Euglenoid movement inEuglena fusca: Evidence for sliding between pellicular strips

Summary InEuglena fusca, each pellicular strip carries a row of particles on its surface. The relative displacement of particles on adjacent strips was analysed by video-microscopy and evidence was obtained that adjacent pellicular strips slide relative to each other during euglenoid movement.E. fusca shows two types of euglenoid movement, oscillatory bending and rounding-up of the cell body. During oscillatory bending, the maximum velocity of sliding was 0.4 m/s and the maximum displacement distance between adjacent strips 2.3 m about their mean position. WhenE. fusca exhibited rounding-up of the cell body, particle displacement again occurred and the angle of the pellicular strips to the long axis of the cell body increased because of pellicular sliding. As a result the distance between the cell's anterior and posterior tips was reduced. There was no change in distance either between rows of particles or between particles within the same row. The findings are incompatible with theories of euglenoid movement requiring local contraction of pellicular strips and point to the likely existence of active sliding between adjacent strips.     

Pellicle structure in Euglena

The pellicle of Euglena has been investigated by anoptral and phase contrast light microscopy, and by electron microscopy of osmium-fixed, Epon-embedded, lead-stained sections and of carbon/platinum replicas.

Observations on living cells show that the pellicular striatiors of Euglena spirogyra trace a left-handed (S) helix in a majority of cells, and a right-handed (Z) helix in only 5 to 30% of the cells in any one culture. All cells of E. spirogyra var. fusca have a left-handed (S) helix. Ornamentation on the pellicle takes the form of rows of knobs in various patterns. The living pellicle can be dissociated into long flat strips.

Electron microscopy shows that each pellicular strip has an elaborate cross-sectional shape, features of which are accessory teeth and ribs and a continuous ridge which articulates in a groove running along the edge of the next strip. The strips can move against one another, presumably by the ridges sliding in the grooves, and it is suggested that the joints might be lubricated by mucilage supplied from the helically disposed muciferous bodies. One single and one pair of fibrils or tubes, 200–250 Å in diameter, are regularly arranged parallel to each pellicular strip. A continuous tripartite plasmalemma, 80–100 Å thick, lies externally to the strips; external to this membrane lie the pellicular knobs. Each cell has from 35 to 45 pellicular strips, reducing to a few at the posterior end of the cell by successive fusions. Similar fusions occur at the anterior end of the cell, mainly within the canal.

These observations are compared with those made on the euglenoid pellicle by previous authors, and the following problems are discussed: direction of helix; the nature and cause of ornamentation; euglenoid movement with reference to fibrils, cytoplasmic flow, pellicle flexibility and the proteinaceous nature of the pellicle; helical and bilateral symmetry in the cell; and cet growth and division.     

On the mechanism of trap closure of Venus flytrap (Dionaea muscipula Ellis)

