scholarly journals FACTORS AFFECTING THE REDISTRIBUTION OF SURFACE-BOUND CONCANAVALIN A ON HUMAN POLYMORPHONUCLEAR LEUKOCYTES

1974 ◽  
Vol 62 (2) ◽  
pp. 351-365 ◽  
Author(s):  
Graeme B. Ryan ◽  
Joan Z. Borysenko ◽  
Morris J. Karnovsky
Keyword(s):  
Concanavalin A ◽  
Polymorphonuclear Leukocytes ◽  
Cytochalasin B ◽  
Immobilized Cells ◽  
Fluorescein Isothiocyanate ◽  
Cross Linking ◽  
Con A ◽  
Factors Affecting ◽  
Motile Cells ◽  
Cellular Locomotion

Human neutrophil polymorphonuclear leukocytes (PMN) were studied to determine the influence of cellular locomotion upon the redistribution and capping of concanavalin A (Con A). Con A was detected by fluorescence (using Con A conjugated to fluorescein isothiocyanate [Con A-FITC]), or on shadow-cast replicas (using Busycon canaliculatum hemocyanin as a marker for Con A). After labeling with Con A 100 µg/ml at 4°C and warming to 37°C, locomotion occurred, and the Con A quickly aggregated into a cap at the trailing end of the cell. When locomotion was inhibited (with cytochalasin B, or by incubation in serum-free medium at 18°C) Con A rapidly formed a cap over the central region of the cell. Iodoacetamide inhibited capping. PMN labeled with FITC, a monovalent ligand, developed caps at the tail only on motile cells; FITC remained dispersed on immobilized cells. PMN exposed to Con A 100 µg/ml at 37°C bound more lectin than at 4°C, became immobilized, and showed slow central capping. The Con A soon became internalized to form a perinuclear ring. Such treatment in the presence of cytochalasin B resulted in the quick formation of persistent central caps. Colchicine (or prior cooling) protected PMN from the immobilizing effect of Con A, and tail caps were found on 30–40% of cells. Immobilization of colchicine-treated cells caused Con A to remain in dispersed clusters. Thus, capping on PMN is a temperature- and energy-dependent process that proceeds independently of cellular locomotion, provided a colchicine-sensitive system is intact and the ligand is capable of cross linking receptors. On the other hand, if the cell does move, it appears that ligands may be swept into a cap at the tail whether cross-linking occurs or not.

scholarly journals Reciprocal transmembranous receptor-cytoskeleton interactions in concanavalin A-activated platelets.

1985 ◽  
Vol 101 (3) ◽  
pp. 993-1000 ◽  
Author(s):  
M E Wheeler ◽  
J M Gerrard ◽  
R C Carroll
Keyword(s):  
Platelet Activation ◽  
Concanavalin A ◽  
Prostaglandin E1 ◽  
Cytochalasin B ◽  
Specific Interaction ◽  
Human Platelets ◽  
Cross Linking ◽  
Con A ◽  
Glycoprotein Complex ◽  
Activated Platelets

Concanavalin A (Con A) has been used to activate platelets, inducing a specific interaction between the glycoprotein IIb-IIIa complex and the cytoskeleton of the activated platelet. In agreement with this, we have shown that Con A activates human platelets, initiating phosphorylation, secretion, and cytoskeletal formation. Con A and cytochalasin B were used to demonstrate a reciprocal interaction of the glycoprotein complex with the platelet cytoskeleton. Additionally, we have shown that a similar reciprocity is provided by the multivalent fibrin-fibrinogen platelet interaction found in the thrombin-induced clot. Con A differs from other activators in precipitating an apparent cytoskeletal core despite a complete inhibition of platelet activation by prostaglandin E1. We suggest, from this result, that Con A may be cross-linking a membrane-associated cytoskeletal complex present in the unactivated platelet.

scholarly journals Concanavalin A induces microtubule assembly and specific granule discharge in human polymorphonuclear leukocytes.

