Abstract: The intercellular localization of enzymes involved in starch metabolism and the kinetic properties of ADPglucose pyrophosphorylase were studied in mesophyll protoplasts and bundle sheath strands separated by cellulase digestion of Zea mays L. leaves. Activities of starch synthase, branching enzyme, and ADPglucose pyrophosphorylase were higher in the bundle sheath, whereas the degradative enzymes, starch phosphorylase, and amylase were more evenly distributed and slightly higher in the mesophyll. ADPglucose pyrophosphorylase partially purified from the mesophyll and bundle sheath showed similar apparent affinities for Mg2+, ATP, and glucose-1-phosphate. The pH optimum of the bundle sheath enzyme (7.0-7.8) was lower than that of the mesophyll enzyme (7.8-8.2). The bundle sheath enzyme showed greater activation by 3-phosphoglycerate than did the mesophyll enzyme, and also showed somewhat higher apparent affinity for 3-phosphoglycerate and lower apparent affinity for the inhibitor, orthophosphate. The observed activities of starch metabolism pathway enzymes and the allosteric properties of the ADPglucose pyrophosphorylases appear to favor the synthesis of starch in the bundle sheath while restricting it in the mesophyll.

Abstract: In shoots of the dodder Cuscuta odorata mycoplasma-like organisms (MLO) were observed to occur in nearly all mature sieve elements. Their frequency within the sieve elements varied from a few organisms to high concentrations. In addition, MLO, were found in other cell types. By identifying these cells and investigating the location in the vascular tissue three different types of cells infected with MLO could be distinguished: (1) phloem parenchyma cells, (2) parenchyma cells separating the five vascular bundles of C. odorata and (3) cells in the outer region of the vascular system next to the cells looking like an endodermis. Within the cells of the types 2 and 3, MLO occurred in large numbers and at different morphological structures., Therefore, we assume that the MLO multiply in these cells.

Abstract: Intact mesophyll and bundle sheath chloroplasts wee isolated from the NADP-malic enzyme type C4 plants maize, sorghum (monocots), and Flaveria trinervia (dicot) using enzymic digestion and mechanical isolation techniques. Bundle sheath chloroplasts of this C4 subgroup tend to be agranal and were previously reported to be deficient in photosystem II activity. However, following injection of intact bundle sheath chloroplasts into hypotonic medium, thylakoids had high Hill reaction activity, similar to that of mesophyll chloroplasts with the Hill oxidants dichlorophenolindophenol, p-benzoquinone, and ferricyanide (approximately 200 to 300 micromoles O2 evolved per mg chlorophyll per hour). In comparison to that of mesophyll chloroplasts, the Hill reaction activity of bundle sheath chloroplasts of maize and sorghum was labile and lost activity during assay. Bundle sheath chloroplasts of maize also exhibited some capacity for 3-phosphoglycerate dependent O2 evolution (29 to 58 micromoles O2 evolved per milligram chlorophyll per hour). Both the mesophyll and bundle sheath chloroplasts were equally effective in light dependent scavenging of hydrogen peroxide. The results suggest that both chloroplast types have noncyclic electron transport and the enzymology to reduce hydrogen peroxide to water. The activities of ascorbate peroxidase from these chloroplast types was consistent with their capacity to scavenge hydrogen peroxide.

Abstract: Oosterhuis, D. M. and Wullschleger, S. D. 1987. Water flow through cotton roots in relation to xylem anatomy.—J. exp. Bot. 38: 1866-1874. The effect of root anatomy on water flow was studied in 7-d-old cotton (Gossypium hirsutum L.) seedlings grown in solution culture. The total water flux of the intact root system was measured using a pressure chamber. Then successive terminal root sections were removed at 2,6,10 and 12 cm behind the root tip and the flux was remeasured after each successive cut was made. Xylem development at different distances behind the root apex was studied with a microscope using sections cut free-hand and stained with toluidine blue. Water flux increased with the removal of successive terminal root sections and this coincided with the degree of basipetal primary xylem development. The large increase in water flux at 10 to 12 cm was associated with secondary xylem development and increased xylem vessel number. A comparison of water flow and xylem anatomy between roots with tetrarch (Stoneville 506 and Deltapine 41) and pentarch (T25 strain) vascular bundle arrangements showed no significant differences in the measured values of water flux for the primary root. Water flux, estimated using Poiseuille's equation and measured xylem dimensions, was greater for the tetrarch roots, primarily because of the larger diameter of individual vessel elements. The increased number of vessel elements in the pentarch primary root of T25 did not result in any apparent decrease in axial resistance to water flow.