The rapid trap closure of Dionaea muscinula Ellis has been explained by either a loss of turgor pressure of the upper epidermis, which should thus become flexible, or by a sudden acid-induced wall loosening of the motor cells. According to our experiments both explanations are doubtful. Objections against the turgor mechanism come from the determination by extracellular measurements from the upper epidermis of action-potential amplitudes before and after trap closure. Neither time course nor amplitude of the action potentials are altered by trap closure. In contrast a rise in the apoplastic concentration of K⁺ or Na⁺, which are the only ions present in the trap in osmotically significant concentrations, from 1 to 10 mM reduces the action-potential amplitudes by 25% and 15%, respectively. Furthermore, after trap closure the upper epidermal cells retain a considerable cell sap osmolality of 0.41 mol·kg^-1 which equals that of the mesophyll cells as determined by incipient plasmolysis. A sudden cell-wall acidification causing movement is improbable since an acidification of the apoplast from pH 6 to pH 4 reduces action-potential amplitudes by 33% whereas the amplitudes measured extracellylarly from the mesophyll and lower epidermis remain unchanged by trap closure. In addition, buffering the apoplast at pH 6 does not prevent movement in traps which have been incised several times from the margin to the midrib to facilitate buffer diffusion into the mesophyll. Even an alkalinization of cell walls of plasmolysed leaf segments to pH 9 does not prevent considerable extensions of the mesophyll and subsequent movement of the specimens during deplasmolysis.These experiments make it very likely that the mesophyll cells are already extensible but are kept compressed in the open trap, thus developing tissue tension. The mechanism which prevents their extension as long as the trap is open can so far only be explained for traps which have been paralysed by a long-term incubation in 1 mM La³⁺. Leaf strips taken from stimulated, closed traps, comprising the lower epidermis and some mesophyll, prove to be highly extensible if they are stretched perpendicular to the midrib on a constant-load extensiometer. By contrast, strips taken from the lower side of paralysed traps are as rigid as those from the upper side of both stimulated and paralysed traps. From observations of semithin cross sections in a polarizing microscope, it is concluded that the extensibilities of these tissue strips are mainly determined by the cell walls of the upper epidermis plus a layer of adjacent mesophyll and by the lower epidermis, respectively, since these are the only cell walls with a preferential microfibril orientation in the direction of the applied stress.Abbreviations E _m membrane potential - E _s surface potential - Mes 2-(N-morpholino)ethanesulfonic acid - Tris 2-amino-2(hydroxymethyl)-1,3-propanediol     

Separation of cell enlargement and division in bean leaves

A method is presented for inducing cell enlargement in intact leaves and leaf strips of Phaseolus vulgaris L. without the complication of cell division. Primary bean leaves complete cell division and stop growing after 10 d in dim red light. Transfer to white light induces expansion (50% in 24 h) which is entirely the consequence of cell enlargement. Leaf strips from red-light-grown seedlings placed in white light and provided external solutes (10 mM KCl+10 mM sucrose) expand at the same rate as intact leaves in the light. This system makes possible future investigation of the mechanism of leaf cell enlargement.     

Electrical characteristics of stomatal guard cells: The ionic basis of the membrane potential and the consequence of potassium chlorides leakage from microelectrodes

The membrane electrical characteristics of stomatal guard cells in epidermal strips from Vicia faba L. and Commelina communis L. were explored using conventional electrophysiological methods, but with double-barrelled microelectrodes containing dilute electrolyte solutions. When electrodes were filled with the customary 1–3 M KCl solutions, membrane potentials and resistances were low, typically decaying over 2–5 min to near-30 mV and <0.2 k·cm² in cells bathed in 0.1 mM KCl and 1 mM Ca²⁺, pH 7.4. By contrast, cells impaled with electrodes containing 50 or 200 mM K⁺-acetate gave values of-182±7 mV and 16±2 k·cm² (input resistances 0.8–3.1 G, n=54). Potentials as high as (-) 282 mV (inside negative) were recorded, and impalement were held for up to 2 h without appreciable decline in either membrane parameter. Comparison of results obtained with several electrolytes indicated that Cl^- leakage from the microelectrode was primarily responsible for the decline in potential and resistance recorded with the molar KCl electrolytes. Guard cells loaded with salt from the electrodes also acquired marked potential and conductance responses to external Ca²⁺, which are tentatively ascribed to a K⁺ conductance (channel) at the guard cell plasma membrane.Measurements using dilute K⁺-acetate-filled electrodes revealed, in the guard cells, electrical properties common to plant and fungal cell membranes. The cells showed a high selectivity for K⁺ over Na⁺ (permeability ratio P_Na/P_K=0.006) and a near-Nernstian potential response to external pH over the range 4.5–7.4 (apparent P_H/P_K=500–600). Little response to external Ca²⁺ was observed, and the cells were virtually insensitive to CO₂. These results are discussed in the context of primary, charge-carrying transport at the guard cell plasma membrane, and with reference to possible mechanisms for K⁺ transport during stomatal movements. They discount previous notions of Ca²⁺-and CO₂-mediated transport control. It is argued, also, that passive (diffusional) mechanisms are unlikely to contribute to K⁺ uptake during stomatal opening, despite membrane potentials which, under certain, well-defined conditions, lie negative of the potassium equilibrium potential likely prevailing.Abbreviations and symbols EGTA ethylene glycol-bis(-aminoethyl ether)-N,N,N,N-tetraacetic acid - Hepes 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid - Mes 2-(N-morpholino) propanesulfornic acid - E equilibrium potential - G_m membrane conductance - R_in input resistance - V_m membrane potential     

