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  • Wiley-Blackwell  (17,778)
  • 1980-1984  (17,778)
  • 1975-1979
  • 1955-1959
  • 1984  (8,849)
  • 1982  (8,929)
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  • 1980-1984  (17,778)
  • 1975-1979
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  • 1
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 2 (1982), S. 225-242 
    ISSN: 0886-1544
    Keywords: spermatozoa ; calcium ion transport ; motility regulation ; cholinergic agonists ; ouabain ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Behavioral responses of mature spermatozoa treated with neurotropic factors suggest that calcium entry and intracellular transport may be regulated by a cholinergic mediated program. To test the validity of this proposed mechanism, the effect of several agents on Ca distribution in the sperm cell was examined cytochemically.Sites of Ca accumulation were visualized in thin sections of bull spermatozoa by the application of a modification of Gomori's histochemical procedure for phosphatases. Intact bull sperm cells incubated at room temperature in a buffered balanced salt solution containing 5 mM/liter of CaCl2 showed small, randomly scattered deposits of the reaction product. Similarly treated sperm cells, plasmolyzed in hypoosmotic KCl, revealed a greatly increased amount of deposit associated with the cell membranes (mitochondrial surfaces and plasmalemma), the axonemal complex components, and satellite fibers adjacent to the outer dense fibers. Preincubation of intact cells in nicotine or eserine considerably enhanced the entry of calcium into the cell and its association with the membranes and other intracellular organelles. Decamethonium, an irreversible depolarizer and blocker of cholinergic receptors, interfered with the uptake and intracellular distribution of the calcium. Ouabain, the digitalis glycoside that decreases progressive motility of bull sperm and inhibits Na-, K-ATPase, appears to block Ca efflux, causing an intense accumulation of electron-opaque particles in the plasma membrane while smaller numbers of particles are distributed sparsely throughout the cell interior.The cytochemical results showing enhanced calcium entry in the presence of cholinergic agents, depressed intracellular calcium in cells treated with cholinergic receptor blocker, and intense accumulation of calcium within the cell membrane in the presence of ouabain are consistent with spermatozoan behavioral responses to these agents. These observations support the concept that neurotropic factors may be involved in regulating transmembrane and intracellular transport of ions in control of sperm cell function.
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  • 2
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 2 (1982), S. 309-315 
    ISSN: 0886-1544
    Keywords: tropomyosin ; avian muscular dystrophy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The isotype pattern of tropomyosin was investigated in normal and dystrophic avian pectroal muscle using two-dimensional gel electrophoresis. Previous reports have shown that adult pectoral muscle of chickens contains only the α-subunit of tropomyosin and a breast-type troponin-T (TN-T), whereas pectoral fetal muscle contains both α- and β-tropomyosin and leg-type TN-T. The change from the fetal to the adult forms begins shortly after hatching. It has been previously reported that avian dystrophic pectoral muscle contains both the leg- and breast-type TN-T; we show that in avian dystrophic muscle there is also persistent expression of the β-subunit of tropomyosin.
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  • 3
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 2 (1982) 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 4
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    Cell Motility and the Cytoskeleton 2 (1982), S. 287-308 
    ISSN: 0886-1544
    Keywords: actin-binding protein ; Dictyostelium ; cytoskeleton ; amoeboid movement ; calcium ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A protein from Dictyostelium discoideum with an apparent subunit molecular weight of 95,000 daltons (95K protein) was previously identified as an actin-binding protein ‘Hellewell and Taylor, 1979’. In this paper, we present a method for purifying the protein, and characterize some important aspects of its structure and function. Purification of the 95K protein is achieved by fractionation with ammonium sulfate followed by chromatography on DEAE-cellulose, gel filtration on 6% agarose, and final purification on hydroxyapatite. The 95K protein is a dimer, composed of apparently identical subunits. It is a rod-shaped molecule, 38 nm in length, with a Stokes radius of 74 Å. In these structural properties, the 95K protein is similar to muscle and nonmuscle α-actinins. The 95K protein and filamin are equally competent, when compared on a weight basis, to enhance the apparent viscosity of actin as determined by falling ball viscometry. The apparent viscosity of mixtures of the 95K protein and actin is dramatically reduced at pH greater than 7.0 or free ‘Ca2+’ greater than 10-7 M. We also examine the mechanism by which calcium regulates the interaction of the 95K protein and actin. A change in free ‘Ca2+’ induces no detectable change in the quaternary structure of the 95K protein. Our experiments indicate that the 95K protein does not dramatically alter the length distribution of actin filaments in the presence of micromolar free ‘Ca2+’. A large fraction of the 95K protein cosediments with actin in the presence of low free ‘Ca2+’ (ca. 3 × 10-8M), but not in the presence of high free ‘Ca2+’ (ca. 4 × 10-6M). We conclude that increased free ‘Ca2+’ inhibits gelation of actin by the 95K protein by reducing the affinity of the 95K protein for actin. We propose that 95K protein is an important component of the cytoskeletal/contractile system in D. discoideum amoebae.
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  • 5
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    Cell Motility and the Cytoskeleton 2 (1982), S. 343-354 
    ISSN: 0886-1544
    Keywords: NBD-phallacidin ; actin ; ocular tissues ; wound repair ; stress fibers ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The fluorescent derivative of the actin-binding toxin phallacidin, 7-nitrobenz-2-oxa-1,3 diazole phallacidin, has been used to cytologically demonstrate the presence of actin in lens epithelium, corneal endothelium, and retinal pigment epithelium. In these noninjured tissues, no stress fibers are observed and fluorescence is confined mainly to an area at or near the cell membrane, although some diffuse cytoplasmic staining can also be seen. However, following injury to either the lens epithelium or corneal endothelium of rats and frogs, stress fibers are detected, but only in those cells that migrate into the wound area. Cells on the periphery of each tissue do not partake in would repair and thus maintain their normal appearance. After the tissue has regenerated, stress fibers disappear, and those cells involved in the injury response return to their normal morphology.When rabbit corneal endothelium is placed in tissue culture, stress fibers are observed as the cells migrate away from the initial explant. Upon reaching confluency, these cells spread out and each is surrounded by thick actin-containing bands. Furthermore, they exhibit some stress cables within their cytoplasm. This is in contrast to their appearance in vivo where stress fibers are absent and fluorescence is limited to a region near the cell membrane.
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  • 6
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    Cell Motility and the Cytoskeleton 2 (1982), S. 369-383 
    ISSN: 0886-1544
    Keywords: motility ; flagella ; cilia ; microtubules ; Gregarines ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The male gametes of the parasitic protozoan, Lecudina tuzetae, have a motile flagellum with a “6 + O” ultrastructure ‘Schrével and Besse, 1975’. These gametes were isolated from the cysts in which they develop and were observed and photographed under a variety of conditions. The flagella beat continuously, without stopping and starting, with a beat period of about 2 sec. They can beat in solutions whose viscosities are greater than 0.5 Nsm-2 (l Nsm-2 = 103 cP). The waveform can be approximated by a series of helical arcs and interconnecting straight regions that travel from the base to the tip. The helical regions have a radius of curvature of 3.2 μm and subtend a final angle of 1.7 radians. The straight portions are 2.0 μm in length. There are two sets of opposing bends, but they do not originate in the same plane. The resulting waveform is an approximately helical coil, with a pitch of 9.8 μm, a pitch angle of 0.6 radian and a peak-to-peak amplitude of 2.3 μm. The sense of the coil is left handed. The axoneme twists during beating. The main differences between the movement of this flagellum and that of typical 9 + 2 flagella are a low beat frequency and three-dimensional bends that produce relatively little forward movement of the cell. Twisting is discussed as a means of discriminating between some types of models of flagellar motility.