Abstract: Three polypeptides of 33 kD, 23 kD and 16 kD are released from maize photosystem II particles by alkaline Tris solution treatment and shown to cross-react with antisera to purified spinach oxygen-evolving (OE) polypeptides of 34 kD, 23 kD and 17 kD, respectively. They are not located exclusively in mesophyll cells but each is about 4 times more abundant in the thylakoid membranes of mesophyll than bundle sheath cells of etiolated, greening and green leaves. Three maize cDNA clones (OE33, OE23, OE16) have been identified by hybrid-selection, in vitro translation and immunoprecipitation with antisera against spinach OEs. Transcripts of all three genes are already detectable in both mesophyll and bundle sheath cells of etiolated leaves; however, they accumulate transiently and coordinately in mesophyll cells but remain at a constant low level in bundle sheath cells upon illumination of dark-grown maize seedlings. Moreover, the level of each protein increases in mesophyll cells following the accumulation of transcripts during greening and remains high in late greening and green leaves, despite the decline in each corresponding mRNA. The accumulation of all three OE proteins is also stimulated by light in bundle sheath cells without increases in their corresponding mRNAs. The preferential localization of these three proteins in mesophyll cells is due to both transcriptional and translational regulation.

Abstract: Experiments were conducted with several Panicum species (representing the different C(4) subtypes) to examine the light modulation of sucrose phosphate synthase (SPS) activity and the effect of illumination on the distribution of SPS activity between mesophyll cells (MC) and bundle sheath cells (BSC). Activity of SPS in the light decreased in the order: C(4) > C(3)-C(4) intermediate > C(3). In illuminated leaves, SPS activities were similar among the three C(4) subtypes, but SPS activity was higher for NAD-malic enzyme (NAD-ME) species with centripetal chloroplasts in BSC (NAD-ME(P) species) than for NAD-ME species with centrifugal chloroplasts in BSC (NAD-ME(F) species). Transfer of plants into darkness for 30 minutes resulted in decreased SPS activity for all species tested except Panicum bisulcatum (C(3) species) and Panicum virgatum (NAD-ME(P) species) which showed little or no change. All C(4) subtypes had some SPS activity both in MC and BSC. In the light, SPS activity was mainly in the MC for NADP-ME, NAD-ME(F) and phosphoenolpyruvate carboxykinase species, while it was mainly in the BSC for NAD-ME(P) species. In the dark, for all C(4) subtypes, SPS activity in the MC was decreased to a greater extent than that in the BSC. It is intriguing that NAD-ME(F) and NAD-ME(P) species differed in the activity and distribution of SPS activity between MC and BSC, although they are otherwise identical in the photosynthetic carbon assimilation pathway. Diurnal changes in SPS activity in the MC and BSC were also examined in maize leaves. SPS activity in the MC in maize leaves was high and relatively constant throughout the middle of the light period, dropped rapidly after sunset and increased again prior to the light period. On the other hand, SPS activity in the BSC was lower and changed more coincidently with light intensity than that in the MC. The results suggested that light activation of SPS activity located in the BSC may require higher irradiance for saturation than the SPS in the MC. We conclude that SPS may function in both MC and BSC for sucrose synthesis in the light, particularly at high light intensity, while in the dark, the major function may be in the BSC during starch degradation.

Abstract: Leaves of the susceptible soybean cv. Clark were inoculated with virulent strains of Xanthomonas campestris pv. glycines, causative agent of bacterial pustule disease in soybean. After 7 d, leaf tissue bearing pustules was prepared for transmission and scanning electron microscopy. Pustules developed in the spongy parenchyma. Vascular-associated cells, including bundle sheath extensions, were not affected. The heaviest concentration of bacteria was in the intercellular spaces in or near pustules and bacterial extracellular polysaccharide was present. Bacteria were widely dispersed among the palisade cells, but the palisade parenchyma was not affected. There was no envelopment of bacteria at host cell wall surfaces and no bacteria were observed in the xylem or phloem elements of the minor leaf veins. Pustules were comprised of large mesophyll cells with uniform shape, unlike control spongy parenchyma cells which were irregular in shape. Affected cells had more endoplasmic reticulum and more ribosomes, both free and membrane-bound, than the control, and had few chloroplasts. Deep, focal invaginations of plasma membrane and tonoplast appeared to be involved in endocytic transport of membrane lamellae, small membrane vesicles, and non-organellar cytoplasmic constituents to the central vacuole.

