DOCSLIB.ORG

The Cellular Response to Cytochalasin B: a Critical Overview

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
The Cellular Response to Cytochalasin B: a Critical Overview Cytologia 39: 709-727 , 1974 The Cellular Response to Cytochalasin B: A Critical Overview Michelle Copeland Children'sCancer Research Foundation , 35 BinneyStreet, Boston, Massachusetts02115 , U. S. A. ReceivedApril 3, 1973 The cytochalasins comprise a novel class of fungal metabolites which have attracted a great deal of recent interest as a result of the numerous apparently di versecytological effects which are elicited by their presence, both in vivo and in vitro. Six cytochalasins have been isolated, to date, from the cultures of Helminthospor ium dematioidem, Metarrhizium anisopliae, and Rosellina necatrix, although some what similar biological activity has been associated with the fermentation products of several additional fungi (Aldridge et al. 1967). The six currently isolable cyto chalasins are designated A, B, C, D, E, and F. In terms of their biological activity, i nsofar as this is known at present, these six cytochalasins appear qualitatively simi lar to one another, although the potencies of the isomeric cytochalasins C and D Aldridge et al., 1967 Chem. Comm. 1: 26 Fig. 1. The chemical structure of cytochalasin B. exceedthose of cytochalasins A and B by a factor of approximately ten (Carter 1967). Themost readily isolated of these is the B variant , which has been shown to be iden tical with the antibiotic "phomin" (Aldrige and Turner 1968) (Fig. 1). The addi tional features of appropriate potency , essential nontoxicity at low concentrations and reversibility of action have made cytochalasin B the subject of most laboratory studies of the biological phenomena produced by the cytochalasins. Preliminary investigators employed the term"cytochalasin" (from the Greek cytos: cell;+chalasis: relaxation) to denote the cytological changes induced by these metabolic end-products . These substances appeared to "relax" the cell 710 Michelle Copeland Cytologia 39 through a mechanism which may have involved an inhibition of the normal process of cell motility or a structural variation of the cytoskeleton. A more thorough consideration of this phenomenon will be presented later in this discussion. A reasonable approach to the problem of formulating a realistic mechanism for the action of cytochalasin B is to attempt to establish a functional relationship between the apparently diverse cytological effects which it has been demonstrated to elicit. Among the more prominent of these effects are several which are chara acterized as morphological in nature, such as cytoskeletal variation, multinucleation, Fig. 2. Large quantities of granular endoplasmic reticulum and annulate lamellae are seen in the cytoplasm of this multinucleated cell; this fibroblast cell was exposed to cytochalasin B (1ƒÊg/ml) for 6-7 days. Scale line is 1um. Courtesy of Dr. Awtar Krishan. diffuse organelle alteration, nuclear extrusion, and surface phenomena of diminished membrane ruffling; other effects are described as functional or physiological, and include inhibition of cell motility, of mitotic reproduction and of phagocytosis and pinocytosis, as well as alteration of surface adhesion properties and of micro filaments and depression of synthesis or uptake of membrane components. One rather convincing element which is consistent with some of these processes is an alteration in the properties of the cell plasma membrane and it is not without evi dence that this avenue may be viewed as central to the activity of cytochalasin B. 1974 Cytochalasin B 711 Alteration of cell morphology The diverse cellular effects of cytochalasin B were first systematically noted by Carter (1967) in experiments with fibroblasts in culture . His initial observation of these cells treated with cytochalasin B included diminished levels of both normal motile activity and characteristic membrane ruffling . At concentrations of 1 .0 ƒÊ g/ml of cytochalasin B these fibroblasts were also noted to assume a flattened configuration, and to occupy a larger surface area on glass after treatment . Kro shan (1971) corroborated this early evidence for membrane involvement and sup plemented it with the observation that fibroblasts exposed to similar concentra tions of cytochalasin B transformed from the normal spindle into flat , epithelioid shapes through contraction of elongated