Calcium bridges are not load-bearing cell-wall bonds in Avena coleoptiles

I examined the ability of frozen-thawed Avena sativa L. coleoptile sections under applied load to extend in response to the calcium chelators ethyleneglycol-bis-(-aminoethylether)-N,N,N,N-tetraacetic acid (EGTA) and 2-[(20bis-[carboxymethyl] amino-5-methylphenoxy)methyl]-6-methoxy-8-bis [carboxymethyl]aminoquinoline (Quin II). Addition of 5 mM EGTA to weakly buffered (0.1 mM, pH 6.2) solutions of 2(N-morpholino) ethanesulfonic acid (Mes) initiated rapid extension and wall acidification. When the buffer strength was increased (e.g. from 20 to 100 mM Mes, pH 6.2) EGTA did not initiate extension nor did it cause wall acidification. At 5 mM Quin II failed to stimulate cell extension or wall acidification at all buffer molarities tested (0.1 to 100 mM Mes). Both chelators rapidly and effectively removed Ca²⁺ from Avena sections. These data indicate that Ca²⁺ chelation per se does not result in loosening of Avena cells walls. Rather, EGTA promotes wall extension indirectly via wall acidification.Abbreviations EGTA ethyleneglycol-bis-(-aminoethylether)-N,N,N,N-tetraacetic acid - Quin II 2-[(2-bis-[carboxymethyl]amino-5-methylphenoxy)methyl]-6-methoxy-8-bis(carboxymethyl)aminoquinoline - Mes 2(N-morpholino)ethanesulfonic acid     

Role of acid efflux during growth promotion of primary leaves of Phaseolus vulgaris L. by hormones and light

The white-light-(WL) induced enlargement of dicotyledonous leaf cells is known to occur via an acid-growth mechanism; i.e., WL causes leaf cells to excrete protons which lead to an increase in wall extensibility and thus cell enlargement. Gibberellic acid (GA₃) and N⁶-benzyladenine (BA) also induce leaf cell enlargement. To see if they also act via acid-induced cell wall loosening, a comparison has been made of WL-, GA₃-and BA-induced growth of strips, taken from primary leaves of bean (Phaseolus vulgaris L.) plants raised in continuous red light for 10 d. White light, GA₃ and BA all increased wall extensibility as measured by the Instron technique, and this change preceded the increase in growth rate. However, whereas WL induced significant proton excretion, neither GA₃ nor BA caused any acidification of the apoplast. Furthermore, neutral buffers, which effectively inhibited the growth induced by WL, were without effect on growth promoted by either GA₃ or BA. These results indicate that while WL, GA₃ and BA all initiate growth in bean leaves by altering cell-wall properties, GA₃ and BA do so through some wall loosening mechanism other than wall acidification. Neither gibberellin nor cytokinin is likely to play a major role in light-induced cell enlargement of dicotyledonous leaves.Abbreviations BA N^o-benzyladenine - FC fusicoccin - GA₃ gibberellic acid - RL red light - SK medium 10 mM sucrose+10mM KCl - WL white light     

Single-cell, real-time measurements of extracellular oxygen and proton fluxes fromSpirogyra grevilleana

