Membrane potentials and ion permeabilities in flexor cells of the laminar pulvini of Phaseolus coccineus L.

The internal potential of flexor cells in slices of the laminar pulvini of Phaseolus coccineus has been recorded by standard microelectrode techniques in 100 eq m^-3 external salt solutions of various ionic compositions. The measured values are between-15 and-60 mV depending on the external medium. Treating the results with the Goldman equation yields the following relative permeabilities: K⁺, 1.00; Na⁺, 0.24; Cl^-, 0.19; NO ₃ ^- , 1.6. The membrane potential was only slightly sensitive to external pH and Ca²⁺. Metabolic inhibitors (azide, cyanide and salicylhydroxamic acid, carbonyl cyanid m-chlorphenyl hydrazone) caused only slight depolarizations (ca. 4 mV), which differed from the ion-induced changes by their slow time courses. The results are consistent with the hypothesis that the relatively impermeable Cl^- is actively transported and osmotically efficient, whereas the well-permeable K⁺ passively follows Cl^- to maintain electroneutrality and is osmotically of only minor significance.Abbreviations SHAM salicylhydroxamic acid - CCCP carbonyl cyanid m-chlorphenyl hydrazone     

Effects of white,blue, red light and darkness on pH of the apoplast in the Samanea pulvinus

Leaflet movements in Samanea saman (Jacq.) Merrill are driven by fluxes of K⁺, anions, and water through membranes of motor cells in the pulvinus (R.L. Satter et al., 1974, J. Gen. Physiol. 64, 413–430). Extensor cells take up K⁺ and swell in white light (WL) while flexor cells take up K⁺ and swell in darkness (D). Excised strips of extensor and flexor motor tissue acidify their bathing medium under conditions that normally promote increase in K⁺ in the intact tissue, and alkalize the medium under conditions that normally induce decrease in K⁺ (A. Iglesias and R.L. Satter, 1983, Plant Physiol. 72, 564). To obtain information on pH changes in the whole pulvinus, we measured effects of light on pH of the apoplast, using liquid membrane microelectrodes sensitive to H⁺. We report the following: (1) The pH of the extensor apoplast was higher than that of the flexor apoplast in WL and in D (pH gradient of 1.0 units in WL and 2.0 units in D). Apoplastic pH might affect K⁺ transport through the plasma membranes of Samanea motor cells, since the conductance, gating, and selectivity of ionic channels in other systems depend upon external pH. (2) Extensor cells acidified and flexor cells alkalized their environment in response to irradiation with WL, while the reverse changes occurred in response to D. These results are consistent with the results of Iglesias and Satter (1983), and support the physiological relevance of data obtained with excised tissue. (3) The pH changes in response to irradiation with red light were similar to those obtained with D; also, the pH changes in response to blue light were similar to those obtained with WL. The pulvinus closed in red light as in darkness and opened in WL, but failed to open in blue light. The advantages and limitations of apoplastic pH measurements for assaying H⁺ transport are discussed.Abbreviations BL blue light - D darkness - RL red light - WL white light     

Cell walls as reservoirs of potassium ions for reversible volume changes of pulvinar motor cells during rhythmic leaf movements

The laminar pulvinus of primary leaves of Phaseolus coccineus L. was investigated with respect to the total K⁺ content, the apoplastic K⁺ content, and the water potential of extensor and flexor sections in relation to the leaf positions in a circadian leaf-movement cycle, as well as the cation-exchange properties of isolated extensor- and flexor-cell walls. Turgid tissue showed a high total but low apoplastic K⁺ content, shrunken tissue a low total but high apoplastic K⁺ content. Thus, part of the K⁺ transported into and out of the swelling or shrinking protoplasts is shuttled between the protoplasts and the surrounding walls, another part between different regions of the pulvinus. The K⁺ fraction shuttled between protoplasts and walls was found to be 30–40% of the total transported K⁺ fraction. Furthermore, 15–20% of the total K⁺ content of the tissue is located in the apoplast when the apoplastic reservoir is filled, 5–10% when the apoplastic reservoir is depleted. The ion-exchange properties of walls of extensor and flexor cells appear identical in situ and in isolated preparations. The walls behave as cation exchangers of hhe weak-acid type with a strong dependence of the activity of fixed negative charges as well as of the K⁺-storing capacity on pH and [K⁺] of the equilibration solution. The high apoplastic K⁺ contents of freshly cut tissues reflect the cation-storing capacity of the isolated walls. We suggest that K⁺ ions of the Donnan free space are used for the reversible volume changes (mediating the leaf movement) mainly by an electrogenic proton pump which changes the pH and-or the [K⁺] in the water free space of the apoplast.Abbreviations and symbols DFS Donnan free space - DW dry weight - pK negative logarithm of the equilibrium constant K of the acidic group - WFS water free space - water potential; Indices - cw cell wall - t tissue     