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  • 7
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    Cell Motility and the Cytoskeleton 2 (1982), S. 429-443 
    ISSN: 0886-1544
    Keywords: 21S dynein ; tubulin ; binding stoichiometry ; ATP sensitivity ; binding cooperativity ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The binding properties of Tetrahymena 21S dynein to doublet A and B subfiber microtubules were analyzed by both a turbidimetric assay (Δ A350 nm) and electron microscopy. KCl-extracted, sucrose-gradient, purified 21S dynein binds to each of the two kinds of axonemal microtubules in both ATP-insensitive and ATP-sensitive modes, even though only a single type of binding occurs to each of the subfibers in situ. Total dynein bound to axonemal microtubules is a composite of binding that is sensitive to dissociation by ATP and binding that is insensitive to ATP. Each exhibits a different binding profile. Total binding exhibits a sigmoid profile (h = 1.93) and saturates at 1.49 mg D/mg T. ATP-sensitive binding likewise exhibits a sigmoid profile (h = 2.66) but saturates at 1.06 mg D/mg T. Binding occurs with a similar affinity for both A and B subfibers. The Hill coefficient (h) for ATP-sensitive binding implies positive cooperativity between binding events. ATP-insensitive binding was studied independently in 20 μM ATP, 10 μM vanadate, which blocks ATP-sensitive binding. ATP-insensitive binding exhibits a hyperbolic profile (h = 1.0) and likewise occurs along each of the two kinds of axonemal tubules. Binding saturates at 0.87 mg D/mg T. The binding data suggest that the tubulin dimer has conserved both ATP-sensitive and ATP-insensitive binding sites for 21S dynein, even though the sites may not be expressed in vivo.
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  • 8
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    Cell Motility and the Cytoskeleton 2 (1982), S. i 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 9
    ISSN: 0886-1544
    Keywords: flagella ; cilia ; trachea ; microtubules ; crowns ; microtubule assembly ; caps ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The distal tips of the central pair and A-microtubules are capped in mammalian and avian tracheal cilia. The capping structures are similar to those found in protozoan cilia and flagella [Dentler, 1981], and consist of a central microtubule cap that links the central microtubules to the membrane or to the ciliary crown and A-microtubule plugs that insert into the lumen of each of the A-microtubule plugs is bound to the central microtubule cap by distal filaments. The ends of the central and outer doublet microtubules are tightly bound to the cap in both intact and in demembranated and reactivated tracheal cilia. Analysis of the displacement of the microtubule tips in cilia fixed at various bend angles revealed that the displacements of A-microtubules are only partially in agreement with those predicted by the sliding filament model [Satir, 1968]. These results are discussed with respect to the regulation of microtubule sliding in capped cilia and the role of the microtubule capping structures in microtubule assembly.
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  • 10
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    Cell Motility and the Cytoskeleton 2 (1982), S. 19-24 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 11
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    Cell Motility and the Cytoskeleton 2 (1982), S. 165-168 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 12
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    Cell Motility and the Cytoskeleton 2 (1982), S. 199-204 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 13
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    Cell Motility and the Cytoskeleton 2 (1982), S. 71-75 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 14
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    Cell Motility and the Cytoskeleton 4 (1984), S. 129-135 
    ISSN: 0886-1544
    Keywords: amoeboid motion ; chemoattractants ; chemotaxis ; Dictyostelium ; filopodia ; folic acid ; pterins ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Living vegetative D. discoideum amoebae were studied to determine whether their filopodia respond to folic acid, a chemoattractant for these cells. Exponentially growing amoebae (ca. 10 μm diameter) exhibit 5-30 μm long filopodia; at stationary phase, aggregation competent amoebae have numerous multibranched filopodia up to 100 μm long. Folic acid was observed to stimulate production, elongation, and branching of filopodia with its effects progressively changing as the amoebae approach aggregation. Filopodial construction was also found to be dependent upon Mg2+ levels. The significance of these results is discussed with respect to progressive changes within the vegetative phase as well as to the mechanisms of amoeboid movement, pseudopodial activity, and chemotaxis.
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  • 15
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    Cell Motility and the Cytoskeleton 4 (1984), S. 1-5 
    ISSN: 0886-1544
    Keywords: motility ; power output ; muscle ; flagella ; cytokinetic furrow ; mitotic spindle ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Cellular motile systems as diverse as muscle and the mitotic spindle have been compared by their specific power output: the maximum power they develop per unit of engine volume. Striated muscles and flagella have high specific output; their performance is comparable to that of typical automobile engines. The cytokinetic furrow and the mitotic spindle have very much lower specific power output. The furrow's output is 7,000 times lower than muscle and the spindle's is 300,000 times lower. Different macromolecules have been used to generate power in systems with similar output (muscles and flagella) and, conversely, the same macromolecular motor has been used in systems with very different output (muscles and cytokinetic furrows). The common feature amid this diversity is adaptation to a particular biological role, which specific power output reflects very well. High values are found where a powerful, compact engine should be advantageous, while low values are found where precision, not power, matters most.
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  • 16
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    Cell Motility and the Cytoskeleton 4 (1984), S. 76-76 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 17
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    Cell Motility and the Cytoskeleton 2 (1982), S. 73-82 
    ISSN: 0886-1544
    Keywords: desmids ; videomicrography ; photokinesis ; photophobic response ; photophosphorylation ; photosynthesis ; phototaxis ; uncouplers ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The effects of the uncouplers CCCP and DNP on photokinesis, phototaxis, and the photophobic response in the desmid Cosmarium have been studied both in population systems and by videomicrographic, single-cell analysis. Light-dependent motility is specifically inhibited by both uncouplers, indicating that photokinesis is driven by photophosphorylation. In population experiments, phototaxis and accumulations in light traps due to photophobic responses are inhibited by drug concentrations comparable to those that inhibit photokinesis. Analysis of single-cell behavior demonstrated, however, that neither photophobic responses elicited by an increase in light intensity (step-up response) nor by a decrease (stepdown response) are inhibited, as long as the reduced motility allows the organisms to cross a light--dark border. Phototactic orientation is not impaired by DNP in the single cell analysis, but CCCP significantly reduced the degree of orientation. The results indicate that, although chlorophyll is the photoreceptor for all three photoresponses, at least the photophobic response is independent of both the photosynthetic electron transport chain and photophosphorylation.
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  • 18
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    Cell Motility and the Cytoskeleton 2 (1982), S. 91-92 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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  • 19
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    Cell Motility and the Cytoskeleton 2 (1982) 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 20
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    Cell Motility and the Cytoskeleton 2 (1982), S. 93-101 
    ISSN: 0886-1544
    Keywords: cell motility ; singlet microtubules ; dynein ; coccid insect ; aflagellate spermatozoa ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: In this report we demonstrate that in the coccid insects Pseudococcus, Phenacoccus, and Planococcus the whole spermatozoon is made up by a nuclear central core surrounded by two complete and one incomplete turns of concentric microtubule palisades. Microtubules of the outer row are linked by a system of short projections 6 nm long; those of the inner row are linked to each other by similar arms; a second system of 6 nm arms links the tubules of each inner row to those of the respective outer row. All these systems of arms are longitudinally spaced every ∼ 12 nm. The motility of this spermatozoon is due to waves progressing from the posterior extremity to the anterior one. By SDS polyacrylamide gel electrophoresis a group of high molecular weight polypeptides is detected, one of which migrates in coincidence with the A dynein band from sea urchin sperm. Our data suggest that occurrence in coccid spermatozoon of a motility due to singlet tubules-dynein interaction.