pseudopodia. These discoid fibroblasts became multinucleated and accumulated Golgi membranes, rough endoplasmic reticulum and annulate lamellae such that after fifteen days of culture in medium Fig. 3. A large cytochalasin-induced multinucleated fibroblast cell with 9 nuclei . Individual variation in the nuclear size is evident. Exposed to cytochalasin B (1ƒÊg/ml) for 6-7 days. Scale line is 1ƒÊm. Courtesy of Dr. Awtar Krishan. containing cytochalasin B, the cells attain diameters of approximately 300 microns and possessed eight or nine nuclei each (Figs. 2 and 3). The possibility that these alterations in cell structure might be due to an augmentation by cytochalasin B of the synthesis of cellular elements has been effectively eliminated by the demonstra tion that the rates of DNA, RNA and protein syntheses are unaffected by cyto chalasin B (Estensen 1971). Krishan's data are consonant with a process in which the drug influences merely the plasma membrane such that the maintenance of even normal metabolic activities results in large-scaled cytoeconomics. That is, quite simply, that cells behave metabolically as if they had been separated from one another following normal mitosis, and that each member "cell" of the cytochalasin B-induced conglomerate contributed to the mass that quantity of synthetic pro ducts which it would have formed in the independent state. This consideration points out an important aspect of the analysis of the activities of cytochalasin B; 712 Michelle Copeland Cytologia 39 that is, the drug may have a single direct action on a component of, say, the plasma membrane and yet produce in vitro variations which suggest a diversity of roles. An additional example of the application of this principal has been provided by Krishan in an explanation of the conspicuously large number of RNA virus-like particles found within the giant fibroblasts produced by cytochalasin B. Krishan has realistically suggested that the larger fibroblast cells were preferentially suited to the proliferation of endogenous L-cell virus-like particles as well as exogenously introduced virus-like reo and Herpes viruses in that the hosts contained large cyto plasmic masses, unusually large nuclei, relatively hypertrophied endoplasmic reti culum, and numerous other cell organelles in high numbers. Arborization and cellular adhesion Insight into the processes resulting in the observations of Carter and Krishan have been provided by Sanger and Holtzer (1972), who have validated an alteration by cytochalasin B of the cytoskeleton. Exposure of myoblasts, fibroblasts and stellate chondroblasts for 48 hours to 5ƒÊg/m1 of cytochalasin B caused the cells to assume a branched gross morphology and induced a virtual immotility. Reversal of this "arborization" could be achieved by withdrawal of cytochalasin B from the medium. It was noted by these authors that the resumption of normalized cell morphology and considerable recovery of cellular motility did not entail retraction of the extruded processes, the overall dimensions of which remain unchanged. The cells apparently lose their arborized morphology by gradually filling the area between the processes with outflowing lobopodia, the forms of which are indis tinguishable from the advancing tip of amoebae, and no membrane "ruffling" pheno mena were associated with the lobopodial protrusion. Additional experiments were conducted by these researchers in which specific metabolic inhibitors were employed to assess the role of cytochalasin B in the arborization process. Ar borized cells incubated with cycloheximide (an inhibitor of protein synthesis) and 5ƒÊg/ml of cytochalasin B for 30 minutes and then transferred to a medium con taining only cycloheximide underwent normal reversal of shape. From this datum it can be concluded that the motile lobopodial activity responsible for normaliza tion of shape is not dependent upon protein synthesis. On the other hand, during exposure of arborized cells to sodium cyanide, an inhibitor of oxidative phos phorylation, the branched conformation is maintained until this inhibitor is with drawn. It is apparent, therefore, that protrusion of lobopodia is dependent upon energy production and independent of protein synthetic activity. In addition, it should be mentioned that not all of the cell types treated with cytochalasin B ex hibited arborization under the conditions employed. Replicating chick embryo erthrocytes undergoing nuclear division show