Role of phloem in sucrose transport by Ricinus cotyledons

Sucrose is taken up and accumulated by cotyledons of Ricinus communis L. Autoradiographic studies reveal a predominant accumulation of sucrose in the phloem of the cotyledons. The export of sucrose from the cotyledons to hypocotyl and roots proceeds in the phloem by mass flow. These results, taken together with previous data, are experimental evidence for proton-sucrose symport as the mechanism of phloem loading.     

Inositol 1,4,5-trisphosphate may mediate closure of K+ channels by light and darkness in Samanea saman motor cells

Leaflet movements of Samanea saman (Jacq.) Merr. depend in part upon circadian-rhythmic, light-regulated K⁺ fluxes across the plasma membranes of extensor and flexor cells in opposing regions of the leaf-moving organ, the pulvinus. We previously showed that blue light appears to close open K⁺ channels in flexor protoplasts during the dark period (subjective night) (Kim et al., 1992, Plant Physiol 99: 1532–1539). In contrast, transfer to darkness apparently closes open K⁺ channels in extensor protoplasts during the light period (subjective day) (Kim et al., 1993, Science 260: 960–962). We now report that both these channel-closing stimuli increase inositol 1,4,5-trisphosphate [Ins(1,4,5)P₃] levels in the appropriate protoplasts. If extensor cells are given a pulse of red light followed by transfer to darkness, channels still apparently close (Kim et al. 1993) but changes in Ins(1,4,5)P₃ levels are complex with an initial decrease under red light followed by accumulation. Neomycin, an inhibitor of polyphosphoinositide hydrolysis, inhibits both blue-light-induced Ins(1,4,5)P₃ production and K⁺-channel closure in flexor protoplasts and both dark-induced Ins(1,4,5)P₃ production and K⁺ channel closure in extensor protoplasts. The G-protein activator, mastoparan, mimics blue light and darkness in that it both increases Ins(1,4,5)P₃ levels and closes K⁺ channels in the appropriate cell type at the appropriate time. These results indicate that phospholipase C-catalyzed hydrolysis of phosphoinositides, possibly activated by a G protein, is an early step in the signal-transduction pathway by which blue light and darkness close K⁺ channels in S. saman pulvinar cells.Abbreviations DiS-C₃-(5) 3,3-dipropylthiadicarbocyanine iodide - F measure change in Dis-C₃-(5) fluorescence - F_o initial Dis-C₃-(5) fluorescence - Ins(1,4,5)P₃ inositol 1,4,5-trisphosphate - PtdIns(4,5)P₂ phosphatidylinositol 4,5-bisphosphate - rbc red blood cell Supported by grants from NSF (IBN 9206179 and MCB 9305154) and U.S.-Israel Binational Agricultural Research and Development Fund (IS-1670-90RC) to R.C.C. We thank the University of Connecticut Biotechnology Center for the use of a fluorescent spectrophotometer.     

Isolation of functional extensor and flexor protoplasts fromPhaseolus coccineus L. pulvini: potassium induced swelling

Methods are described for the isolation of functional protoplasts from secondary pulvinus tissue (flexor and extensor) and from leaf mesophyll tissue of primary leaves ofPhaseolus coccineus L. Integrity of the protoplasts was shown by vital staining and their ability to evolve oxygen in the light. Extensor-cell protoplasts increased their volume for up to 60% upon addition of 10 mM KCl. This K⁺-induced swelling was accompanied by increased rates of proton extrusion.     