1976 ◽  
Vol 68 (3) ◽  
pp. 781-787 ◽  
Author(s):  
S Hoffstein ◽  
R Soberman ◽  
I Goldstein ◽  
G Weissmann
Keyword(s):  
Concanavalin A ◽  
Binding Sites ◽  
Polymorphonuclear Leukocytes ◽  
Cytochalasin B ◽  
Microtubule Assembly ◽  
Human Neutrophils ◽  
Con A ◽  
Specific Granules ◽  
Specific Granule ◽  
Human Polymorphonuclear Leukocytes

Human neutrophils stimulated by concanavalin A (Con A, 100 microng/ml) contained markedly enhanced numbers of microtubules and discharged peroxidase-negative (specific) but not peroxidase-position (azurophile) granules. Release of lysozyme from specific granules was dose and time dependent, could be inhibitied by alpha-methyl-D-mannoside, and enhanced by cytochalasin B. Many microtubules were associated with internalized plasma membrane bearing Con A binding sites.

Mobility of concanavalin A receptors and distribution of cytoplasmic actin in odontogenic epithelial and mesenchymal cells

1978 ◽  
Vol 33 (1) ◽  
pp. 329-340
Author(s):  
S.S. Prime ◽  
B.H. Toh
Keyword(s):  
Epithelial Cells ◽  
Concanavalin A ◽  
Cytochalasin B ◽  
Fluorescein Isothiocyanate ◽  
Tooth Germ ◽  
Cell Types ◽  
Mesenchymal Cells ◽  
Staining Procedure ◽  
Con A ◽  
Cytoplasmic Actin

The distribution of concanavalin A (Con A) surface receptors and cytoplasmic actin in the same cell was studied in monolayer cultures of 2 odontogenic epithelial cells of different developmental age and in ecto-mesenchymal cells derived from the same tooth germ. Con A receptors were demonstrated by fluorescein-isothiocyanate-labelled Con A (FITC-Con A) and cytoplasmic actin by a specific anti-actin autoantibody (AAA) traced with a rhodamine-labelled goat anti-human globulin (R-AHG). All 3 cell types, incubated with FITC-Con A at 37 degrees C for increasing time periods, showed progressive changes in staining patterns from clusters, caps to perinuclear globules. Capping was seen in the majority of immature epithelial cells at 120–180 min, in cells of more mature epithelium at 180–240 min and in ecto-mesenchymal cells at 240–360 min. Binding of FITC-Con A to cell surfaces resulted in sequential changes in AAA staining from filamentous to an aggregated or diffuse pattern, co-capping of aggregated or diffusely stained areas with those capped by FITC-Con A, presence of aggregated or diffusely stained areas in sites similar to the perinuclear globules stained by FITC-Con A, to final re-emergence of filamentous staining. Prior treatment of cells with cytochalasin B or colchicine promoted capping in epithelial but not in ecto-mesenchymal cells while presence of either drug throughout the staining procedure inhibited capping. The results show that Con A receptors are more mobile in epithelial compared to ecto-mesenchymal cells and in immature epithelial cells compared to their more mature counterparts, and that binding and mobility of Con A receptors on the cell surface is associated with redistribution of cytoplasmic actin. The cytochalasin B and colchicine experiments suggest that both microfilaments and microtubules may have synergistic roles in the opposing functions of receptor anchorage and mobility, and that the relative receptor immobility of ectomesenchymal compared to epithelial cells may be attributed to firmer receptor anchorage to the cytoskeleton.

scholarly journals Antagonism by dibutyryl adenosine cyclic 3',5'-monophosphate and testololactone of concanavalin A capping.

