have undergone various degrees of secondary fusion A condition of two totally separate bundles extending the entire length of the leaf was not encountered Our observations confirm the characterization of Austrobaileya as an example of "second rank" level of leaf vasculature Nodal anatomy emphasizes the extremely isolated taxonomic position of Austrobaileya within the primitive dicotyledons THE DISCOVERY ofthe nodal anatomical pattern in which a unilacunar node is associated with two leaf traces, each related to an independent portion of the stem vascular cylinder, brought about a reassessment of ideas relating to the evolution of nodal vasculature in dicotyledons Suggestions were made that the unilacunar, twotrace nodal condition was primitive in dicotyledons (Canright, 1955; Marsden and Bailey, 1955; Bailey, 1956) This idea was subsequently accepted by a number of individuals (Eames, 1961; Cronquist, 1968; Foster and Gifford, 1974) One of the primary evidences used in support of the primitive nature of the 2:1 node is its occurrence (or modifications thereof) in various members of the primitive Ranalean complex, including the monotypic family Aus

Abstract: Our current understanding of the photosynthetic process in species utilizing the C4 photosynthetic pathway suggests that photosynthetic efficiency should be enhanced by: 1) maximizing the conductance of the gas phase transport pathway from the leaf exterior to the mesophyll cell surfaces; 2) maximizing cytoplasmic connections and metabolite transport between bundle sheath and mesophyll parenchyma cells; and 3) minimizing the conductance of the gas phase transport pathway from the bundle sheath cells to the leaf exterior. In this study we have examined several species in the Poaceae with C4 photosynthesis to determine if there is any evidence for anatomical specialization which would lead to enhanced photosynthetic efficiency by these processes. Observations with light and scanning electron microscopes revealed specializations in mesophyll cell morphology and arrangement which include branched cells forming intercellular channels. These specializations are hypothesized to contribute to photosynthetic efficiency through its influence on the above transport processes.

Abstract: Incisions used to offer radioisotopes and conditioning chemicals were monitored for their effect on the transport system, using movement of nC-photosynthate and 14C-sucrose to study phloem transport in Helianthus and India ink to study xylem water movement in Heracleum. It was found that cuts made in air were more effective than cuts made under water, in part because both radioisotopes and conditioning chemicals penetrated more readily when cuts were made in air. Incisions made while the stem was under a backpressure, created by a pressure bomb, neutralized the release of pressure by the cut as seen in the movement of India ink in the xylem and biopotential changes as seen in vascular bundles.

Abstract: In this paper, we report an observation with light and electron microscope on the process of the secondary wall thickening and lignin accumulation in the cells of the 4th internode from the top of rice culm, var. Koshihikari. Materials for ultra thin sections were fixed in 2-4% glutaraldehyde, postfixed with 1-2% osmium tetroxide, and embedded in Spurr's resin. Meshes were stained with uranium and lead. For histochemical detections of lignin, free hand sections, 60-80 μm in thickness, were treated with Maule's reagents (M-reaction) and phloroglucinol-HCl (P-reaction). The secondary wall of the cortical fibers makes a start of thickening just after the cell has completed its elongation. It usually consists of three layers; S1, S2 and S3. In crease in secondary wall thickness is associated with increase in number of Golgi apparatus, Golgi vesicles, exocytosis and invagination. The ribosomes are often present as polysomes on the rough endoplasmic reticulum, some of which connect with the plasmalemma (Figs. 1-4). The fiber cells have smaller diameter and thicker walls in the outer rows near the epidermis than those in the inner rows which sometimes lack S3 layer. The secondary walls of the tips of fiber cells thicken more slowly and are thinner than those of the central part (Fig. 5). The epidermal long cells have a large number of small papillae, and a number of large ones on both sides of the short cells, but a fewer papillae on the small vascular bundle (Fig. 6). The aspects of cell organelles at their wall thickening stage are similar to those in the cortical fibers (Figs. 8-10). At this stage, silica deposition cetripetally progresses from the inside of cuticular layer in all the outer walls of the long cells, especially in those of the silica cells (Figs. 9-12). As for the parenchyma cells, a large vacuole is formed from an early stage of cell development, and conspicuous invaginations are often seen accompanying with ribosomes (Fig. 13). No lignin is detected in elongating cell walls except those of the protoxylem vesseles. On and after beginning of the secondary wall thickening, M-reaction begins by appearing (Fig. 14A). On and after beginning of the S2 layer thickening, P-reaction follows up the traces of M-reaction. M-reaction decreases in the part where P-reaction is positive (Fig. 14B). In the 4th mature internode, P-reaction is generally intensive in thc epidermis, cortical fibers and small vascular bundle. M-reaction is thc most intensive in the large vascular bundle sheath (Fig. 14C). In a fiber cell, P-reaction is the most intensive in the middle lamella-primary wall complex and decreases centripetally. On the contrary, M-reaction is intensive on the faint parts of P-reaction, but both reactions are not obvious in the parenchyma. In briefly saying, P- and M-reactions relate complementally each other. The secondary wall thickening and the lignin accumulation in the 4th internode of the cultivar terminate about 5 days after heading and entirely complete after next 10 days.