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  • 21
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    Cell Motility and the Cytoskeleton 2 (1982), S. 103-113 
    ISSN: 0886-1544
    Keywords: actin ; cleavage ; fluorescein-labeled phalloidin ; microinjection ; phalloidin ; sand dollar eggs ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Effects of microinjection of phalloidin on fertilization and cleavage of sand dollar (Clypeaster japonicus and Scaphechinus mirabilis) eggs were studied. The drug, previously injected into unfertilized eggs, showed no effect on the elevation of the fertilization membrane upon insemination up to an intracellular concentration of 50 μM. However, the movement of the egg pronucleus to the sperm pronucleus was inhibited and the fusion of pronuclei did not occur. The subsequent development no longer took place. When phalloidin was injected into fertilized eggs, the thickness of the cortical layer increased and the microvilli became conspicuous. Both nuclear division and cleavage were inhibited at the intracellular concentration of more than 20 μM, though the latter seemed to be more sensitive to phalloidin than the former.Fluorescein-labeled phalloidin (FL-phalloidin) was injected into eggs in order to investigate F-actin localization by fluorescence microscopy. In both unfertilized and fertilized eggs, FL-phalloidin was localized in the cortical layer within 1 min after injection. It was also localized in the cortical layer as radially oriented rodlike structures when injected into fertilized eggs before the disappearance of the nuclear membrane. No distinct fluorescence was detected in the mitotic apparatus or in the cleavage furrow. FL-phalloidin redistributed gradually into egg cytoplasm. In unfertilized eggs, fluorescent rods were found especially in the egg pronucleus 30 min after injection.
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  • 22
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    Cell Motility and the Cytoskeleton 2 (1982), S. 131-147 
    ISSN: 0886-1544
    Keywords: higher land plant contractile system ; actin activation of myosin ; S-1 decoration of actin ; polymerization of actin ; calcium sensitivity of actomyosin interaction ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: This paper describes the initial isolation of actin- and myosin-like proteins from the cytoplasm of the endocarp tissue cells of the fruit of the tomato, Lycopersicon esculentum. Low ionic strength buffers extracted the 42,000 molecular weight tomato actin in the depolymerized form. Tomato actin can be polymerized in 0.1 M KCl, 2 mM MgCl2 to form 6 nm diameter filaments resembling rabbit skeletal muscle F-actin in their ultrastructure and pattern of decoration with rabbit myosin subfragment-1 (S-1). Tomato F-actin activates the low ionic strength Mg2+ ATPase of rabbit S-1 up to ten-fold. High ionic strength extracts of tomato yield a myosinlike enzyme whose ATPase activity in 0.5 M KCl is maximal in the presence of K+-EDTA and is repressed in the presence of Mg2+. The column-purified enzyme forms a complex with rabbit F-actin, which can be dissociated by Mg2+ ATP. The low ionic strength Mg2+ ATPase of tomato myosin can be activated ten-fold by rabbit actin and up to nineteen-fold by tomato actin. No activation of the tomato myosin by rabbit F-actin occurs in the absence of free calcium ions.
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  • 23
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    Cell Motility and the Cytoskeleton 2 (1982), S. 149-161 
    ISSN: 0886-1544
    Keywords: cell locomotion ; gastrulation ; contact paralysis ; collagen substratum ; serum factors ; morphogenetic movements ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Prospective mesodermal cells of Xenopus laevis gastrulae showed substantial locomotion in vitro, averaging 4.3 μM/min, when dissociated and cultured on a glass surface coated with collagen and fetal bovine serum. The cell translocate by making lamellipodia and filopodia whereas the main cell body remains rounded. When two mesodermal cells made contact with each other, they showed contact paralysis of lamellipodial activity. In contrast, when mesodermal cells contact ectodermal cells, contact paralysis does not occur. Rather, migrating mesodermal cells continue to translocate. The locomotion in vitro appears to mimic that in vivo during gastrulation, because of the similarities of the rate of movement and the cell shape in culture and in embryos. Neither prospective ectodermal cells from gastrulae nor prospective mesodermal cells from blastulae showed locomotion under the same culture conditions.
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  • 24
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    Cell Motility and the Cytoskeleton 2 (1982), S. 191-194 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 25
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    Cell Motility and the Cytoskeleton 4 (1984), S. 431-441 
    ISSN: 0886-1544
    Keywords: dynein ; chromatophores ; permeabilization ; melanosomes ; motility ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Teleost chromatophores are filled with individual pigment granules that rapidly aggregate to the cell center or become dispersed throughout the cytoplasm in response to environmental stimuli. Microtubules appear to be required for pigment aggregation (movement toward the cell center), and recent findings have suggested that a dynein-like ATPase may participate in force production. Based on previous studies, however, it has been argued that pigment aggregation does not require energy directly, a view that supports the involvement of an elastic component in granule movement. To examine this point further, we have reinvestigated the energy requirements for pigment aggregation using both intact cells and detergent-permeabilized cell models of Fundulus melanophores. Poisons of oxidative phosphorylation, namely, 2,4 dinitrophenol and NaCN, reversibly inhibit melanosome aggregation in response to adrenaline. Inhibition of movement results directly from depletion of intracellular ATP, since pigment translocation can be reactivated in permeabilized cells by the addition of exogenous ATP to the lysis buffer. Non-hydrolyzable analogues, including β,γ-imidoadenosine-5′-triphosphate (AMPPNP), β,γ-methylene adenosine-5′-triphosphate (AMPPCP), and ATPγS, will not substitute for ATP in reactivation of movement. Similarly, other nucleotides such as ADP, AMP, GTP, CTP, and ITP, have limited ability to support melanosome aggregation in metabolically poisoned cells subjected to detergent lysis. ATP itself has no effect on intact cells. These results indicate that melanosome aggregation is ATP-dependent and energy-driven, and are consistent with a role for a force-transducing ATPase in particle movement.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 257-272 
    ISSN: 0886-1544
    Keywords: cellular slime mold ; microtubule-organizing centers ; tubulin ; microtubules ; polymerization ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Nucleus associated bodies (NABs) were isolated from Dictyostelium discoideum or Dictyostelium mucoroides and their ability to nucleate microtubules in vitro was examined.NABs were localized at the tapered ends of the nuclei and released from lysed cells in complex with the nuclei. Microtubules radiating from the NAB could also be isolated with the complex under microtubule stabilizing conditions. The ultrastructure of the isolated NAB showed it to be composed of a core structure surrounded by an amorphous matrix.The ability of isolated NABs to nucleate microtubules in vitro was demonstrated by incubation with exogenous brain microtubule protein. Microtubule assembly was easily visualized by dark-field or immunofluorescence microscopy. Polymerization of microtubules seemed to be initiated not from the core structure but from the surrounding matrix.The number of microtubules polymerized from the NAB was directly counted in whole-mount preparations by electron microscopy, which provided a quantitative assay for the NAB activity. The nucleating activity of NAB was quite unstable and its half-life was calculated as about 5 hours. The activity was sensitive to protease digestion and was also temperature sensitive but could be stabilized by addition of glycerol or storage at - 80°C or in liquid nitrogen. These characteristics are analogous to those of the centrosomes in cultured mammalian cells and a possible explanation of their similarity is discussed.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 317-332 
    ISSN: 0886-1544
    Keywords: cytoskeleton ; platelets ; actin-binding protein ; actin ; myosin ; thrombin activation ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: When human blood platelets were immersed in an ice-cold solution containing 1% Triton ×-1200, 40 mM KCl, 10 mM EGTA, 10 mM imidazole-HCl, and 2 mM NaN3 pH 7.0, a flocculent precipitate appeared immediately in the tube. This precipitate was collected at 3,000g and SDS-polyacrylamide gel analysis showed it to consist mainly of actin, α-actinin, actin-binding protein (ABP), and varying amounts of myosin.Any modifications of this solution used to isolate the platelets' Triton-insoluble cytoskeleton caused profound changes in the nature of the cytoskeleton isolated. Increasing the KCl concentration resulted in a lower yield of cytoskeletal actin and ABP. Inclusion of EDTA in the solution resulted in an increased amount of myosin associated with the cytoskeleton, whereas including MgATP decreased the myosin yield.Experiments with the purified proteins showed that ABP and myosin can each protect the actin from depolymerizing when dialyzed into the Triton solubilization solution. In addition, it was found that when platelets were stimulated with thrombin for 2 min prior to the addition of the Triton solution, 3-4 times more myosin was associated with the cytoskeletal precipitate.The results suggest, therefore, that any variations in solution conditions used for isolating the cytoskeleton from resting platelets, which results in alterations in the amount of ABP, may have profound effects on the state of actin polymerization. Likewise, in thrombin-activated platelets, it is suggested that the increased association of myosin with the cytoskeleton results in a greater stabilization of the F-actin associated with the cytoskeleton. These factors must be considered when interpreting the results regarding the nature of actin transformations in the resting and activated platelet.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 385-391 