neither cytokinesis nor arborization; whereas presumptive myoblasts arborize upon treatment with cytochalasin B, their fusion products, myotubes, do not; furthermore, the nonfunctional chondrocyte, lacking a maturation capsule, does exhibit arborization, while the functional chon droblast encapsulated in chondroitin sulfate does not branch. It has been sug gested that these differing responses to cytochalasin B may reflect fundamental differences in cell surfaces. Myoblasts, for example, are thought to undergo con siderable surface alteration following fusion into multinucleate myotubes, and 1974 Cytochalasin B 713 this change may effect variation in the cellular response to cytochalasin B . Simi larly, while the nonfunctional chondrocyte is freely exposed to the drug , the en capsulated chondroblast may not
Load more

Recommended publications
  • The Sensitivity of Developing Cardiac Myofibrils to Cytochalasin-B (Electron Microscopy/Polarized Llght/Z-Bands/Heartbeat) FRANCIS J
    Proc. Nat. Acad. Sci. USA Vol. 69, No. 2, pp. 308-312, February 1972 The Sensitivity of Developing Cardiac Myofibrils to Cytochalasin-B (electron microscopy/polarized llght/Z-bands/heartbeat) FRANCIS J. MANASEt,* BETH BURNSIDEt't, AND JOHN STROMAN* * Departments of Anatomy and Pediatrics, Harvard Medical School, Boston, Massachusetts 02115; Department of Cardiology and Pathology, The Children's Hospital Medical Center, Boston, Mass. 02115; and t Department of Anatomy, Harvard Medical School, Boston, Mass. 02115 Communicated by Keith R. Porter, November 12, 1971 ABSTRACT Developing cardiac muscle cells of 11- to (about 30-40 hr of incubation) were selected. The splanchno- 13-somite chick embryos are sensitive to cytochalasin-B. the In cultured chick embryos, ranging in development from pleure of the developing yolk sac was removed, exposing 11 to 13 somites, hearts stop beating in the presence of this heart. If care is taken to prevent damage to the anterior in- agent. Both polarized light and electron microscopic testinal portal, this surgical procedure does not interfere with examination show that cytochalasin-B disrupts existing normal development during the time periods used in this myofibrils and inhibits the formation of new ones. Dis- study. In experimental series, the culture medium was re- crete Z-bands are not present in treated heart cells and thick, presumably myosin, filaments are found in dis- placed with either 0.1 mM (50 ug/ml) or 20 ,gM (10 jsg/ml) array. These effects are reversible; after cytochalasin-B is cytochalasin-B in Tyrodes solution plus 1% dimethyl sulf- removed from the medium, heartbeat recovers and myo- oxide (Me2SO) for specified incubation times.
    [Show full text]
  • SAFETY DATA SHEET Revision Date 04/30/2021 Print Date 09/25/2021
    Version 6.3 SAFETY DATA SHEET Revision Date 04/30/2021 Print Date 09/25/2021 SECTION 1: Identification of the substance/mixture and of the company/undertaking 1.1 Product identifiers Product name : Cytochalasin B, from Drechslera dematioidea Product Number : C6762 Brand : Sigma CAS-No. : 14930-96-2 1.2 Relevant identified uses of the substance or mixture and uses advised against Identified uses : Laboratory chemicals, Synthesis of substances 1.3 Details of the supplier of the safety data sheet Company : Sigma-Aldrich Inc. 3050 SPRUCE ST ST. LOUIS MO 63103 UNITED STATES Telephone : +1 314 771-5765 Fax : +1 800 325-5052 1.4 Emergency telephone Emergency Phone # : 800-424-9300 CHEMTREC (USA) +1-703- 527-3887 CHEMTREC (International) 24 Hours/day; 7 Days/week SECTION 2: Hazards identification 2.1 Classification of the substance or mixture GHS Classification in accordance with 29 CFR 1910 (OSHA HCS) Acute toxicity, Oral (Category 2), H300 Acute toxicity, Inhalation (Category 1), H330 Acute toxicity, Dermal (Category 2), H310 Reproductive toxicity (Category 2), H361 For the full text of the H-Statements mentioned in this Section, see Section 16. 2.2 GHS Label elements, including precautionary statements Pictogram Signal word Danger Sigma - C6762 Page 1 of 10 The life science business of Merck KGaA, Darmstadt, Germany operates as MilliporeSigma in the US and Canada Hazard statement(s) H300 + H310 + H330 Fatal if swallowed, in contact with skin or if inhaled. H361 Suspected of damaging fertility or the unborn child. Precautionary statement(s) P201 Obtain special instructions before use. P202 Do not handle until all safety precautions have been read and understood.