Summary We have adapted the self-referencing microelectrode technique to allow sensitive and noninvasive measurement of oxygen fluxes around single cells. The self-referencing technique is based on the translational movement of a selective microelectrode through the gradient next to the cell wall or membrane. The electrode is moved at a known frequency and between known points. The differential electrode output values are converted into a directional measurement of flux by the Fick equation. By coupling the newly developed oxygen-selective self-referencing electrochemical microelectrode (SREM-O₂) system with self-referencing ionselective proton measurements (SRIS-H⁺) we have characterized oxygen and proton fluxes from a single cell of the filamentous green algaSpirogyra gre illeana (Hass.). Oxygen showed a net efflux and protons showed a net influx when the cell was illuminated. These photosynthesis-dependent fluxes were found to be spatially associated with the chloroplasts and were sensitive to treatment with dichlorophenyldimethylurea. In the dark the directions of oxygen and proton fluxes were reversed. This oxygen influx was associated with mitochondrial respiration and was reduced by 78% when the cells was treated with 0.5 mM KCN. The residual cyanide-resistant respiration was inhibited by the application of 5 mM salicylhydroxamic acid, an inhibitor of the alternative oxidase. Similarly the cytochrome pathway was also inhibited by the presence of 20 M NO, while the cyanide-resistant alternative oxidase was not. These results demonstrate the use of the newly developed SREM-O₂ system to measure and characterize metabolic fluxes at a level of sensitivity that allows for subcellular resolution. These measurements, in conjunction with SERIS-H⁺ measurements, have led to new insights in our understanding of basic cellular physiology in plant cells.Abbreviations SRIS self-referencing ion selective - SREM self-referencing electrochemical microelectrode - ICP inductive coupled plasma spectroscopy     

Biochemical Approaches to Problems of Cellular Patterning*†

SYNOPSIS. The expression of intracellular patterning is perhaps nowhere more impressive than in the arrangements of structural elements associated with the cell surface in protozoa. The view is proposed that biochemical studies of protozoan plasma membranes and associated surface structures represent important contributions of potential significance for the understanding of the perpetuation, and expression of positional information at the intracellular level. Some recent work dealing with the isolation, identification, and metabolism of pellicular proteins in Tetrahymena is presented and discussed. Some integral membrane proteins have been identified by iodination and polyacrylamide gel electrophoresis. Labeling studies suggest heterogeneous turnover rates within the group of presently identified membrane proteins. High molecular weight proteins with some similarity to spectrin have been isolated from Tetrahymena epiplasm. It is suggested that the ciliate epiplasm is one example of membrane-associated, actomyosin-like systems found in a variety of cell types. The epiplasm may play a role in the positioning of surface-associated structures and in the control of cell shape.     

Reduced Na+/K+ ATPase transport activity,resting membrane potential,and bradykinin-stimulated phosphatidylinositol synthesis by polyol accumulation in cultured neuroblastoma cells

In these studies we examined the effect of polyol accumulation on neural cellmyo-inositol metabolism and properties. Neuroblastoma cells were cultured for two weeks in media containing 30 mM glucose, fructose, galactose or mannose with or without 0.4 mM sorbinil or 250 Mmyo-inositol. Chronic exposure of neuroblastoma cells to media containing 30 mM glucose, galactose, or mannose caused a decrease inmyo- inositol content and myo-[2-³H]inositol accumulation and incorporation into phosphoinositides compared to cells cultured in unsupplemented medium or medium containing 30 mM fructose as an osmotic control. These monosaccharides each caused an increase in intracellular polyol levels with galactitol > sorbitol = mannitol accumulation. Chronic exposure of neuroblastoma cells to media containing 30 mM glucose, galactose, or mannose caused a significant decrease in Na⁺/K⁺ ATPase transport activity, resting membrane potential, and bradykinin-stimulated³²P incorporation into phosphatidylinositol compared to cells cultured in medium containing 30 mM fructose. In contrast, basal incorporation of³²P into phosphatidylinositol or basal and bradykinin-stimulated³²P incorporation into phosphatidylinositol 4,5-bisphosphate were not effected. Each of these cellular functions as well asmyo-inositol metabolism and content and polyol levels remained near control values when 0.4 mM sorbinil, an aldose reductase inhibitor, was added to the glucose, galactose, or mannose supplemented media.myo-Inositol metabolism and content and bradykinin-stimulated phosphatidylinositol synthesis were also maintained when media containing 30 mM glucose, galactose, or mannose was supplemented with 250 Mmyo-inositol. The results suggest that polyol accumulation induces defects in neural cellmyo-inositol metabolism and certain cell functions which could, if they occurred in vivo, contribute to the pathological defects observed in diabetic neuropathy.     