    ISSN: 0886-1544
    Keywords: ciliated cell ; basal body apparatus ; microtubules ; microfilaments ; respiratory epithelium ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: This is a descriptive study showing the three-dimensional interrelationship of cytoskeletal elements at the apex of ciliated cells of rat respiratory epithelium. Tissue specimens were serially thin sectioned in various planes and examined by transmission electron microscopy. Thicker sections were also cut at various angles and analyzed stereoscopically. Other specimens were cleared of soluble molecules by glycerination or Triton-X 100 treatment and sectioned as described above. It was found that C microtubules from the triplets of each basal body diverge from the A and B microtubules, run a short distance, and converge at the basal foot. These microtubules or other microtubules arising anew then dispersed deeper into the cytoplasm. The C fibers also interdigitated with other microtubules running perpendicular to them and parallel to the ciliated surface. Ten-nanometer intermediate filaments were organized in parallel sheets between adjacent basal bodies. Sixnanometer actin filaments were distributed throughout the apical cytoplasm. Neighboring basal bodies were linked to one another by microtubules and microfilaments. Basal bodies from each cell appear to be structured for stability, flexibility, and arranged to operate as a single unit.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 445-455 
    ISSN: 0886-1544
    Keywords: clot structure ; platelet contractility ; protein networks ; rheological techniques ; viscoelasticity ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: When citrated plasma is recalcified, it forms a viscoelastic gel-a clot. The relationship between platelet contractility and clot rigidity was studied by using a rheological technique which simultaneously measured both the dynamic rigidity modulus and the contractile force during gel formation with platelet rich plasma (PRP). Protein network formation in a clot was accompanied by a contractile force throughout the clotting process. PRP demonstrated a maximum elastic modulus of 6,000 dynes/cm2 and a maximum contractile force/area of 1,500 dynes/cm2. The values of these parameters for a platelet-free clot (PFP) were 700 dynes/cm2 and less than 100 dynes/cm2 respectively. Sonicated control PRP and PRP from a Glanzmann thrombasthenia patient both clotted in a manner similar to PFP. Metabolic inhibitors, 2-deoxy-D-glucose and KCN (5 mM each), retarded the clotting curves of PRP. Cytochalasin B and E suppressed both structural rigidity and force generation in a concentration-dependent manner similar to their inhibitory effect on actin polymerization in platelets. Colchicine (2.5 mM) or vinblastine (0.11 mM) did not affect these clotting curves. Thrombi-activated, fixed platelets did not generate any force, nor did they significantly increase clot rigidity. Streptokinase induced a concurrent decrease of both rigidity and force in PRP clots. The elastic modulus of a PFP clot could be increased to 2,500 dynes/cm2 by externally straining the network with an axial force/area of 1,500 dynes/cm2. Our results indicate that clot structure formation in PRP is strongly coupled to the contractile force generated by the platelet microfilament system and that this force modulates clot rigidity.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 483-496 
    ISSN: 0886-1544
    Keywords: Cilia ; Ca ; motor control ; ciliates ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have studied quantitative aspects of ciliary motor responses to membrane depolarization in the ciliate Stylonychia using voltage clamp and high-speed cinematograhpy techniques and employing computer-processing methods for evaluation. Depolarizations beyond 4 mV activate the cirri (compound cilia) which are at rest in the absence of a stimulus. The power stroke of activiated cirri is oriented toward the cell anterior. The frequency and duration of beating increase with rising depolarization. With very large positive stimuli (≥ 150 mV) activation of the response is delayed until the end of the voltage step (“off-response”). The peak frequecy is essentially unaltered during sustained depolarization. The frequency drops exponentially following repolarization of the membrane. The time constant of the decay in ciliary activity rises with the amplitude, not with the duration of the depolarization. The ciliary motor response is most adequately represented by the number of evoked ciliary cycles (ciliary work), and appears to be related to the amplitude of the depolarization.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 497-508 
    ISSN: 0886-1544
    Keywords: cilia ; electric motor control ; ciliates ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have studied the motor responses to membrane hyperpolarization of the marginal cirri in Stylonychia using voltage-clamp, high-speed cinematography, and computer-processing techniques. The cirri started beating when voltage step amplitudes rose beyond 5 mV. The power stroke was oriented toward the posterior cell and (hyperpolarizing motor activation). The frequency rose slightly during a voltage step, and decreased with similar rates for 100 ms following the step end. Amplitude and duration of the step tended to increase the motor response of the cirri. The late response declined exponentially. The time constant of the decay rose with the step amplitude. Among three response parameters tested (frequency, duration, number of cycles), the number of evoked ciliary cycles was best correlated with the amplitude of the hyperpolarization. Comparisons with the responses to depolarizing voltage steps reveal similarities in the relaxation of ciliary activity which appears to be uncoupled, in part, from the electric membrane events during the voltage stimulus.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 573-582 
    ISSN: 0886-1544
    Keywords: Euglena flagella ; laser microsurgery ; stigma ; Mg2+ ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: When the area of the stigma of Euglena was irradiated with an infrared laser beam at a dose too low to cause permanent loss of motility, a reduction in flagellar motility was observed only when the external medium contained less than 1 mM Mg2+. At these low Mg2+ concentrations, the laser caused a decrease in flagellar frequency and a tendency for the flagellar waveform to shift towards that taken during reversed swimming. This suggests that the effect of the laser irrdiation was to deplete the cells of Mg2+. After the laser pulse the reversal response remained sensitive to the wavelength of the illuminating light. In white light (420-700 nm) 60% of the Euglena showed a reversed waveform; in orange light (530-700 nm) this increased to 90%. This shows that the photoreceptor was not destroyed by the laser irradiation.These experiments were performed on cells that had been impaled on a microelectrode. If direct electric current was passed into the laser-irradiated cells, the current necessary to cause flagellar arrest was 2 to 4 times less than that for cells not laser irradiated.It is concluded that an internal Mg2+ store is present in the Euglena, localized in the area of the paraflagellar swelling; and that the laser irradiation eliminates this Mg2+ store, but at the power used it does not destroy the ability of the stigmaparaflagella to control the flagellar activity.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 1-5 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 107-112 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 149-152 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 175-180 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 87-93 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 121-126 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 113-119 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 133-136 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982) 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 4 (1984), S. 25-27 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 4 (1984), S. 41-55 
    ISSN: 0886-1544
    Keywords: Leptodiscinae ; Dinoflagellates ; contractility ; non-actin filaments ; Life and Medical Sciences ; Cell & Developmental Biology
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    Topics: Biology , Medicine
    Notes: The Leptodiscinae, a group of marine Dinoflagellates, are good material for the study of contraction though they cannot be collected in abundance. Their cell bodies are flattened anteroposteriorly (Leptodiscus, Leptophyllus, and Leptospathium) and are able to contract suddenly when the surrounding water is disturbed.Electron microscopical observations have shown that the structures responsible for the contraction consist of a layer of parallel filaments located beneath the cell membrane of some specialized parts of the body. These filaments seem to be nonactin (NAF) because of their diameter (2.5-3 nm) and because they are not decorated by heavy meromyosin (HMM). They appear helically coiled and doubly twisted, and form tubular structures when contracted.