Efflux of sucrose from minor veins of tobacco leaves

Mature leaves import limited amounts of nutrient when darkened for prolonged periods. We tested the hypothesis that import is restricted by the apoplast-phloem loading mechanism, ie., as sucrose exits the phloem of minor veins it is retrieved by the same tissue, thus depriving the mesophyll of nutrient. When single, attached, mature leaves of tobacco (Nicotiana tabacum L.) plants were darkened, starch disappeared from the mesophyll cells, indicating that the supply of solute to the mesophyll was limited. Starch was synthesized in mesophyll cells of darkened tissue when sucrose was applied to the apoplast at 0.1–0.3 mM concentration. Efflux from minor veins was studied by incubating leaf discs on [¹⁴C]sucrose to load the minor veins and then measuring subsequent ¹⁴C release. Efflux was rapid for the first hour and continued at a gradually decreasing rate for over 13 h. Net efflux increased when loading was inhibited by p-chloromercuribenzene-sulfonic acid, anoxia, isotope-trapping, or reduction of the pH gradient. Neither light nor potassium had a significant effect on the rate of labeled sucrose release. The site of labeled sucrose release was investigated by measuring efflux from discs in which sucrose had previously been loaded preferentially by either the minor veins or mesophyll cells. Efflux occurred primarily from minor veins.Abbreviations Mes 2(N-morpholino)ethanesulfonic acid - Mops 3(N-morpholino)propanesulfonic acid - PCMBS p-chloromercuribenzenesulfonic acid - SE-CC sieve element-companion cell complex     

Compartmentation of soluble carbohydrates,of starch and of malate in motor organs (pulvini) and other parts of Phaseolus coccineus L. leaves

Quantitative histochemistry was used to investigate the tissue-specific compartmentation of soluble carbohydrates (sucrose, glucose, fructose), starch and malate in the laminar pulvinus, leaf blade and petiole of Phaselous coccineus L. at day and night positions of diurnal leaf movement. Total carbohydrate levels measured in a series of cross sections along individual pulvini of 24-d-old plants showed only small differences between the day and night positions of the respective leaf. In contrast, the level of malate changed during diurnal leaf movement, especially in the central part of a pulvinus. The levels of glucose and fructose in the pulvinus increased towards the transition zones between the pulvinus and lamina, and pulvinus and petiole, and this trend was even more pronounced for starch. By contrast, sucrose levels were highest in the pulvinus proper. The transverse compartmentation of metabolites was studied in distinct, approx. 0.5-mm-thick tissue slices from the central part of a pulvinus. These were dissected further into up to 14 distinct subsamples (bundle, bundle sheath, motor tissues, flanks). Irrespective of the position of the leaf (day or night), the central vascular core and the surrounding bundle sheath had high levels of sucrose (up to 500 mmol-(kg DW)^–1) and low levels of glucose and fructose (below 100 mmol-(kg DW)^–1), while in the cortex the situation was reversed. In the night position the level of sucrose decreased by approx. 30% in the bundle sheath and the central vascular core but not in the other sections. We thus suggest that because of the relatively small diurnal changes in their cortical pools, soluble sugars are not involved in the osmotic processes resulting in leaf movement. In contrast, pulvini from 14-d-old plants showed an interesting diurnal change in starch and malate pools in the outermost layer of the extensor. Here starch increased at night while the malate pool was lowered nearly stoichiometrically. Inverse pool sizes were found in the day position of the respective leaves. Although less significant, the opposite diurnal variation occurred in samples taken from the flexor region. We thus were able to locate areas of different carbohydrate activities in the laminar pulvinus of P. coccineus. The central vascular core, including the bundle sheath, is involved in temporary storage of photoassimilates, and the cortical regions are responsible for osmotically driven leaf movement. The results are discussed with respect to guard-cell physiology.Abbreviations CLP cut-leaf pulvini - ILP intact-leaf pulvini This work was supported by a grant from the Deutsche Forschungsgemeinschaft.     