Abstract: Publisher Summary This chapter describes methods to isolate mitochondria from leaf tissue of three different photosynthetic groups of higher plants: C3, C4, and Crassulacean acid metabolism (CAM) plants. In isolating mitochondria from the leaf mesophyll cells of C3 plants, photosynthetic tissue in which atmospheric CO2 is directly fixed through the reductive pentose phosphate pathway (with the 3-C phosphoglyceric acid as product). In C4 plants, CO2 assimilation occurs through two photosynthetic cell types, the mesophyll and the bundle sheath. Atmospheric CO2 is initially fixed into C4 dicarboxylic acids in the mesophyll cells; the C4 acids then serve as donors of carbon to the reductive pentose phosphate pathway in bundle sheath cells. In CAM plants, CO2 is fixed in the dark through phosphoenolpyruvate (PEP) carboxylase and malic acid is formed and utilized as a carbon donor to the reductive pentose phosphate pathway in the subsequent light period.

Abstract: Alopecurus gerardi, Poa alpina, and Carex curvula are spontaneous, perennial forage plants distributed in the high elevation (2300–3200 m) pasture lands of Piedmont and Valle d'Aosta (Italy). Sedum atratum is an annual succulent which grows at elevations up to 3200 m. The three monocotyledons have, in comparison with corresponding plants from the low-land, peculiar organographic and anatomic structures such as curling leaf lamina, vascular bundle sheath layer with chloroplasts arranged in a centrifugal fashion, low stomatal density (stomata number/cm2: upper epidermis x= 0–11.9; lower epidermis x= 7.66–11.55), thick cuticles, as well as higher values of S (H2O; g/dm2; x= 0.6–0.32), Sm (H2O g/mg Chi.; x= 0.11–0.16), SLW (g f. wt/dm2; x= 0.86–1.36), but lower SLA values (cm2/g f. wt; x= 75.07–116.77). All these data are correlated to water stress. Even though the leaf lamina anatomy possesses some features typical of C4 plants, the presence of starch grains in the mesophyll chloroplasts indicates that these plants are probably C3 ones. In spite of high values of thylacoid grana/thylacoid intergrana ratios, typical of shade plants (mesophyll chloroplasts: x up to 3.81; bundle sheath chloroplasts: x up to 5.3), and Chi a/Chl b ratios (x up to 4.23 in C. curvula), the apparent absence of peroxisomes seem to indicate a very efficient dark phase of photosynthesis. S. atratum, in comparison with the typical CAM succulents, which live in dryer and warmer habitats, has a higher values of stomatal density (upper epidermis, x= 2.59; lower epidermis, x= 3.15) and of SLA (x= 24.98), but lower S (x= 3.83), Sm (x= 1.19) and SLW (x= 4.15).

Abstract: The main vascular connection observed in the fruits of melon, cucumber and Luffa acutangula Roxb. were as follows; peduncle-receptacle-pericarp (exocarp-mesocarp-endocarp)-septum-fruit axis-placenta-seed (ovule). Some vascular bundles were also observed to reach the fruit axis directly from the peduncle, although they were smaller in number and diameter than those running through the receptacle.The basic number of vascular bundles in each part was 10 each in the peduncle, receptacle and exocarp, 1 per carpel in the fruit axis and 1 per ovule in the placenta. Many vascular bundles were observed in the mesocarp and endocarp, but their basic number could not be determined.The direction of vascular bundles in each part was as follows; acropetal in the peduncle, fruit axis, receptacle, exocarp and endocarp, tangential in the mesocarp, centripetal in the septum, and centrifugal in the placenta.There were usually 5 carpels in ‘Wasada-uri’ melon although the number sometimes decreased to 4 or 3 at high night temperatures. In ovaries consisting of 5 carpels, each carpet corresponded to 2 of vascular bundles of receptacles, but in ovaries consisting of 3, 4 or 6 carpels there was no such clear correspondence.In ‘Wasada-uri’ ovaries consisting of 3, 4 and 5 carpels, 4, 8 and 5 types of carpel arrangement, respectively, were observed in relation to petals and calyxes. Among these, the typical ones were III1, IV1 and IV3 and V2 and V4 (refer to Table 2-4).Assuming that the number of carpels increased from 1 to 5 in ontogenetic differentiation as leaves or floral leaves, its sequence and arrangement seemed to be I, II1, III1, IV1 and IV3, V2 and V4 (refer to Fig. 12).From these results, it was suggested that the carpels differ congenitally in time and hence in relative position in their ontogenetic development, which consequently results in differential growth of respective carpels and respective sides of ovary and fruit.