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    Cell Motility and the Cytoskeleton 4 (1984), S. 77-87 
    ISSN: 0886-1544
    Keywords: Chlamydomonas ; flagella ; cell surface ; adhesion ; glycoproteins ; iodination ; lactoperoxidase ; Iodogen ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The Chlamydomonas flagellar surface exhibits interesting adhesive properties that are associated with flagellar surface motility. This dynamic surface property can be exhibited as the binding and movement of small polystyrene microspheres or as the interaction of the flagellar surface with a solid substrate followed by whole cell locomotion, termed “gliding.” In order to identify flagellar surface proteins that mediate substrate interaction during flagellar surface motility, two immobilized iodination systems were employed that mimic the conditions for flagellar surface motility: small polystyrene microspheres derivatized with lactoperoxidase, and large glass beads derivatized with Iodogen. Use of these iodination conditions resulted in preferential iodination of a high-molecular-weight glycoprotein with apparent molecular weight of 300,000-350,000. These results suggest this glycoprotein as a major candidate for the surface-exposed adhesive component that directly interacts with the substrate and couples the substrate to a system of force transduction presumed to be located within the flagellum.
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    Cell Motility and the Cytoskeleton 4 (1984) 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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  • 46
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    Keywords: fibroblast ; permeabilized cell model ; Ca2+-dependent contraction ; calmodulin ; phosphorylation ; myosin light chain ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Human lung fibroblast MRC-5 cells treated with Triton X-100 (MRC-5 cell models) were able to contract in the presence of MgATP and Ca2+ of more than 1 μM. Immunofluorescence microscopy with antibodies to actin and myosin 20,000-dalton (20 Kd) light chain revealed that stress fibers were prominent in MRC-5 cell models. Use of a fluorescent actin probe, 7-nitrobenz-2-oxa-1,3-diazole-phallacidin permitted visualization of contraction of the stress fibers in the presence of MgATP and Ca2+. Of the proteins in MRC-5 cell models, only a myosin 20 Kd light chain was phosphorylated in a Ca2+-dependent manner. This Ca2+-dependent phosphorylation of the 20 Kd light chain closely corresponded with the contraction of MRC-5 cell models: 1) Both phosphorylation of the 20 Kd light chain and contraction of MRC-5 cell models were inhibited by calmodulin antagonists such as N-(6-aminohexyl)5-chloro-1-napthalene sulfonamide. 2) The threshold Ca2+ concentration for phosphorylation of the 20 Kd light chain was similar to that for contraction of MRC-5 cell models. Both were lowered by exogenous calmodulin in a concentration-dependent manner. 3) The 20 Kd light chain was thiophosphorylated by incubation of MRC-5 cell models with an ATP analogue, adenosine 5′-0-(3-thiotriphosphate) only in the presence of Ca2+. After this treatment, MRC-5 cell models lost the Ca2+-dependence for contraction. These results indicate that Ca2+-calmodulin-dependent phosphorylation of myosin 20 Kd light chain is required for contraction of MRC-5 cell models.
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    Cell Motility and the Cytoskeleton 4 (1984), S. 387-401 
    ISSN: 0886-1544
    Keywords: bull sperm flagella ; motility ; time course ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Detailed measurements were made of the time course of the motion of bull spermatozoa. Fourier analysis of the data showed the time course to be basically sinusoidal within 2% to 3%. An asymmetry in the motion was present, resulting in a second harmonic component in the Fourier spectra of normal sperm of approximately 11% of the main component. When the energy metabolism of the sperm was inhibited or when the external viscosity of the medium was raised, the asymmetry was reduced. When the internal Mg2+ content of the sperm was lowered, the asymmetry was increased. The asymmetries and the corresponding second harmonic components in the Fourier spectra were correlated with the overall bend shape of the sperm and with the curvature of the path in which the sperm were swimming. Model calculations showed that the asymmetry could reside in either the internal active moments in the sperms or in the stiffness of the sperm fiagella.
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    Cell Motility and the Cytoskeleton 4 (1984), S. 443-468 
    ISSN: 0886-1544
    Keywords: actin ; microfilaments ; HMM ; phagocytosis ; cytochalasin ; Paramecium ; fluorescence microscopy ; electron microscopy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Using heavy meromyosin (HMM) or the fragment S1 of myosin as probes for actin microfilaments, we studied their organization in Paramecium both by fluorescence and electron microscopy.In interphasic cells, HMM decorates (a) most prominently the periphery of nascent and young food vacuoles and their route during the early phase of their intracellular transit; (b) a thin meshwork radiating from the gullet throughout the cytoplasm; (c) a small area beneath the pore of contractile vacuoles and beneath the cytoproct when open to release food residues. Most of these HMM-decorated structures are in close contact with microtubular arrays. All HMM decoration disappears in dividing cells and in cytochalasin-treated cells. In vivo, the drug immediately blocks food vacuole formation but does not affect cytokinesis, cyclosis, contractile vacuole pulsation, defecation, or nuclear movements.The data show that, as in the cells of other organisms, actin microfilaments form defined arrays that undergo physiologically controlled cycles of assembly/disassembly. These arrays contribute (at least in the phagocytotic process) to diverse types of movement: constriction, membrane fusion, and migration of food vacuoles. However, aside from their massive concentration along the phagocytotic tractus, actin microfilaments are neither major structural components of Paramecium cytoplasm nor the only cytoskeletal components ensuring motility or contractility processes.
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    Cell Motility and the Cytoskeleton 4 (1984), S. 197-213 
    ISSN: 0886-1544
    Keywords: gelation ; actin ; filamin ; cytoplasm ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have compared the meniscus depletion assay and falling ball viscometry, two means of assessing the extent of gelation in actin-based systems using mixtures of actin and the actin-binding protein filamin. We examined the effect of varying the concentrations of actin and filamin in both assays. The interaction of actin and filamin was detected only above a threshold concentration of filamin. This threshold concentration was lower for falling ball viscometry than for the meniscus depletion assay at equal actin concentrations. At constant concentrations of filamin, an increase in actin concentration caused an increase in apparent viscosity measured by the falling ball assay, but a decrease in sedimentability detected by the meniscus depletion assay. The rate of sedimentation of actin was dependent on the molar ratio of actin to filamin. At each molar ratio, the sedimentation of actin was not dependent on the specific concentrations of actin and filamin used. The apparent viscosity was dependent on both the molar ratio and the specific concentrations of actin and filamin. To relate the present results to earlier studies, we examined mixtures of actin and filamin using a macroscopic assay of gelation (tube tipping assay), and polarized light microscopy. The effect of increasing filamin concentration in the four assays was compared at three actin concentrations. Mixtures of actin and filamin whose apparent viscosities were low enough to be estimated by falling ball viscometry were optically isotropic fluids that flowed out of inverted test tubes. Mixtures of actin and filamin in the range of sensitivity of the meniscus depletion assay were either viscous fluids or gels, and were either optically isotropic or anisotropic. Thus, the four assays provide different estimates of gelation. Both the meniscus depletion assay and falling ball viscometry can be used to determine relative gelation activity, but neither can be used as a quantitative assay of gelation.