NaCl-stimulated proton efflux and cell expansion in sugar-beet leaf discs

The expansion of illuminated sugar-beet leaf discs floating on aqueous solutions is stimulated by 10 mM NaCl. During expansion, protons are pumped out of the cell and NaCl increases this proton flux by about 40%. The nett flux of K⁺ and Na⁺ into the discs was also evaluated. During the expansion period K⁺ decreases while Na⁺ increases markedly. The results indicate the existence of a sodium-stimulated proton pump which is active during cell enlargement.Abbreviations IAA indole-3-acetic acid - PEG polyethylene glycol     

The regulation of K+ influx in excised barley roots

The electrochemical potential differences for potassium, between excised barley (Hordeum vulgare L.) roots and external media containing 0.05 mM KCl+0.5 mM CaSO₄, were determined over a 4-h period during which initially low-K⁺ roots accumulated K⁺ by pretreatment in 50 mM KCl plus 0.5 mM CaCl₂. This pretreatment resulted in increased internal [K⁺], decreased K⁺ influx (as measured from 0.05 mM KCl+0.5 mM CaSO₄) and decreased values of . These observations indicate that the decline of K⁺ influx associated with increased internal K⁺ concentration cannot be accounted for by passive adjustment to the electrochemical gradient for this ion.     

λ-Glutamylcysteine synthetase in higher plants: catalytic properties and subcellular localization

λ-Glutamylcysteine synthetase activity (EC 6.3.2.2) was analysed in Sephacryl S-200 eluents of extracts from cell suspension cultures ofNicotiana tabacum L. cv. Samsun by determination of λ-glutamylcysteine as its monobromobimane derivative. The enzyme has a relative molecular mass (M_r) of 60000 and exhibits maximal activity at pH 8 (50% at pH 7.0 and pH9.0) and an absolute requirement for Mg²⁺. With 0.2mM Cd²⁺ or Zn²⁺, enzyme activity was reduced by 35% and 19%, respectively. Treatment with 5 mM dithioerythritol led to a heavy loss of activity and to dissociation into subunits (M_r 34000). Buthionine sulfoximine andl-methionine-sulfoximine, known as potent inhibitors of λ-glutamylcysteine synthetase from mammalian cells, were found to be effective inhibitors of the plant enzyme too. The apparent K_m values forl-glutamate,l-cysteine, and α-aminobutyrate were, respectively, 10.4mM, 0.19 mM, and 6.36 mM. The enzyme was completely inhibited by glutathione (K_i=0.42 mM). The data indicate that the rate of glutathione synthesis in vivo may be influenced substantially by the concentration of cysteine and glutamate and may be further regulated by feedback inhibition of λ-glutamylcysteine synthetase by glutathione itself. λ-Glutamylcysteine synthetase is, like glutathione synthetase, localized in chloroplasts as well as in the cytoplasm. Chloroplasts fromPisum sativum L. isolated on a Percoll gradient contained about 72% of the λ-glutamylcysteine synthetase activity in leaf cells and 48% of the total glutathione synthetase activity. In chloroplasts ofSpinacia oleracea L. about 61% of the total λ-glutamylcysteine synthetase activity of the cells were found and 58% of the total glutathione synthetase activity. These results indicate that glutathione synthesis can take place in at least two compartments of the plant cell. Dedicated to Professor A. Prison on the occasion of his 80th birthday     

The influence of potassium content on the kinetics of potassium influx into excised ryegrass and barley roots

The influx of K⁺ into excised roots of barley (Hordeum vulgare L.) and ryegrass (Lolium multiflorum L.) previously grown with or without K⁺ was measured in K⁺ solutions ranging in concentration from 0.01 to 50 mM. In both species the K⁺ influx was lower in the roots with high K⁺ content. The extent of reduction by high internal [K⁺] decreased with external concentration above 1 mM. These results support the contention that at high external concentrations passive diffusion makes significant contributions to observed fluxes.     