Uptake of [14C]sucrose in isolated minor-vein networks of Commelina benghalensis L

Maceration with pectinase (4.5h) of Commelina benghalensis L. leaves stripped at either side yielded isolated vein networks consisting of four to five secondary veins and tertiary cross veins (=minor veins). Examination with Evans Blue and injection of Fluorescein F showed that 80% of the veins were viable. Proof of normal functioning of isolated minor veins was that [¹⁴C]sucrose fed to an apical vein network attached to the remaining intact part of the leaf was absorbed and finally arrived in the petiole. Sucrose uptake by veins obeyed Michaelis-Menten kinetics (K _m 5·10^-4 mol l^-1; V _max (light) 3.2 mol h^-1 g^-1 fresh weight, V _max (dark) 1.5 mol h^-1 g^-1 fresh weight). A linear component, not inhibited by carbonylcyanide m-chlorophenylhydrazone and p-chloromercuribenzenesulfonic acid, was present. Maximal uptake took place at 5 mmol l^-1 K⁺; concentrations of K⁺ higher than 10 mmol l^-1 decreased the rate of uptake. The uptake rates by isolated veins and veins in situ (in disks) were in the same order of magnitude. Altogether, isolated veins promise to be a useful system for the study of loading.Abbreviations CCCP carbonylcyanide m-chlorophenylhydrazone - EDTA ethylenediamine tetraacetic acid - PCMBS p-chloromercuribenzenesulfonic acid     

Evidence for the involvement of a UDP-glucose-dependent group translocator in sucrose uptake into vacuoles of storage roots of red beet

Vacuoles isolated from the storage roots of red beet (Beta vulgaris L.) accumulate sucrose via two different mechanisms. One mechanism transports sucrose directly, and its rate is increased by the addition of MgATP. The other mechanism utilizes uridine diphosphate glucose (UDP-glucose) to synthesize and simultaneously transport sucrose phosphate and sucrose into the vacuole. This group translocation mechanism has also been found in sugarcane vacuoles. As in sugarcane, the beet group translocator does not require fructose 6-phosphate, nor is the latter substance transported into the vacuole. The uptake of UDP[¹⁴C]glucose in inhibited by high concentrations of osmoticum.Abbreviations EDTA ethylenediaminetetraacetic acid - Hepes 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid - UDP uridine 5-diphosphate     

Intercellular compartmentation of basic carbon pathways in motor organs (pulvini) of leaves of Phaseolus coccineus L.

The activities of enzymes which catalyze one step in each of the five major carbon pathways in green plants were measured in secondary pulvini and other tissues of Phaseolus coccineus L. leaves. We were able to detect activities of fumarase (EC 4.2.1.2; tricarboxylic-acid pathway), NAD-glyceraldehyde-phosphate dehydrogenase (NAD-GAPDH, EC 1.2.1.12; glycolysis), 6-phosphogluconate dehydrogenase (6-PGDH, EC 1.1.1.44; oxidative pentose-phosphate pathway), ribulose-1, 5-bisphosphate carboxylase (Rubisco, EC 4.1.1.39; photosynthetic carbon-reduction pathway), and of hydroxypyruvate reductase (HP-R, EC 1.1.1.81; photosynthetic carbon-oxidation pathway). On a protein basis the activities of Rubisco and HP-R in pulvinar regions were very low (below 1 and 2 mol · (kg protein) ^–-1 · h^–-1, respectively), but the activities of fumarase and NAD-GAPDH were between 10- and 5-fold higher compared with the laminar tissue (up to 7 and 50 mol · (kg protein)^–-1 · h^–-1, respectively). Similarly, the protein specific activities of 6-PGDH were increased in the pulvinus (3–4 compared with approx. 1 mol · (kg protein)^–-1 · h^–-1 in the leaf blade). No differences in specific activities were detected between day and night positions of the leaves. By applying quantitative histochemical techniques we determined the longitudinal and transversal compartmentation of the activities of fumarase, NAD-GAPDH, and 6-PGDH in pulvinar tissues. Levels of activity of all three enzymes increased towards the middle part of the pulvinus. Here, expressed on a dry-weight (DW) basis, the analysis of cross sections showed highest activities in the outer parts of the extensor in the order given, approx. 0.6, 5, and 0.25 mol · (kg DW)^–-1 · h^–-1 for fumarase, NAD-GAPDH and 6-PGDH. When related to protein, levels of activity were comparably high within the inner parts of extensor and flexor, and partly also in the abaxial part of the bundle (fumarase, 6-PGDH). The tissue-specific compartmentation of the respective activities is discussed in relation to leaf movement and shows parallels with guard-cell function.Abbreviations Chl chlorophyll - DW dry weight - GAPDH glyceraldehyde-phosphate dehydrogenase - HP-R hydroxypyruvate reductase - Rubisco ribulose-1,5-bisphosphate carboxylase - 6-PGDH 6-phosphogluconate dehydrogenase This investigation was supported by a grant from the Deutsche Forschungsgemeinschaft.     