    [Show full text]
  • Identification of New GLUT2-Selective Inhibitors Through in Silico Ligand
    www.nature.com/scientificreports OPEN Identifcation of new GLUT2‑selective inhibitors through in silico ligand screening and validation in eukaryotic expression systems Sina Schmidl1,7, Oleg Ursu2,6,7, Cristina V. Iancu3, Mislav Oreb1, Tudor I. Oprea2,4* & Jun‑yong Choe3,5* Glucose is an essential energy source for cells. In humans, its passive difusion through the cell membrane is facilitated by members of the glucose transporter family (GLUT, SLC2 gene family). GLUT2 transports both glucose and fructose with low afnity and plays a critical role in glucose sensing mechanisms. Alterations in the function or expression of GLUT2 are involved in the Fanconi– Bickel syndrome, diabetes, and cancer. Distinguishing GLUT2 transport in tissues where other GLUTs coexist is challenging due to the low afnity of GLUT2 for glucose and fructose and the scarcity of GLUT‑specifc modulators. By combining in silico ligand screening of an inward‑facing conformation model of GLUT2 and glucose uptake assays in a hexose transporter‑defcient yeast strain, in which the GLUT1‑5 can be expressed individually, we identifed eleven new GLUT2 inhibitors (IC50 ranging from 0.61 to 19.3 µM). Among them, nine were GLUT2‑selective, one inhibited GLUT1‑4 (pan‑Class I GLUT inhibitor), and another inhibited GLUT5 only. All these inhibitors dock to the substrate cavity periphery, close to the large cytosolic loop connecting the two transporter halves, outside the substrate‑binding site. The GLUT2 inhibitors described here have various applications; GLUT2‑specifc inhibitors can serve as tools to examine the pathophysiological role of GLUT2 relative to other GLUTs, the pan‑Class I GLUT inhibitor can block glucose entry in cancer cells, and the GLUT2/GLUT5 inhibitor can reduce the intestinal absorption of fructose to combat the harmful efects of a high‑fructose diet.
    [Show full text]
  • In Vitro Micronucleus Test
    DRAFT GUIDELINE (2nd version) 21 December 2006 OECD GUIDELINE FOR THE TESTING OF CHEMICALS DRAFT PROPOSAL FOR A NEW GUIDELINE 487: In Vitro Micronucleus Test INTRODUCTION 1. The in vitro micronucleus assay is a genotoxicity test system used for the detection of micronuclei in the cytoplasm of interphase cells. These micronuclei may originate from acentric fragments (chromosome fragments lacking a centromere) or whole chromosomes that are unable to migrate with the rest of the chromosomes during the anaphase of cell division. The assay detects the activity of both clastogenic and aneugenic chemicals (1, 2) in cells that have undergone cell division after exposure to the test substance. Development of the cytokinesis-block methodology, by addition of the actin polymerisation inhibitor cytochalasin B during the targeted mitosis, allows the identification and selective analysis of micronucleus frequency in cells that have completed one cell division as such cells are binucleate (3, 4). However, the Test Guideline also allows the use of protocols without cytokinesis block, provided there is evidence that the majority of cells analysed are likely to have undergone cell division. 2. The immunochemical labelling (FISH) of kinetochores, or hybridisation with general or chromosome specific centromeric/telomeric probes can provide useful information on the mechanism of micronucleus formation (5). Use of cytokinesis block facilitates the acquisition of the additional mechanistic information (e.g., chromosome non-disjunction) that can be obtained by FISH-techniques (6- 15). 3. The micronucleus assay has a number of advantages over metaphase analysis performed to measure chromosome aberrations in the OECD Guidelines 473 and 475 (16, 17).