1975 ◽  
Vol 66 (2) ◽  
pp. 392-403 ◽  
Author(s):  
B Storrie
Keyword(s):  
Concanavalin A ◽  
Binding Sites ◽  
Cytochalasin B ◽  
Cold Treatment ◽  
Intracellular Camp ◽  
Camp Concentration ◽  
Con A ◽  
Cell Rounding ◽  
Intracellular Camp Concentration

Exposure of CHO-K1 cells in vitro to dibutyryl adenosine cyclic 3',5'-monophosphate (DBcAMP) plus testololactone produces a rapid, reversible antagonism of ligand-induced collection of initially dispersed concanavalin A (Con A) binding sites into a caplike mass. Morphologically, as Con A capping occurs, the cells become less spread and then round completely. With prolonged Con A exposure, cells cultured in either the absence or the presence of DBcAMP plus testololactone cap and round. Capping is blocked by cold treatment and respiratory inhibitors. Colcemid at concentrations greater than 1 muM promotes both Con A capping and cell rounding. Cytochalasin B at similar concentrations inhibits both capping and cell rounding. Treatment of cells with Con A has little effect on intracellular cAMP concentration. Possible mechanisms by which cAMP may modulate the movement of Con A binding sites are discussed.

scholarly journals Concanavalin A binding to human erythrocytes leads to alterations in properties of the membrane skeleton

1987 ◽  
Vol 241 (2) ◽  
pp. 521-525 ◽  
Author(s):  
S M Gokhale ◽  
N G Mehta
Keyword(s):  
Concanavalin A ◽  
Membrane Skeleton ◽  
Cross Linking ◽  
Dependent Manner ◽  
Con A ◽  
Normal Cells ◽  
Skeletal Protein ◽  
Low Ionic Strength ◽  
Dose Dependent ◽  
Resistance To Heat

Three properties related to the erythrocyte membrane skeleton are found to be altered after the binding of concanavalin A (Con A) to erythrocytes or their isolated membranes. Con A binding to normal erythrocytes imparts resistance to heat (49 degrees C)-induced fragmentation of the cells. The fragmentation, due to denaturation of spectrin at 49 degrees C, is prevented by Con A in a dose-dependent manner, but levels off at concentrations of Con A in excess of 100 micrograms/ml. The binding of Con A to ghosts isolated from normal, trypsin- or Pronase-treated cells prevents (completely or substantially) the elution of the skeletal protein complex when the membranes are extracted under low-ionic-strength conditions in the cold. The Con A-agglutinated membranes of trypsin- and Pronase-treated, but not normal, cells show cross-linking of skeletal proteins and band 3 with dimethyl adipimidate, a 0.86 nm (8.6 A)-span bifunctional reagent. The extent of cross-linking is greater in the Pronase-treated membrane than in the less-agglutinable trypsin-treated membranes. The results show that, after Con A has bound, rearrangements occur in the membrane that alter properties of the skeletal proteins. Additionally, redistribution of the skeletal proteins and the Con A receptor occurs in the lectin-agglutinated membranes.

Concanavalin A binding to amphibian embryo and effect on morphogenesis

Development ◽  
1979 ◽  
Vol 51 (1) ◽  
pp. 63-72
Author(s):  
J. C. Boucaut ◽  
B. Bernard ◽  
M. Aubery ◽  
R. Bourrillon ◽  
Ch. Houillon
Keyword(s):  
Concanavalin A ◽  
Direct Interaction ◽  
Fluorescein Isothiocyanate ◽  
Inhibitory Effect ◽  
Vitelline Membrane ◽  
Con A ◽  
Amphibian Embryo ◽  
Before And After ◽  
Three Stages ◽  
Pleurodeles Waltlii

The effect of Concanavalin A (Con A) on morphogenesis in Pleurodeles waltlii has been studied. Embryos were incubated with various concentrations of the lectin for a period of 6 days. Three stages of development were examined, late blastula, young gastrula and late gastrula. In the presence of the lectin at a concentration of 200, 150 or 100 μg/ml morphogenic movements were delayed, altered and finally blocked. At lower concentrations, 50 or 25 μg/ml, there was a slight delay in gastrulation, but in some cases development was normal. These findings indicate that Con A exerted an inhibitory effect on amphibian morphogenesis and there is evidence that the lectin effect was concentration dependent. The effects of Con A were specific since they were totally inhibited by α-methyl-D-mannopyranoside (0·05 m). The viability of the 24 h lectin-treated embryos was demonstrated by washing experiments. Labelled Con A binding to the embryos was investigated before and after discarding the vitelline membrane. The results suggest a direct interaction between Con A and the cell surface and this was confirmed by using fluorescein isothiocyanate Con A.