Mechanics of circadian pulvini movements in Phaseolus coccineus L.

The circadian movement of the lamina of primary leaves of Phaseolus coccineus L. is mediated by antagonistic changes in the length of the extensor and flexor cells of the laminar pulvinus. The cortex of the pulvinus is a concentric structure composed of hexagonal disc-like cells, arranged in longitudinal rows around the central stele. Observations with polarization optics indicate that the cellulose microfibrils are oriented in a hoop-like fashion in the longitudinal walls of the motor cells. This micellation is the structural basis of the anisotropic properties of the cells: tangential sections of the extensor and flexor placed in hypotonic mannitol solutions showed changes only in length. As a consequence a linear correlation between length and volume was found in these sections. Based on the relationship between the water potential (which is changed by different concentrations of mannitol) and the relative volume of the sections and on the osmotic pressure at 50% incipient plasmolysis, osmotic diagrams were constructed for extensor and flexor tissues (cut during night position of the pulvinus). The bulk moduli of extensibility, , were estimated from these diagrams. Under physiological conditions the values were rather low (in extensor tissue below 10 bar, in flexor tissue between 10 to 15 bar), indicating a high extensibility of the longitudinal walls of the motor cells. They are strongly dependent on the turgor pressure at the limits of the physiological pressure range.In well-watered plants, the water potentials of the extensor and flexor tissues were surprisingly low,-12 bar and-8 bar, respectively. This means that the cells in situ are by no means fully turgid. On the contrary, the cell volume in situ is similar to the volume at the point of incipient plasmolysis: the cell volumes of extensor and flexor cells in situ were only 1.01 times and 1.1 times larger, respectively, than at the point of incipient plasmolysis, whereas at full turgidity (cells in water) the corresponding factors were 1.8 and 1.5. It is suggested that the high elasticity of the longitudinal walls, the anisotropy of the cell walls, and the low water potential of the sections which is correlated with slightly stretched cell walls in situ, are favourable and effective for converting osmotic work in changes in length of the pulvinus cells, and thus for the up and down movement of the leaf.Symbols volumetric elastic modulus - _i instantaneous volumetric elastic modulus - _i stationary volumetric elastic modulus - weight-averaged stationary bulk modulus of extensibility - ₀ osmotic pressure of the vacuole of a cell at the point of incipient plasmolysis - weight-averaged osmotic pressure of the vacuoles of the tissue at 50% incipient plasmolysis - water potential     

Sucrose synthase catalyses a readily reversible reaction in vivo in developing potato tubers and other plant tissues

Experiments were carried out to investigate whether sucrose synthase (Susy) catalyses a readily reversible reaction in vivo in potato (Solanum tuberosum L.) tubers, Ricinus communis L. cotyledons, and heterotrophic Chenopodium rubrum L. cell-suspension cultures. (i) The contents of sucrose, fructose, UDP and UDP-glucose were measured and the mass-action ratio compared with the theoretical equilibrium constant. In all three tissues the values were similar. (ii) Evidence for rapid turnover of label in the sucrose pool was obtained in pulse-chase experiments with potato discs and with intact tubers attached to the plant. The unidirectional rates of sucrose synthesis and degradation were considerably higher than the net flux through the sucrose pool in the tubers. (iii) Labelling of the glucosyl and fructosyl moieties of sucrose from [¹⁴C]glucose in the presence of unlabelled fructose provided evidence that Susy contributes to the movement of label into sucrose. Methods for estimating the contribution of sucrose-phosphate synthase and Susy are presented and it is shown that their relative contribution varies. For example, the contribution of Susy is high in developing tubers and is negligible in harvested tubers which contain low Susy activity. (iv) The absolute values of the forward (v⁺¹) and backward (v^?1) reaction direction of Susy are calculated from the kinetic labelling data. The estimated values of v⁺¹ and v^?1 are comparable, and much higher than the net flux through the sucrose pool. (v) The estimated concentrations of the substrates and products of Susy in tubers are comparable to the published K _m values for potato-tuber Susy. (vi) It is concluded that Susy catalyses a readily reversible reaction in vivo and the relevance of this conclusion is discussed with respect to the regulation of sucrose breakdown and the role of Susy in phloem unloading.     