    [Show full text]
  • With Deuterium Oxide, Colchicine, and Cytochalasin B
    Histamine Release from Human Leukocytes: Studies with Deuterium Oxide, Colchicine, and Cytochalasin B ELIZABETH GLLESPIE and LAWRENCE M. LICHTENSTEIN From the Division of Clinical Immunology, The Johns Hopkins University School of Medicine at The Good Samaritan Hospital, Baltimore, Maryland 21239 A B S T R A C T Agents known to interact with either INTRODUCTION microtubules or microfilaments influenced the antigen-in- Antigen-mediated release of histamine from the duced release of histamine from the leukocytes of al- leuko- lergic individuals. Deuterium oxide (D20) cytes of allergic individuals is a good in vitro model for which sta- allergic reactions. The extent of bilizes microtubules and thereby favors their formation release correlates well enhanced histamine release with clinical symptoms (1) and drugs, e.g., isoproterenol markedly. Concentrations and theophylline, which alleviate the as low as 5% increased antigen-induced release somne- in vivo symptoms also inhibit the in vitro release what while concentrations as high as 75% had no effect process (2). At the same time the mechanism by which histamine is released from on release in the absence of antigen. Enhancement oc- leukocytes curred over a wide range of antigen is fundamentally similar to the release process concentrations and in other tissues, with cell-bound was also seen when release was initiated by antibody (IgE) acting antibody to as a IgE or IgG. When the release process was divided into receptor and antigen replacing a hormone or other two stages a D20 activity could be demonstrated only releasing agent. in the second stage. However, when D20 was present in Colchicine, a drug known to disrupt microtubules, has the first stage together with agents which raise cyclic been shown to inhibit the release of a variety of com- AMP levels and thereby inhibit release it partially re- pounds (3-6) including histamine from leukocytes (7).
    [Show full text]
  • Visualization of a System of Filaments 7-10 Nm Thick in Cultured Cells of An
    Proc. Natl. Acad. Sci. USA Vol. 74, No. 6, pp. 2490-2494, June 1977 Cell Biology Visualization of a system of filaments 7-10 nm thick in cultured cells of an epithelioid line (Pt K2) by immunofluorescence microscopy (intermediate filaments/tonofilaments/keratin/autoantibodies/mitotic drugs) MARY OSBORN*, WERNER W. FRANKEt, AND KLAUS WEBER* * Max Planck Instittit fiir Biophysikalische Chemie, G6ttingen; and t Deutsches Krebsforschungszentrum, Heidelberg, Federal Republic of Germany Communicated by J. Brachet, April 5, 1977 ABSTRACT During our studies with antibodies against MATERIALS AND METHODS structural proteins of the cytoskeleton of eukaryotic cells we have observed that sera from many normal rabbits decorate a Pt K2 cells, originally derived from rat kangaroo kidney epi- fiber system in cells of the established rat kangaroo cell line Pt thelium and Nettro 2a cells, a neuroblastoma line from mouse, K2. The display and organization of these fibers are different were from the American Type Culture Collection. Pt K2 cells from those of microfilament bundles (decorated by antibody were maintained in Eagle's minimal essential medium sup- to actin) and microtubules (decorated by antibody to tubulin). plemented with nonessential amino acids, glutamine, sodium This new fiber system can be further distinguished by its resis- tance to reorganization when cells are treated with Colcemid pyruvate, and 10% (vol/vol) fetal calf serum. Neuro 2a cells or cytochalasin B. The decoration of this fiber system is not were grown in Dulbecco's modified Eagle's medium containing detected if Pt K2 cells are fixed with formaldehyde. Such sera 10% fetal calf serum. Growth conditions for mouse 3T3 cells also appear to decorate swirls of perinuclear fibers in mouse have been described (6).