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    Cell Motility and the Cytoskeleton 4 (1984), S. 183-196 
    ISSN: 0886-1544
    Keywords: tubulin ; assembly ; mitotic apparatus ; bimane ; fluorescence microscopy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Fluorescent derivatives of cellular proteins that retain their native characteristics have become useful probes to investigate the dynamics of specific cytoskeletal proteins. In the experiments reported here, a previously characterized fluorescent derivative of tubulin, bimane-tubulin [Wadsworth and Sloboda, 1982a], was used to investigate microtubule assembly in vitro. The results demonstrate that bimanetubulin was competent to assemble onto a variety of organizing centers in vitro, including microtubule organizing centers (MTOCs) present in homogenates of sea urchin eggs, isolated mitotic apparatuses (MAs), and lysed mitotic cells. When homogenates of fertilized sea urchin eggs containing MTOCs were incubated with bimane-tubulin at 37°C, discrete areas of linear fluorescence were observed. Only diffuse fluorescence was observed when calcium or colchicine was added to the homogenate or if the temperature was maintained at 0°C. Negative-stain electron microscopy of the fluorescent arrays revealed morphologically normal microtubules radiating from electron dense regions. When mitotic spindles, isolated in glycerol containing buffers and therefore cold stable, were incubated with bimane-tubulin, linear fluorescence was observed emanating from the spindle poles but not from the region occupied by the kinetochores. MAs incubated with bimane-labeled bovine serum albumin or bimane-labeled microtubule-associated proteins showed only diffuse fluorescence. However, when mitotic cells which were hypotonically lysed in the absence of detergents or microtubule stabilizing solvents, were perfused with bimane-tubulin intense fluorescence was observed in the asters and throughout the spindle. Two experiments suggested that the fluorescence observed in the results outlined above was due to the assembly of normal microtubules from the fluorescent subunits. First, the observed fluorescence was sensitive to cold temperataure, which is known to disassemble microtubules. Second, when the isolated, fluorescent MAs were examined by thin section electron microscopy, microtubules of normal diameter were seen. No aggregated material appeared associated with the walls of the microtubules, which might have been expected if the fluorescent protein was nonspecifically adsorbed to the microtubules. The results of these experiments demonstrate that isolated, stabilized MAs support the growth of new microtubules from the spindle poles while labile spindles, present in lysed cells, incorporate fluorescent tubulin throughout the spindle and asters. The significance of these results for hypotheses concerning microtubule assembly and disassembly during mitosis is discussed.
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    Cell Motility and the Cytoskeleton 4 (1984), S. 241-247 
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    Keywords: cytoskeleton ; centrosome ; tonofilaments ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We present observations on the relative location of the centriole and keratin filament cap in motile PtK1 cells. Subconfluent cells were double labeled with anticentriole and antikeratin sera. These preparations revealed that the centriole is separate from, but neighboring, the keratin filament cap. Serial ultrathin sections confirm this observation. These observations are consistent with the idea that the microtubule organizing center and intermediate filament distribution center are not identical or concentric in PtK1 cells.
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    Cell Motility and the Cytoskeleton 4 (1984), S. 403-404 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 4 (1984), S. 417-430 
    ISSN: 0886-1544
    Keywords: flagella ; image analysis ; microcomputer ; motility ; parameter estimation ; Simplex method ; spermatozoa ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Parameters to describe flagellar bending patterns can be obtained by a microcomputer procedure that uses a set of parameters to synthesize model bending patterns, compares the model bending patterns with digitized and filtered data from flagellar photographs, and uses the Simplex method to vary the parameters until a solution with minimum root mean square differences between the model and the data is found. Parameters for Chlamydomonas bending patterns have been obtained from comparison of shear angle curves for the model and the data. To avoid the determination of the orientation of the basal end of the flagellum, which is required for calculation of shear angles, parameters for sperm flagella have been obtained by comparison of curves of curvature as a function of length for the model and for the data. A constant curvature model, modified from that originally used for Chlamydomonas flagella, has been used for obtaining parameters from sperm flagella, but the methods can be applied using other models for synthesizing the model bending patterns.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 173-181 
    ISSN: 0886-1544
    Keywords: cytoplasmic streaming ; motive force ; mitotic cycle ; Physarum polycephalum ; migration ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The relationship between mitosis and cytoplasmic streaming in the plasmodium of Physarum polycephalum was investigated by simultaneously conducting the following three experiments: (1) identification of the mitotic stages under phase contrast optics, (2) measurement of the rate of oriented migration of the plasmodium on an agar ribbon, and (3) measurement of the motive force of cytoplasmic streaming by the double-chamber method of Kamiya.The migration of the plasmodium almost stopped during synchronous mitosis and the motive force of the flow decreased to 1/4 of the normal level in this period. Gelation of the endoplasm did not occur during the mitotic period, and thus the cessation of the plasmodial migration must have been caused by the diminution of the motive force responsible for cytoplasmic streaming.
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    Cell Motility and the Cytoskeleton 2 (1982) 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 183-195 
    ISSN: 0886-1544
    Keywords: crane flies ; meiosis ; spermatocytes ; chromosome movement ; nuclear envelope ; prophase ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Meiotic prophase in spermatocytes of the crane fly, Nephrotoma suturalis, involves both the condensation and the movement of bivalent chromosomes. Since crane flies have only four bivalents that appear highly condensed during late prophase, changes of position and orientation of those bivalents relative to one another can be seen easily in living cells. Chromosome movement during the final 1 to 2 hr of diakinesis was analyzed in detail. Maximal velocities of prophase movements were between 0.1 and 1 μM/min. Metakinetic movements during prometaphase have similar velocities. To assess the physiological basis of prophase movements, experiments employing cyanide and cold treatment were performed. Prophase movements were abolished completely by cyanide, and, for the most part, the velocities of chromosomes in the cold at 2°C and 6°C were less than that of untreated cells at 22°C. The results suggest that prophase movements are energy dependent and may involve an enzyme-catalyzed process occurring in close association with the nuclear envelope.