Orcokinin immunoreactivity in the accessory medulla of the cockroach Leucophaea maderae

The accessory medulla is the master circadian clock in the brain of the cockroach Leucophaea maderae and controls circadian locomotor activity. Previous studies have demonstrated that a variety of neuropeptides are prominent neuromediators in this brain area. Recently, members of the orcokinin family of crustacean neuropeptides have been identified in several insect species and shown to be widely distributed in the brain, including the accessory medulla. To investigate the possible involvement of orcokinins in circadian clock function, we have analyzed the distribution of orcokinin immunostaining in the accessory medulla of L. maderae in detail. The accessory medulla is densely innervated by approximately 30 orcokinin-immunoreactive neurons with cell bodies distributed in five of six established cell groups in the accessory medulla. Immunostaining is particularly prominent in three ventromedian neurons. These neurons have processes in a median layer of the medulla and in the internodular neuropil of the accessory medulla and send axonal fibers via the posterior optic commissure to their contralateral counterparts. Double-labeling experiments have revealed the colocalization of orcokinin immunostaining with immunoreactivity for pigment-dispersing hormone, FMRFamide, Mas-allatotropin, and γ-aminobutyric acid in two cell groups of the accessory medulla, but not in the ventromedian neurons or in the anterior and posterior optic commissure. Immunostaining in the ventromedian neurons suggests that orcokinin-related peptides play a role in the heterolateral transmission of photic input to the pacemaker and/or in the coupling of the bilateral pacemakers of the cockroach.This study was supported by the Deutsche Forschungsgemeinschaft, grant HO 950/9.     

蔗糖对列当科两种根部半寄生植物吸器发生的促进作用

吸器是寄生植物的特征器官,研究影响其发生的因素,有助于了解寄生关系的建立和调控过程。该研究以两种列当科(Orobanchaceae)根部半寄生植物甘肃马先蒿(Pedicularis kansuensis)和松蒿(Phtheirospermum japonicum)为材料,通过皿内培养试验,分析了蔗糖、DMBQ(2,6-二甲氧基-对-苯醌,一种高效的列当科根部半寄生植物吸器诱导化合物)和寄主植物诱导下两种根部半寄生植物吸器发生情况。结果表明：(1)蔗糖显著促进两种根部半寄生植物吸器发生,无寄主存在时,2%蔗糖处理使甘肃马先蒿和松蒿吸器发生率分别提高39.9%和20.2%。(2)蔗糖明显提升寄主植物对两种根部半寄生植物的吸器诱导水平,添加蔗糖后,寄主诱导的甘肃马先蒿单株吸器数和具木质桥的吸器比例分别增加5.7个/株和17.9%,松蒿吸器发生率和具木质桥的吸器比例分别提升76.7%和16.2%。(3)蔗糖对松蒿吸器发生的促进作用与已知吸器诱导化合物DMBQ相当,均能诱导50%以上的植株产生吸器。(4)培养基中添加4%蔗糖对两种根部半寄生植物的吸器诱导效果最好,其中甘肃马先蒿吸器发生率为56%...     

Microautoradiographic studies of the role of mesophyll and bundle tissues of the Ricinus cotyledon in sucrose uptake

The aim of the study was to show which tissues and cell types of the cotyledon of Ricinus communis L. are responsible for uptake of sucrose by H⁺-sucrose symport. The cotyledons were incubated in labelled sucrose for up to 20 min and then the amount of radioactivity in each cell type of the cotyledon was assessed by microautoradiography. It was found that 50% of the label was present in the spongy mesophyll, and 10–15% was in the bundles, the epidermal layers and the palisade parenchyma. The sieve tubes contained only 2–3% of the label. The addition of sucrose to cotyledons depolarized the membrane of spongy-mesophyll cells by 33 mV. Therefore, it was concluded that the previously found H⁺-sucrose symport is at least partly located at the spongy mesophyll. No precursor-like behaviour of the label in mesophyll or bundle-sheath cells was observed in pulse-chase experiments, which indicates a direct uptake of sucrose by the sieve tube-companion cell complex from the apoplast.This work was funded by Deutsche Forschungsgemeinschaft. The valuable help by Ina Möller, Elke Schmidt, Christian Schobert (all from Bayreuth, FRG), Dr. Dieter Gradmann (Göttingen, FRG), Dr. Jörg Tittor (MPI Biochemie München, FRG), Dr. Pavlovkin (Plant Pathology, Bratislava), Dr. K. Köhler (Botany Department Würzburg, FRG) and the intense discussions with Dr. Enno Brinkmann (Bayreuth) are gratefully acknowledged. The technical assistance by Beatrix Tannhäuser-Hofmann and Hildegard Stork was of great help for this work.     