    [Show full text]
  • (Bhk-~21) Cells
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by PubMed Central THE EFFECTS OF CYTOCHALASIN B ON THE MICROFILAMENTS OF BABY HAMSTER KIDNEY (BHK-~21) CELLS ROBERT D . GOLDMAN From the Department of Biology, Case Western Reserve University, Cleveland, Ohio 44106 ABSTRACT Attempts were made to test the motile functions of bundles of microfilaments found in baby hamster kidney (BHK-21) cells, by using cytochalasin B (CB) . It was found that individual cells respond differently to the drug . These differential effects are quite obvious in both light and electron microscope preparations . Some cells contain normal bundles of microfilaments even after 24 hr in CB, and other cells form muscle-like configurations which also contain arrays of microfilaments . These varied effects suggest the existence of several types of microfilaments in BHK-21 cells, and make the interpretation of the motif role of microfilaments difficult to evaluate at the present time . INTRODUCTION Cytochalasin B (CB)I (1) causes the disruption of MATERIALS AND METHODS microfilaments in the cleavage furrow of marine Cell Cultures eggs end HeLa cells (2-4), and the reversible dis- organization of microfilaments in several types of BHK-21/C13 cells (11) were grown in BHK-21 embryonic cultures (5-8) . It has been suggested medium supplied by Grand Island Biological Com- that these microfilaments are involved in primi- pany (Grand Island, N . Y.). The medium was sup- plemented with 10%o tryptose phosphate broth, 10%1% tive contractile processes and that they are similar calf serum, and 100 units/ml of penicillin and strep- to actin (8-10) .
    [Show full text]
  • Asbestos Induces Apoptosis of Human and Rabbit Pleural Mesothelial Cells Via Reactive Oxygen Species
    Asbestos induces apoptosis of human and rabbit pleural mesothelial cells via reactive oxygen species. V C Broaddus, … , J D Ernst, A M Boylan J Clin Invest. 1996;98(9):2050-2059. https://doi.org/10.1172/JCI119010. Research Article Mesothelial cells, the progenitor cell of the asbestos-induced tumor mesothelioma, are particularly sensitive to the toxic effects of asbestos, although the molecular mechanisms by which asbestos induces injury in mesothelial cells are not known. We asked whether asbestos induced apoptosis in mesothelial cells and whether reactive oxygen species were important. Pleural mesothelial cells (rabbit or human) were exposed to asbestos (crocidolite, amosite, or chrysotile) or control particles at moderate doses (1-10 microg/cm2) over 24 h and evaluated for oligonucleosomal DNA fragmentation, loss of membrane phospholipid asymmetry, and nuclear condensation. Asbestos fibers, not control particles, induced apoptosis in mesothelial cells by all assays and induction of apoptosis was dose dependent for all types of asbestos, with crocidolite (5 microg/cm2) inducing 15.0+/-1.1% (mean+/-SE; n = 12) apoptosis versus control particles < 4%. Apoptosis induced by asbestos, but not by actinomycin D, was inhibited by extracellular catalase, superoxide dismutase in the presence of catalase, hypoxia (8% oxygen), deferoxamine, 3-aminobenzamide [an inhibitor of poly(ADP-ribosyl) polymerase], and cytochalasin B. Only catalase and cytochalasin B decreased fiber uptake. We conclude that asbestos induces apoptosis in mesothelial cells via reactive oxygen species. Escape from this pathway could allow the abnormal survival of mesothelial cells with asbestos-induced mutations. Find the latest version: https://jci.me/119010/pdf Asbestos Induces Apoptosis of Human and Rabbit Pleural Mesothelial Cells Via Reactive Oxygen Species V.