Lectin-mediated agglutination of amphibian embryonic cells

1977 ◽  
Vol 27 (1) ◽  
pp. 227-243
Author(s):  
B.R. Fraser ◽  
S.E. Zalik
Keyword(s):  
Concanavalin A ◽  
Wheat Germ ◽  
Cytochalasin B ◽  
Ricinus Communis ◽  
Soya Bean ◽  
Embryonic Cells ◽  
Glutaraldehyde Fixation ◽  
Con A ◽  
Neuraminidase Treatment ◽  
Ricinus Communis Agglutinin

Dissociated blastula cells of Xenopus laevis are agglutinated with wheat germ agglutinin (WGA), Ricinus communis agglutinin (RCA), concanavalin A (Con A) and, to a lesser extent with soya bean agglutinin (SBA). They are not agglutinated with fucose-binding protein. Neuraminidase treatment of cells enhances their agglutinability with RCA and SBA, but has no effect on Con A- and WGA-mediated agglutinability. Treatment of cells with procaine, or xylocaine, has no effect on the cells' agglutinability or on the extrusion of lobopodia. Treatment with colchicine or cytochalasin B either separately or simultaneously has no effect on lectin-mediated agglutinability. Cells treated with cytochalasin B or colchicine and cytochalasin B simultaneously lack lobopodial extensions, while colchicine alone has no effect on these structures. Phenothiazine tranquillizers inhibit agglutination mediated by all of the above mentioned lectins. Lobopodial extensions are absent in cells treated with these compounds. Glutaraldehyde fixation inhibits RCA and WGA mediated agglutinability and reduces the Con A-mediated agglutinability. Results suggest that in this system microtubules and microfilaments are not involved in lectin-mediated agglutination.

scholarly journals Kinetic evidence for a common mechanism of capping on lymphocytes

1982 ◽  
Vol 204 (1) ◽  
pp. 229-237 ◽  
Author(s):  
Anthony N. Corps ◽  
James C. Metcalfe ◽  
Tullio Pozzan
Keyword(s):  
Concanavalin A ◽  
Cytochalasin B ◽  
Specific Effect ◽  
Common Mechanism ◽  
Cross Linking ◽  
Detectable Effect ◽  
Membrane Flow ◽  
Surface Immunoglobulin ◽  
Fixed Times ◽  
Concanavalin A Receptors

1. Differences in the rates at which ligands cap various receptors on the same cells, and their sensitivity to various drugs, have been interpreted as evidence that there are distinct mechanisms for ‘fast’ and ‘slow’ cap formation. We have examined the factors which determine the rate of cap formation of three receptors on mouse splenic lymphocytes or thymocytes, and compared the effects of cytochalasin B or colchicine under conditions where the different receptors cap at similar rates. 2. When surface immunoglobulin, concanavalin A receptors, or θ antigen are induced to cap at their maximal rates by appropriate concentrations of one or more cross-linking ligands, the half-time for maximal capping of each receptor population is between 1.5 and 3.0min at 37°C. Slower rates of cap formation are obtained by using non-optimal concentrations of the cross-linking ligands. 3. When the three receptors were induced to cap at similar rates (either maximal or slower), 10μm-cytochalasin B caused a similar decrease in the rate of cap formation for each receptor, without affecting the eventual extent of capping. At comparable capping rates on control cells, colchicine (10μm) increased the rate of cap formation for surface immunoglobulin and concanavalin A receptors to a similar extent, without affecting the eventual extent of cap formation. In contrast, colchicine had no detectable effect on the capping of θ antigen. 4. From these results, we conclude that there are no intrinsic differences in the rates at which different receptors can be induced to cap that can be used to diagnose differences in their mechanisms of cap formation. The observation that ligand concentration and the drugs acting on the cytoskeleton generally affect the rate but not the extent of cap formation accounts for the wide variation in reported effects of the drugs on cap formation measured at fixed times. The receptor-specific effect of colchicine on surface immunoglobulin and concanavalin A receptors, but not θ antigen, is not readily compatible with models of cap formation which depend on lipid or membrane flow.

scholarly journals Transmembrane linkage between surface glycoproteins and components of the cytoplasm in neutrophil leukocytes.