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    Keywords: intermediate filament ; desmosomes ; epidermal keratinocytes ; nuclear envelope ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have shown, by indirect immunofluorescence microscopy using an antiserum against the mouse keratin subunit K2 and by electron microscopy, that transformed (PAM) and primary (PME) mouse epidermal cells possess extensive net works of IF bundles. Following trypsinization and replating of PAM cells, IF bundles are seen to move as a continuous net work from a perinuclear zone into the peripheral cytoplasmic regions. In PAM cells lysed in high-ionic-strength solutions containing Triton ×-100 and DNAase-1, IF bundles appear to be closely associated with nuclear envelope remnants and, in some cases, appear to be attached to nuclear pore complexes. PME cells cultivated in low Ca2+-containing medium possess perinuclear birefringent arrays of IF bundles. Within 2 hours of switching the cells to normal Ca2+ levels, the PME IF bundle network moves towards and establishes contact with the cell surface as desmosomes form. Live cells observed by phase contrast and fixed cells observed by immunofluorescence microscopy demonstrate that desmosomes can be distinguished as dark bands separating neighboring cells. There is little difference between the major proteins seen in SDS-polyacrylamide gel profiles of isolated IF bundle net works from PME cells before and after the Ca2+ switch. Therefore, a reorganization of relatively insoluble membrane-associated protein following the Ca2+ switch may be involved in desmosome formation. The isolated IF networks from PAM cells differ in protein composition compared to the PME IF networks. This may be related to the greatly reduced number of desmosomes in PAM cells. The IF bundle system in epidermal cells appears to be involved in shape formation, shape maintenance, the establishment of desmosomes, nuclear centration, and cell-cell contact.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 243-255 
    ISSN: 0886-1544
    Keywords: directional macrophage motility ; electric fields ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Extracellularly applied electric fields (〈 12 V/cm) strongly influence murine resident peritoneal macrophages (Mø) to undergo directional protrusive pseudopodial activity to wards the positive pole of the electric fields in the absence of exogenously applied chemotactic ligands. Internal and external morphological features were not grossly disrupted by the fields. Directional motility induced by the electric fields was inhibited in the presence of 1.0 mM La3+ or 2.5 mM Mg2+ and 5.0 mM EGTA. Effects of the fields were latent in the inhibited cells and directional motility was expressed after termination of the field and removal of the inhibitors. Receptors for the lectins concanavalin A (Con A) and phytohemagglutinin (PHA-L) were uniformly distributed on the surfaces of Mø with no exposure to electric fields. After exposure to the fields, Con A receptors were preferentially distributed on regions of the Mø surface facing the negative pole and PHA-L receptors were preferentially distributed on those regions facing the positive pole. The possibility that directional Mø motility is regulated by the molecular topography of the cell surface is discussed.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 405-427 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Calmodulin (CaM) isolated from freshwater mussel gill has been shown to be present in fractions derived solely from epithelial cells of whole gill tissue. This CaM has been used to activate bovine brain phosphodiesterase (PDE) and the dose dependence of trifluoperazine-dihydrochloride (TFP)-mediated inhibition of activation has been investigated. The dose-response curve yields an apparent K1 of 20 μM and suggests that at low concentration (up to 40 μM) TFP inhibits PDE activity in the presence of Ca2+, not by direct action on the enzyme, but by a shunt that negates the effect of mussel gill CaM.To determine whether CaM is physiologically significant for ciliary activity in the mussel gill, the effect of TFP upon the Ca2+-dependent ciliary arrest response of the lateral (L) cells of the gill has been examined. Detergent-treated permeabilized epithelial L cell models are scored for ciliary activity after transfer to reactivation solutions that vary in their free Ca2+ and TFP content. At 10-5 M free Ca2+ (pCa 5), maximum recovery from arrest is observed for TFP concentrations in the range of 25 to 30 μM. TFP-mediated recovery from arrest is never complete, suggesting that in the presence of sufficient Ca2+ the drug irreversibly damages a certain fraction of axonemes. At pCa 7 the cilia reactivate and 25-30.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 471-482 
    ISSN: 0886-1544
    Keywords: calmodulin ; myosin ; antibody ; immunofluorescence ; amoeba ; Dictyostelium ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A rabbit antiserum was raised against calmodulin from the eukaryotic microorganism Dictyostelium discoideum. In double immunodiffusion experiments, the antiserum formed an immunoprecipitation line with Dictyostelium calmodulin but not bovine brain calmodulin; competition radioimmunoassays showed no cross reactivity between the antiserum and calmodulins from bovine brain and spinach. The calmodulin content of vegetative Dictyostelium amoebae, determined by competition radioimmunoassay, was 0.5 μg/mg protein; similar levels were found in developing cells. The antiserum was used to visualize the distribution of calmodulin in Dictyostelium amoebae by indirect immunofluorescence. Cells were examined under various conditions: in suspension, attached to a substrate, and while phagocytosing yeast cells. In all cases, anticalmodulin staining was concentrated in the cell cortex. Parallel experiments using a monoclonal antibody against Dictyostelium myosin showed that this protein is also enriched in the cortical region of the cell.
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    Cell Motility and the Cytoskeleton 2 (1982) 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. xvii 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 599-614 
    ISSN: 0886-1544
    Keywords: monoclonal antibodies to tubulin ; radioimmune assay ; immunoautoradiography ; Western blots ; immunofluorescence ; tubulin heterogeneity ; eukaryotic flagellar motility ; immunomotility ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Two monoclonal antibodies reactive for α-tubulin but not for β-tubulin have been prepared, characterized in terms of their relative binding to tubulins from differnt sources by a solid-phase binding assay, immunoautoradiography, and indirect immunofluorescence, and utilized to study flagellar motility. Our results demonstrate that α-tubulins from different species, and even from different tissues of the same species, are nonidentical. Especially interesting was the observation that one of the antibodies, Ab2, immunofluorescently stained microtubules of chick embryo fibroblast cells, but was completely unreactive for microtubules of rat kangaroo (PtK2) fibroblasts; a different antibody, Ab1, stained both cell types. Results of these and additional experiments clearly show that Ab1 and Ab2 recognize discrete and different epitopes on α-tubulin.Monoclonal antitubulins Ab1 and Ab2 each inhibited the bend amplitude of reactivated sea urchin spermatozoa without affecting beat frequencies or the ability of the outer doublet microtubules to slide past each other in elastase-digested models. These results, together with those obtained previously using rabbit polyclonal antitubulin antibodies [Asai and Brokaw, 1980], demonstrate that inhibition of bend amplitude is a common property of antitubulin antibodies and is not due to the binding of antibodies to one specific site on the axoneme. Our results suggest that tubulin subunit conformational changes may occur on the outer doublet lattice and may be integrally involved in the mechanism and control of flagellar bending.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 13-18 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 35-39 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 59-65 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 101-106 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 211-215 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982) 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 393-403 
    ISSN: 0886-1544
    Keywords: motility ; Ca2+ ; ionophores ; spirulina subsalsa ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Motility of the marine filamentous cyanobacterium Spirulina subsalsa is both Ca2+ and Na+ dependent, and replacement of Na+ by mannitol arrests it. The data presented suggest that Ca2+ interacts with sites on the surface of the cell membrane. The inhibitory effect of dicyclohexylcarbodiimide (DCCD) hints at the possibility that the role of Ca2+ may be associated with a membrane bound Ca-ATPase. Motility is pH dependent, being nil at pH 〈 6.5 and 〉 10.0, with an optimum at 8.5. Norepinephrine abolishes most of the inhibitory effect of low pH on motility. Ca2+ has an “all-or-none” effect on motility that is triggered at 5 mM. Acetylcholine lowers the threshold of Ca2+ necessary for triggering motility.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 457-470 
    ISSN: 0886-1544
    Keywords: fragmin ; critical actin concentration ; nucleation ; filament growth ; pointed end ; barbed end ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: As reported previously, fragmin isolated form Physarum plasmodia restricts the polymerization of actin to produce short F-actin filaments in the presence of Ca2+ ions. Here it is shown that when actin is polymerized at low concentrations of salts, fragmin increases the critical concentration of actin for polymerization. This effect of fragmin on the critical concentration is independent of the molar ratio of fragmin to actin. The addition of actin monomers onto heavy meromyosin-decorated F-actin fragments treated with fragmin occurs unidirectionally at the pointed end of each fragment. These results suggest that fragmin binds to the barbed ends of F-actin filaments and inhibits association and dissociation of actin monomers at this end. Fragmin accelerates the initial stage of polymerization of actin. When a constant amount of G-actin is polymerized in the presence of small amounts of fragmin, the inverse of the half-polymerization time increases in proportion to the square root of the amount of fragmin added. This means that fragmin acts as a potent promoter of the nucleation step in actin polymerization. Both functions of fragmin-promotion of nucleation and capping at the barbed end of F-actin-require micromolar concentrations of Ca2+.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 509-523 