    [Show full text]
  • A Novel Inhibitor of Glucose Uptake Sensitizes Cells to FAS-Induced Cell Death
    3546 A novel inhibitor of glucose uptake sensitizes cells to FAS-induced cell death Tabitha E. Wood,1,2 Shadi Dalili,1,2 transport as a mechanism to sensitize cells to death Craig D. Simpson,1 Rose Hurren,1 Xinliang Mao,1 receptor stimuli. Thus, fasentin is a novel inhibitor of Fernando Suarez Saiz,1 Marcela Gronda,1 glucose transport that blocks glucose uptake and high- Yanina Eberhard,1 Mark D. Minden,1 Philip J. Bilan,3 lights a new mechanism to sensitize cells to death ligands. Amira Klip,3 Robert A. Batey,2 [Mol Cancer Ther 2008;7(11):3546–55] and Aaron D. Schimmer1 1Princess Margaret Hospital, Ontario Cancer Institute; Introduction 2Department of Chemistry, University of Toronto; 3Cell Biology Resistance to stimuli of the death receptor pathway of Program, Hospital for Sick Children, Toronto, Ontario, Canada caspase activation can render malignant cells resistant to chemotherapy, immune surveillance, and anchorage-inde- Abstract pendent cell death, a process termed anoikis (1, 2). In addition, such resistance would also limit the therapeutic Evasion of death receptor ligand-induced apoptosis is an utility of agonistic monoclonal antibodies targeting tumor important contributor to cancer development and progres- necrosis factor (TNF) apoptosis-inducing ligand receptors sion. Therefore, molecules that restore sensitivity to death that are now in clinical trial (3). Therefore, small molecules receptor stimuli would be important tools to better that restore sensitivity of tumor cells to TNF family death understand this biological pathway and potential leads receptors would be useful probes to understand this for therapeutic adjuncts. Previously, the small-molecule pathway and potentially useful therapeutic adjuncts for N-[4-chloro-3-(trifluoromethyl)phenyl]-3-oxobutanamide the treatment of malignancy.
    [Show full text]
  • A Light-Microscope Study of the Action of Cytochalasin B on the Cells and Isolated Cytoplasm of the Characeae
    J. Cell Sci. 10, 811-819 (1972) 8ll Printed in Great Britain A LIGHT-MICROSCOPE STUDY OF THE ACTION OF CYTOCHALASIN B ON THE CELLS AND ISOLATED CYTOPLASM OF THE CHARACEAE R.E.WILLIAMSON The Botany School, Downing Street, Cambridge, England SUMMARY The reversible inhibition of cytoplasmic streaming in cells of Nitella translucent by cyto- chalasin B was found to be accompanied by an increase in the number of mini-vacuoles in the endoplasm. Fibrils thought to drive the streaming were still present in inhibited cells. In droplets of cytoplasm obtained from cut cells of Chara corralina, the final number of fibrils formed was not sensitive to cytochalasin B, although the motility of these fibrils was highly so. The movement of organelles in the absence of visible fibrils and the rotation of chloroplasts were also inhibited. The evidence for the involvement of the microfilament system in cyto- plasmic streaming is discussed. The way in which such microfilaments seem to be operating in characean cells differs from the most likely mechanism for their operation in other cytochalasin- sensitive processes. INTRODUCTION Many investigations of cytoplasmic streaming in plant cells have made use of the large size and regular streaming pattern of characean cells. In such cells the interface between the stationary cortical gel and the flowing cytoplasm has been identified as the site at which the motive force for the streaming is generated (Kamiya & Kuroda, 1956). An electron-microscope study (Nagai & Rebhun, 1966) revealed bundles of micro- filaments, each filament being some 5 nm in diameter. The location of these micro- filaments at the interface where the motive force is produced, their orientation parallel to the direction of streaming, and their resemblance to filaments implicated in motile processes in other cells, led to their being favoured as the system responsible for generating the motive force.