Abstract: Thorpe, M. R. and Minchin, P. E. H. 1987. Effects of anoxia on phloem loading in C3 and C4 species.—J. exp. Bot. 38: 221-232. Changes in phloem loading rate were inferred from observations of1 'C export from attached leaves of C3 and C4 monocots and dicots. Loading decreased under anoxia in C3 leaves, but not in general in the leaves of either C4 monocots or dicots in the light. However, loading rate in the C4 leaves did reduce if the leaf was also darkened or received no C02. We suggest that insensitivity to anoxia in C4 leaves is due to oxygen from photosynthesis which is retained in the bundle sheath at a concentration sufficient to energize a phloem loading system. There may also be another system which is insensitive to anoxia since the effects of shade and C02 deprivation were not always seen, and loading was not

Abstract: Photosystem II (PS II) activity and the localization of ribulose-l,5-bisphosphate (RuBP) carboxylase (EC 4.1.1.39) were studied in primary leaves of young maize plants (Zea mays L. cv. Fronica) by tetra-nitro-blue-tetrazoliumchloride reduction and immunolocalization, respectively. In tissue of 3-day-old plants all chloroplasts were structurally identical. From day 4 they developed into their typical appearance of mesophyll and bundle sheath chloroplasts. First PS II-activity was present in both types of chloroplasts. From day 4 it disappeared in bundle sheath chloroplasts concomitant with the loss of grana. RuBP carboxylase on the other hand was only present in bundle sheath chloroplasts at all stages of development. Thus, the control of the development of the photosystems and the Calvin cycle enzymes seem to differ.

Abstract: It is well known that PSII activity in maize bundle sheath chloroplasts is much lower than in mesophyll chloroplasts (1,2). Therefore, an input of PSII electron transport can not provide adequate level of ATP and NADPH for reductive carbon metabolism (3,4). Up to date it is postulated that the requirement for ATP in bundle sheath chloroplasts is mainly met by cyclic photophosphorylation maintained by PSI activity (5). It is also postulated that malic acid via decarboxylation servs as CO2 donor and as a reducing agent for NADP+ and consequently for PSI electron transport (6).

Abstract: C4 plants are generally characterized by a spatial separation of the C4 and C4 modes of carbon fixation into bundle sheath cells and mesophyll cells, respectively. In contrast Cam plants typically exhibit a temporal separation of the C3 and C4 modes with initial carbon fixation by the C, pathway at night and decarboxylation of the C4 products and refixation or the released CO2 via the reductive pentose phosphate pathway during the day.

Abstract: This study was carried out to investigate the anatomical characteristics, the radial and axial variation of the cell dimension and bulk density for four bamboo species used as the major material of bamboo products in Korea; Phyllostachys(P.bambusoides, P.edulis, P.nigra var.henonis, and Pnigra). The results obtained were summarized as follows; 1) In the outer part of culm, the mophological difference of cell diameter and distribution were not acknowledged. However, gum-like substance was occurred In the epidermis, hypodermis, cortex and metaxylem vessel of P.nigra, while it was occured only in the metaxylem vessel of P.edulis. 2) The protoxylem vessel in the central zone of culm-wall could be in the two types; the first type consisted of 4-5 small cell pieces occurred in P.edulis and P.nigra var. henonis, the second type of 1-2 large cell pieces in P.bambusoides and P.nigra. And transition of cell diameter from the vascular bundle sheath to the ground tissue was gradually enlarged in P.nigra and P.edulis but abruptly in P.bambusoides and P.nigra var. henonis. 3) Thin-wall tyloses were well developed in the outer and central zone of culm-wall in four species. 4) Compared with the central part of culm-wall, the number of parenchyma cells and the amount of the vascular bundle sheaths per unit area were a few in the inner part. The inner part nearest to the pith cavity was consisted of thick-well parenchyma cell. 5) The dimensional variations of metaxylem vessel and parenchyma cell in the radial direction were decreased from the inner part to the epidermis in all species observed. 6) The fiber length was the maximum in the central zone and its diameter was increased from the inner part to the outer part. In the axial variation. fiber length was slightly increased from the base and then decresed slowly toward the top, and its diameter was reverse. 7) The axial variation of the bulk density was continuously increased from the base toward the top and its radial variation was rapidly increased from the inner part to the epidermis.