Abstract: Primary leaf plastid development and photosynthetic activity in Echinochloa (barnyard grass) were monitored during a 5 day time course of germination and growth in a nitrogen atmosphere. After 8 hours imbibition in the light but without oxygen, the proplastids were spheroidal and contained a few rudimentary stromal thylakoids. However, during the next 126 hours of anaerobiosis, plastid size and stromal thylakoid numbers increased markedly. Under anaerobiosis plastid morphology was also characterized by the development of prolamellar bodies, previously reported only in etiolated tissues, while changes in starch grains and plastoglobuli indicated carbohydrate and lipid metabolism, respectively. Upon exposure to air the arrested development of chloroplasts was reversed. Compared to etioplasts when exposed to light, however, there was a lag for the first 18 hours before the plastids developed rapidly into typical, C4 mesophyll and bundle sheath chloroplasts.

Abstract: O-acetyl-L-serine sulfhydrylase is an enzyme of assi-milatory sulfate reduction. It catalyses the formation of cysteine from O-acetyl-L-serine and sulfur reduced to the thiol level. The pathway begins with the formation of adenosine 5′-phosphosulfate (APS) from ATP and SO4 via ATP sulfurylase (E. C. 2. 7. 7. 4). In C4 plants ATP sulfurylase is predominantly localized in the bundle sheath cells (1, 2), while O-acetyl-L-serine sulfhydrylase was detected in meso-phyll protoplasts and bundle sheath strands (2). We have shown that various forms of O-acetyl-L-serine sulfhydrylase exist in spinach (3) and Phaseolus vulgaris (4). These findings taken together prompted us to examine, whether different forms of the enzyme occured in mesophyll cells and bundle sheath cells of the C4 plant Zea mays. For comparison, we included wheat (Triticum aestivum), a C3 grass.

Abstract: Intraspecific variability in Alloteropsis semialata (R. Br.) Hitchc. is shown ultrastructurally with particular regard to the structure of the bundle sheath and the development of the dimorphic chloroplasts in both the C 3 and C 4 forms of this species. A. semialata is known to possess two distinct anatomical forms — Kranz and non-Kranz — within one species and occurring within a single ecological niche. Kranz and non-Kranz anatomy is known to be correlated with C 3 and C 4 physiology respectively. Transverse sections of leaf portions taken at midsheath, at the ligule and at midlamina show plastids with different morphologies at different ontogenetic stages. Plastid form is related to the stage of development, the influence of light on the emerging leaf and the C 3 or C 4 form of A. semialata . Stages from amyloplast to chloroplast are investigated with regard to fine structure. Leaf transverse sections are examined microscopically and formation of new bundles, chiefly in the lamina is traced. Differences in anatomy and distribution of vascular bundles are more evident in the lamina than in the colourless leaf sheath. Both C 3 and C 4 forms of A. semialata are found to show chloroplast dimorphism in vascular bundle sheath and mesophyll cells. This is shown to differ in the two forms. The specialized chloroplasts of the Kranz sheath are shown to develop in the inner or mestome sheath, and not in the parenchyma sheath as in some other members of the Kranz Panicoideae. Features of A. semialata, such as the double bundle sheath, granal chloroplasts and large numbers of mitochondria in bundle sheath cells, in the C 4 form, question the current classification of this grass as a malate former.

Abstract: Differences in ease of digestion of bundle sheath cell walls of leaf blades of Panicum spp. are shown to be associated with differences in bundle sheath cell wall structure, including the presence or absence of a suberized lamella. These structural differences are correlated with photo-synthetic type (viz. C3, intermediate C3/C4, and the C4 types, PCK, NADP-ME and NAD-ME), as are mesophyll: bundle sheath area ratios.