    [Show full text]
  • Effects of Cytochalasin B Upon Microfilaments Involved in Morphogenesis of Salivary Epithelium Brian S
    Proceedings of the National Academy of Sciences Vol. 66, No. 2, pp. 360-364, June 1970 Effects of Cytochalasin B upon Microfilaments Involved in Morphogenesis of Salivary Epithelium Brian S. Spooner and Norman K. Wessells DEPARTMENT OF BIOLOGICAL SCIENCES, STANFORD UNIVERSITY, STANFORD, CALIFORNIA Communicated by Colin S. Pittendrigh, March 27, 1970 Abstract. Cytochalasin B causes cultured mouse salivary gland epithelium to lose its characteristic shape and to cease undergoing morphogenesis. The drug causes disorganization of the 50 A microfilaments in epithelial cells that are thought to control cell shape because of contractile properties. Upon removal of cytochalasin, epithelia regain normal shape and resume morphogenesis. Exceptionally large bundles of microfilaments appear in such recovered epithelial cells at points where their contractile activity could account for the changes in cell and tissue shape. Morphogenesis of many developing organs is a manifestation at the population level of alterations in shape of individual cells. Recent evidence has implicated microtubules' and putative contractile microfilaments2 as the intracellular organelles involved in maintenance or change in cell shape. The latter filaments average 40-50 A in diameter and apparently insert in the junctional complex regions of epithelial cells (thus differing from the 100 A filaments that loop through desmosomes). Microfilaments are also found in the microspikes and "fluttering membrane" locomotory organelle of motile cells3 and of elongating axons,4 and in the contractile ring responsible for cytokinesis during cell divi- sion.5 Schroeder6 has found that the contractile ring filaments of cleaving marine eggs disappear after treatment with cytochalasin B,7 and that cytokinesis of such eggs ceases.
    [Show full text]
  • Glucose Transport in Human Skeletal Muscle Cells in Culture. Stimulation by Insulin and Metformin
    Glucose transport in human skeletal muscle cells in culture. Stimulation by insulin and metformin. V Sarabia, … , L A Leiter, A Klip J Clin Invest. 1992;90(4):1386-1395. https://doi.org/10.1172/JCI116005. Research Article Primary human muscle cell cultures were established and the regulation of glucose transport was investigated. Primary cultures were allowed to proceed to the stage of myotubes through fusion of myoblasts or were used for clonal selection based on fusion potential. In clonally selected cultures, hexose (2-deoxy-glucose) uptake into myotubes was linear within the time of study and inhibitable by cytochalasin B (IC50 = 400 nM). Cytochalasin B photolabeled a protein(s) of 45,000- 50,000 D in a D-glucose-protectable manner, suggesting identity with the glucose transporters. In the myotube stage, the cells expressed both the GLUT1 and GLUT4 glucose transporter protein isoforms at an average molar ratio of 7:1. Preincubation in media of increasing glucose concentrations (range 5-25 mM) progressively decreased the rate of 2- deoxyglucose uptake. Insulin elevated 2-deoxyglucose uptake in a dose-dependent manner, with half maximal stimulation achieved at 3.5 nM. Insulin also stimulated the transport of the nonmetabolizable hexose 3-O-methylglucose, as well as the activity of glycogen synthase, responsible for nonoxidative glucose metabolism. The oral antihyperglycemic drug metformin stimulated the cytochalasin B-sensitive component of both 2-deoxyglucose and 3-O-methylglucose uptake. Maximal stimulation was observed at 8 h of exposure to 50 microM metformin, and this effect was not prevented by incubation with the protein-synthesis inhibitor cycloheximide.
    [Show full text]