Abstract: Synthesis of starch in leaves of Zea mays, an NADP-ME type C4 plant, is normally restricted to the bundle sheath cells (1). The absence of starch accumulation in maize mesophyll may be related to the distribution of starch metabolizing enzymes between the bundle sheath and mesophyll cells, although this has not been convincingly established (1–3). The stromal enzyme, ADPglucose pyrophosphorylase catalyzes the formation of ADPglucose, the glucosyl donor for starch synthase, and is believed to be the principal site of regulation for starch biosynthesis. This enzyme from all plants studied to date is activated by 3-phosphoglycerate (3-PGA) and inhibited by orthophosphate (Pi) (14). The rate of starch synthesis is believed to be coupled to the rate of photosynthesis and to the rate of photosynthate export from the cell through the interaction of this enzyme and the stromal ratio of 3-PGA/Pi. Activity of this enzyme from whole leaf extracts of sorghum, also a NADH-ME type C4 plant, is also allosterically regulated by 3-PGA and Pi (5). However, significance of the kinetic properties of bundle sheath and mesophyll specific ADPglucose pyrophosphorylases to localization of starch synthesis in leaves of maize or other C4 plants has not been previously investigated.

Abstract: A histochemical method for the determination of IAA-oxidase has been used in sections of various aerial parts of winter wheat plants. High IAA-oxidase activity was localized in the cell walls of sclerenchyma near the periphery of the stem, in the vascular bundle sheath of sclerenchyma and in xylem, both in the stem and in the leaf. The cell wall—bound IAA-oxidase activity therefore appeared in lignifying tissues. The staining was very weak or absent in the cell walls of parenchyma tissues and phloem. The positive reaction of the cytosol at the bulbous ends of guard cells and in the leaf primordia is presumed to be due to cytosolic IAA-oxidase. These results are discussed in relation to peroxidase localization and to our previousin vitro studies.

Abstract: Catalase is an abundant enzyme in the peroxisomes of leaves of C3 plants (1), where it plays a vital role in decomposition of H2O2 arising from the oxidation of photorespiratory glycolate. Although the leaves of some C3 plants appear to possess only one form of catalase (2, 3), multiple forms of leaf catalase have been reported in various C3 (4, 5, 6) and C4 (7) plants. Two of the catalase forms in maize leaves exhibit cell-type specificity, one residing in the cells of the bundle sheath, and the other in the mesophyll (8). Since photorespiration in maize would be confined to the bundle sheath, it appears that the mesophyll-specific catalase isozyme must perform some other function. The physiological significance of multiple forms of catalase in C3 leaves remains unclear. We have undertaken a characterization of leaf catalase in several C3 plants, with particular emphasis on Nicotiana, where multiple forms of the enzyme undergo marked changes during seedling development and major shifts in relative distribution in response to manipulation of photorespiratory activity.

Abstract: In vitro translation of polyA+ mRNAs isolated from purified maize bundle sheath and mesophyll cells results in the production of distinctive, cell-specific polypeptides. Immunoprecipitation experiments show that translatable polyA+ mRNAs for phosphoenolpyruvate carboxylase (PEPC), pyruvate orthophosphate dikinase (PPDK) and NADP-malate dehydrogenase (MDH) are prominent in mesophyll but not bundle sheath cells. On the contrary, those for sedoheptulose-1,7-bisphosphatase (SBP), fructose-1,6-bisphosphatase (FBP), NADP-malic enzyme (ME) and the small subunit of ribulose-1,5-bisphosphate carboxylase (RuBPC SS) are present only in bundle sheath cells. Moreover, polyA+ mRNAs encoding the 33 kD, 23 kD and 16 kD polypeptides of the oxygen-evolving complex (OE33, OE23 and OE16) and the light-harvesting chlorophyll a/b binding protein of photosystem II (LHCP II) are much more abundant in mesophyll than in bundle sheath cells. Northern blot analyses with cDNA clones of PEPC, PPDK, ME, RuBPC SS, OE33, OE23, OE16 and LHCP II are consistent with the conclusion that the cell-specific expression of these genes is regulated at the RNA level. The RNA level differences are especially dramatic in dark-grown maize seedlings after illumination for 24 h.