The density of apical cells of dark-grown protonemata of the mossCeratodon purpureus

Summary Determinations of plant or algal cell density (cell mass divided by volume) have rarely accounted for the extracellular matrix or shrinkage during isolation. Three techniques were used to indirectly estimate the density of intact apical cells from protonemata of the mossCeratodon purpureus. First, the volume fraction of each cell component was determined by stereology, and published values for component density were used to extrapolate to the entire cell. Second, protonemal tips were immersed in bovine serum albumin solutions of different densities, and then the equilibrium density was corrected for the mass of the cell wall. Third, apical cell protoplasts were centrifuged in low-osmolarity gradients, and values were corrected for shrinkage during protoplast isolation. Values from centrifugation (1.004 to 1.015 g/cm³) were considerably lower than from other methods (1.046 to 1.085 g/cm³). This work appears to provide the first corrected estimates of the density of any plant cell. It also documents a method for the isolation of protoplasts specifically from apical cells of protonemal filaments.Abbreviations BSA bovine serum albumin - ER endoplasmic reticulum - V_v volume fraction     

Fine structure of laboratory cultured Distigma proteus and cytochemical localization of acid phosphatase

The phylogenetic relationship and origin of the euglenoids are controversial at present. It is not clear which of the extant genera may be most primitive, although Distigma proteus has been suggested as an early progenitor by some researchers. Scanning (SEM) and transmission (TEM) electron microscopic data were obtained in an effort to clarify some of the major taxonomic characteristics of this organism. SEM analyses of cells fixed during euglenoid movement (metaboly) show that the spirally arranged pellicular strips in the expanded regions of the organism have a lower pitch than those in the more constricted regions. This finding reveals the mechanical basis for euglenoid contortional movements. Mitochondria observed by TEM contain discoidal cristae, but some mitochondria are particularly large (e.g., 1.5 μm) and contain concentrically arranged, multiply layered cristae located deeper within the matrix, as in Khawkinea sp. Acid phosphatase reaction product is located in the cisternae and peripheral saccules throughout the Golgi stack and is regularly observed in the cisternae of the ER located beneath the pellicular ridges. Vacuoles varying in size, containing acid phosphatase reaction product, occur particularly near the periphery of the cell including the region surrounding the flagellar pouch. Occasional deposits of the reaction product within what appear to be membranous extrusions occur at the outer surface of the cell and may be secreted from sub-pellicular organelles. © 1994 Wiley-Liss, Inc.     

Reduction of Cr(VI) by Desulfovibrio vulgaris and Microbacterium sp.

Many industrial wastes contain Cr(VI), a carcinogen and mutagen, the toxicity of which can be ameliorated by reduction to Cr(III). Microbacterium sp. NCIMB 13776 andDesulfovibrio vulgaris NCIMB 8303 reduced Cr(VI) to Cr(III) anoxically using 25 mM sodium citrate buffer (pH 7), with 25 mM sodium acetate and 25 mM sodium formate as electron donors at 30 °C, under which conditions the rates of reduction of 500 M sodium chromate were 77 and 6 nmol h^–1 mg dry cell wt for D. vulgaris and Microbacterium sp., respectively, these being increased to 127 and 17 nmol h^–1 mg dry cell wt in the presence of 20 mM MOPS/NaOH buffer.     