1981 ◽  
Vol 90 (3) ◽  
pp. 743-754 ◽  
Author(s):  
P Sheterline ◽  
C R Hopkins
Keyword(s):  
Plasma Membrane ◽  
Concanavalin A ◽  
Fluorescein Isothiocyanate ◽  
Total Cell ◽  
Membrane Glycoproteins ◽  
Con A ◽  
Fluorescence And Electron Microscopy ◽  
Nonionic Detergent ◽  
Enzyme Markers ◽  
Neutrophil Leukocytes

An experimental approach is described that enables the analysis of interactions between exogenous surface ligands and components of the cytoplasm in neutrophil leukocytes. Neutrophils treated with the nonionic detergent Lubrol PX, under controlled conditions, yield intact detergent-insoluble ghosts. Morphological analysis of neutrophil ghosts shows that they retain the original dimensions of the cell and consist almost entirely of a peripheral filamentous network, representing the submembranous cortical web, concentric to nuclear remnants. All intracellular membrane-bounded organelles, plasma membrane, and background cytoplasmic electron density are absent. Biochemical analysis of the ghosts shows that less than 10% of enzyme markers for the soluble and granule fractions remain, and that greater than 90% of total cell phospholipid is removed during detergent extraction. The major proteins remaining in the ghosts comigrate, on polyacrylamide gels in the presence of SDS, with chicken gizzard actin, myosin, filamin, and a 110-kdalton protein. Patches and caps induced on neutrophils with either fluorescein isothiocyanate-concanavalin A or ferritin-concanavalin A retain their original location and morphology on ghosts after lysis, as determined by both fluorescence and electron microscopy. In similar experiments, but using 125I-labeled lectins, 37% of total cell bound concanavalin A (Con A) and 25% succinylated Con A remain attached to the ghosts. A major 125I-labeled membrane glycoprotein (80 kdaltons) is associated with ghosts prepared from intact neutrophils iodinated in the presence of exogenous lactoperoxidase. Further 125I-labeled membrane glycoproteins (217, 170, and 147 kdaltons) become associated with ghosts prepared from iodinated cells treated before lysis with Con A, but not with succinylated Con A. These data taken together suggest that linkages exist in neutrophils between proteins exposed on the outer surface of the plasma membrane and the peripheral filamentous network independent of the presence of lipid bilayer. The implications of these findings for surface motile phenomena will be discussed.

scholarly journals Effects of glutathione-oxidizing agents on microtubule assembly and microtubule-dependent surface properties of human neutrophils.

1976 ◽  
Vol 71 (3) ◽  
pp. 921-932 ◽  
Author(s):  
J M Oliver ◽  
D F Albertini ◽  
R D Berlin
Keyword(s):  
Concanavalin A ◽  
Peripheral Blood ◽  
Polymorphonuclear Leukocytes ◽  
Human Peripheral Blood ◽  
Microtubule Assembly ◽  
Human Neutrophils ◽  
Hexose Monophosphate ◽  
Con A ◽  
Oxidizing Agents ◽  
The Face

In human peripheral blood polymorphonuclear leukocytes and lymphocytes, GSH-oxidizing agents promote the movement of surface-bound concanavalin A (Con A) into caps and inhibit the assembly of microtubules (MT) that is normally induced by Con A binding. Con A capping and inhibition of MT assembly occur when GSH levels in cell suspensions are decreased by 30-70%, and return to GSH to control levels is accompanied by the appearance of cytoplasmic MT and by inhibition of the capping response with Con A. Oxidation of GSH markedly stimulates the hexose monophosphate shunt, and regeneration of GSH occurs rapidly. The data indicate that MT cannot be assembled or maintained in the face of decreased GSH levels. Thus, GSH homeostasis becomes critical during physiological events such as phagocytosis which simultaneously induce the assembly of MT and the production of agents like H2O2 that can oxidize GSH.

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