Intercellular compartmentation of sucrose synthesis in leaves of Zea mays L.

The incorporation of ¹⁴C into sucrose and hexose phosphates during steady-state photosynthesis was examined in intact leaves of Zea mays L. plants. The compartmentation of sucrose synthesis between the bundle sheath and mesophyll cells was determined by the rapid fractionation of the mesophyll and comparison of the labelled sucrose in this compartment with that in a complete leaf after homogenisation. From these experiments it was concluded that the majority of sucrose synthesis occurred in the mesophyll cell type (almost 100% when the time-course of sucrose synthesis was extrapolated to the time of ¹⁴C-pulsing). The distribution of enzymes involved in sucrose synthesis between the two cell types indicated that sucrose-phosphate synthetase was predominantly located in the mesophyll, as was cytosolic (neutral) fructose-1,6-bisphosphatase activity. Stromal (alkaline) fructose-1,6-bisphosphatase activity was found almost exclusively in the bundle-sheath cells. No starch was found in the mesophyll tissue. These data indicate that in Zea mays starch and sucrose synthesis are spatially, separated with sucrose synthesis occurring in the mesophyll compartment and starch synthesis in the bundle sheath.     

Propagated fluctuations of the electric potential in the apoplasm of Lepidium sativum L. roots

The electric potential on the surface of the Lepidium sativum L. root apex was recorded by means of six non-polarizable electrodes. Nonevoked fluctuations of the potential with amplitudes below 0.1 mV were observed. The fluctuations could be reversibly inhibited either by ether vapor or by anoxia caused by N₂. They did not occur in killed roots. Cross-correlation analysis of the fluctuations from six electrodes located one above another along the 3-mm apical region showed a pattern of time delay which indicates that the fluctuations may be the consequence of signals propagated in the root with a velocity of 3–9 mm · s^–1 in a basipetal direction from the root cap. We hypothesize that the fluctuations are due to signals of an unknown nature propagated along an intrasymplasmic continuous system, the symreticulum, composed of the cortical ER of individual cells and desmotubules passing through the plasmodesmata.Abbreviations AC alternating current - AP action potential - ACF autocorrelation function - CCF cross-correlation function - DC direct current - EEP extracellular electric potential This research was supported by Bundesminister für Forschung und Technologie, Bonn, and Ministerium für Wissenschaft und Forschung, Düsseldorf, (AGRAVIS). We are grateful to Mr. Dipl.-Ing. P. Blasczyk for constructing the amplifiers and for advice in instrumentation, and to Mr. H. Laubach for constructing the mechanical assembly.     

Evidence for proton/sulfate cotransport and its kinetics inLemna gibba G1

Sulfate uptake into duckweed (Lemna gibba G1) was studied by means of [³⁵S]sulfate influx and measurements of electrical membrane potential. Uptake was strongly regulated by the intracellular content of soluble sulfate. At the onset of sulfate uptake the membrane potential was transiently depolarized. Fusicoccin stimulated uptake up to 165% of the control even at pH 8. It is suggested that sulfate uptake is energized in the whole pH range by a 3H⁺/sulfate cotransport mechanism. Kinetics of sulfate uptake and sulfate-induced membrane depolarization in the concentration range of 5 M to 1 mM sulfate at pH 5.7 was best described by two Michaelis-Menten terms without any linear component. The second system had a lower affinity for sulfate and was fully active only at sufficiently high proton concentrations.Abbreviations c _o extracellular sulfate concentration - c _i intracellular sulfate concentration - E _m electrical membrane potential difference - E _m sulfate-induced, maximal membrane depolarization - electrochemical proton gradient - FW fresh weight