Biochemical and Structural Analyses of Microtubules in the Pellicular Membrane of Leishmania tropica1

The structure of the major protein of the pellicular membrane of Leishmania tropica was investigated. This protein is composed of two polypeptides, of ca. 50,000 d molecular weight, that were found to cross-react immunologically with the α and β subunits of pig brain tubulin. The polypeptides and pig brain tubulin subunits were partially digested with S. aureus V8 protease, and the peptides obtained analysed by SDS-polyacrylamide gel electrophoresis. A comparison of the patterns showed that the β subunits of Leishmania and pig tubulin have very similar primary structures, while the α subunits have evolved divergently. These experiments demonstrate that the major polypeptides found in the pellicular membrane of L. tropica are α and β subunits of tubulin. Immuno-electron microscopy indicates that the tubulin is located in the microtubules associated with the pellicular membrane of Leishmania. Arrays of microtubules were prepared by nonionic detergent treatment of the cells and observed by electron microscopy after negative staining. Optical diffraction reveals a 5 nm spacing between protofilaments in the microtubule and a 4 nm axial periodicity corresponding to the tubulin subunits. The pitch of the shallow left-hand three-start helix is 12°. A distance of 47 nm separates each microtubule from the next. These data show that the dimensions and supramolecular organization of the tubulin subunits in the microtubules are identical in the pellicular membrane of L. tropica and in mammalian brain.     

Cytoplasmic streaming in giant algal cells: A historical survey of experimental approaches

Various methods have been used to study cytoplasmic streaming in giant algal cells during the past three decades. Simple techniques can be used with characean internodal cells to modify the cell constitution in various ways to gain insight into the mechanism of cytoplasmic streaming. Another method involves isolatingin vitro a huge drop of uninjured endoplasm, to examine its physical and dynamic properties. The motive force responsible for streaming has been measured by three different techniques with similar results. Subcortical fibrils consisting of bundles of F-actin with the same polarity are indispensable for streaming. Differential treatment of the endoplasm and ectoplasm has shown that putative characean myosin is localized in the endoplasm. Studies of the roles of ATP, Mg²⁺, Ca²⁺, H⁺ etc. in the streaming have been conducted by cellular perfusion, which allows removal of the tonoplast, or by techniques permeabilizing the protoplasmic membrane. A slow version of the movement can even be artificially reproduced by combining characean actinin situ and exogenous myosin in the presence of Mg-ATP. The findings thus far obtained support the hypothesis that cytoplasmic streaming in characean cells is caused by an active shearing force produced by interaction of the actin filament bundles on the cortex with myosin in the endoplasm.     

Competitive growth of slow growingRhizobium japonicum against fast growingEnterobacter andPseudomonas species at low concentrations of succinate and other substrates in dialysis culture

A cultivation system with simultaneous growth of six bacterial cultures in separate bags in dialysis culture was developed. In a medium with no added carbon source (one half concentrated Hoagland solution, water deionized and distilled), cell number ofRhizobium japonicum increased during a 7 day period by a factor of 35, whereas the number ofEnterobacter aerogenes cells decreased to one half. With a concentration of 100 nM succinate as an additional carbon source in the inflow,Rhizobium japonicum 61-A-101 cell number increased by a factor of 50 during an 8 day period, whereas cell number ofEnterobacter cloacae NCTC 10005 only doubled and ofEnterobacter aerogenes NCTC 10006 decreased. At 10 mM concentration of succinate in the inflow, doubling time the twoEnterobacter strains was about 12 h, compared to about 24 h for theRhizobium japonicum strain. Varying the succinate concentration from 10 mM to 100 nM in the inflow,Rhizobium japonicum 61-A-101 surpassed theEnterobacter aerogenes strains in the growth rate between 1 mM and 100 M succinate in the inflowing medium. Three otherRhizobium japonicum strains (fix⁺ and fix^-) did grow with a similar rate as strain 61-A-101 at very low concentrations of substrate. Growth rates for the strains were confirmed by protein data per culture. Growing in competition with twoPseudomonas strains,Rhizobium japonicum RH 31 Marburg (fix^-) did overgrow alsoPseudomonas fluorescens, was however outgrown byPseudomonas putida. In utilizing low concentrations of a¹⁴C labelled organic acid (malonate), three strains ofRhizobium japonicum left 2–4 times smaller amounts of¹⁴C in the medium than two species ofPseudomonas and two species ofArthrobacter.On sabbatical leave at ANU