    ISSN: 0886-1544
    Keywords: cations ; cilia ; dynein ; ATPase ; microtubule sliding ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We recently demonstrated that elevated concentrations (≥ 20 μM) of the dynein substrate MgATP2- inhibit the spontaneous ATP-induced sliding disintegration of isolated, Triton-demembranted Tetrahymena cilia. We have used a turbidimetric assay (ΔA350 nm) and electron microscopy to examine the effect of ATP on sliding disintegration when activated by other divalent cations. Mg2+, Ca2+, and Mn2+ are each capable of activating sliding, but only with Mg2+ and Mn2+ is disintegration inhibited by elevated ATP concentrations (≥ 1 mM). The two major ATPase activities obtained by KCI extraction of Tetrahymena axonemes differ in their cation specificities such that Mg2+ and Ca2+ activate the 21S dynein ATPase with equal efficiency, whereas the 13S axonemal ATPase activity is reduced by ∼ 50% when CaATP2- replaces MgATP2- as substrate. With 1 mM MgATP2- as substrate, 10-7 to 10-2M added CaC12 alleviates the ATP-dependent inhibition of disintegration and likewise represses 13S MgATPase activity. In contrast, free Ca2+ has no effect on either the disintegration response or MgATPase activity. In contrast to Triton-treated cilia, glycerinated cilia, which beat in 1 mM MgATP2-, are inhibited from beating by high CaATP2- concentrations. These substrate specificities suggest that concentration-dependent, substrate inhibition of sliding disintegration may be a manifestation of a physiological mechanism that is mediated by the 13S axonemal ATPase and that may function to modulate sliding during bend formation. However, the effects of added CaCl2 probably do not reflect a physiological mechanism for regulating beat parameters, but rather may result from CaATP2- competing for MgATP2- binding sites on the 13S ATPase, thereby blocking expression of the 13S ATPase.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 7-12 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 25-28 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 47-51 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 77-81 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 95-99 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 525-547 
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    Keywords: cilia ; flagella ; ATPases ; dynein ; Chlamydomonas ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Extraction of isolated, demembranated flagellar axonemes of Chlamydomonas reinhardii with 0.6 M KCl solubilized 77-92% of the total axonemal Mg++ or Ca++-ATPase activity, which sedimented as 18S and 12S peaks in sucrose density gradients. The ATPases of these two peaks were further purified by hydroxyapatite (HAP) column chromatography. The ATPase activity of the 18S peak eluted from the HAP column as a single peak coinciding with the protein peak. The HAP purified 18S ATPase had a specific activity of ∼2.0 ± 0.5 μmoles Pi hydrolyzed min/mg and was associated with four high molecular weight (HMW) polypeptides of ∼ 310,000-340,000 daltons, two intermediate molecular weight (IMW) polypeptides of 78,000 and 69,000 daltons, and eight low molecular weight (LMW) polypeptides of 7,800-19,600 daltons. When the 12S sucrose gradient peak together with a trailing shoulder were chromatographed on HAP, the ATPase activity was eluted in two peaks designated 12S and 10.5S on the basis of the sedimentation properties of their associated polypeptides. The 12S peak contained a single dynein ATPase having a specific activity of ∼ 0.6 ± 0.3 μmoles Pi hydrolyzed min/mg and associated with ∼ 330,000-, 21,700-, and 18, 100-dalton polypeptides. The 10.5S peak contained several high, intermediate, and low molecular weight polypeptides; of these, on HMW polypeptide and one 28,700-dalton polypeptide correlated well with the ATPase activity. The purified ATPases had no polypeptides in common; each therefore represents a discrete dynein. Based on protein recovered in the purified fractions, 18S dynein represents ∼ 9.2% of the total axonemal protein; 12S dynein represents ∼ 4.7% of the axonemal protein.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 185-189 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 225-228 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 137-141 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 4 (1984), S. 169-181 
    ISSN: 0886-1544
    Keywords: cytoskeleton ; motility ; cell spreading ; epithelial cells ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Reorganization of intermediate filaments during cell spreading is examined by immunofluorescence, electron microscopy, and time-lapse video microscopy. A juxtanuclear cap, believed to correspond to the intermediate filament distribution center, was observed to be spatially related to the organization of the intermediate filament network as cells spread. A keratin cap was observed, which appeared spontaneously in motile PtK1 cells. Cap formation may be a consequence of retraction of intermediate filaments from the cytoplasm as cells move. The position of this juxtanuclear cap is related to the direction of movement, located on the side of the nucleus near the advancing edge of the cell. As the cell spreads, the cap disappears as the keratin filament network returns to the cytoplasm. Evidence presented here is consistent with the hypothesis that the distribution center mediates keratin filament organization during cell shape change.
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    Cell Motility and the Cytoskeleton 4 (1984), S. 29-40 
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    Keywords: microfilaments ; microtubules ; contraction ; collagen gel ; fibroblasts ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: In vitro models have been developed recently to study the ability of fibroblasts to generate tensile force within collagen gels. The present study was initiated to assess the role of the cytoskeleton in the cell shape changes and force generation in one such model system. Porcine periodontal ligament fibroblasts (PPLF) were cultured within three-dimensional collagen gels attached to glass coverslips. Fluorescence microscopy, using nitrobenzooxadizole (NBD)-phallacidin labeling for microfilaments and tubulin antibody staining for microtubules, was combined with phase and Nomarski optics to determine the intra- and extracellular architecture of the cells and collagen fibers. Samples were observed from 30 minutes to 24 hours after initiation of cell attachment. During attachment and spreading, NBD-phallacidin staining changed dramatically until large microfilament bundles became prominent. Collagen fiber alignment, compaction, and finally tearing from the coverslip occurred during this time. After release of tension, microfilament bundles were no longer evident. The change in microtubule distribution during these processes was less dramatic, appearing to follow the change in cell shape. These results indicate that microfilaments play an essential role in generating force to align and compact collagen, while microtubules may have a secondary role only.
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    Cell Motility and the Cytoskeleton 4 (1984), S. 57-71 
    ISSN: 0886-1544
    Keywords: actin ; calcium ; coelomocytes ; ionophore ; pH ; shape transformation ; video microscopy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have investigated the ability of the Ca+ + ionophore A23187 to induce the transformation of petaloid sea urchin coleomocytes to the filopodial form. The response of individual cells to different media was observed with time-lapse phasecontrast video microscopy. In the presence of 1 mM CaCl2, isotonic medium containing 1-5 μM A23187 produces a similar shape transformation to that caused by hypotonic shock. Higher concentrations of ionophore (10-20 μM) induce the formation of filopodia that are thinner and less rigid than those generated by hypotonic shock or low doses of ionophore. A23187 also induces shape transformation in highly flattened cells that do not respond fully to hypotonic shock. The induction of cytoplasmic alkalinization by NH4Cl, methylamine-HCl, or the Na+ ionophore monensin does not induce shape transformation, suggesting that increased intracellular pH is not the stimulus for this process. Ultrastructural changes in cytoskeletal organization were examined in negatively stained detergent-extracted cells. Low doses of ionophore produce filopodia that are indistin-guishable from those of hypotonically shocked cells, with actin filament bundles that are straight and cohesive along their entire length. High concentrations of ionophore produce filopodia with filament bundles that branch repeatedly and splay apart near their tips, forming loops and irregular curves. These results suggest that an increase in intracellular free Ca+ + concentration acts as the trigger that stimulates coelomocyte shape transformation, but that abnormally high concentrations of intracellular Ca+ +, produced by high doses of ionophore, interfere with actin filament bundling.
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    Cell Motility and the Cytoskeleton 4 (1984), S. 121-128 
    ISSN: 0886-1544
    Keywords: axonal transport ; ATP ; nucleotides ; saltatory movement ; dynein ; video microscopy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: In a permeabilized axon model, exogenous ATP can reactivate intraaxonal saltatory organelle movements (microscopically visible manifestations of fast axonal transport). We have studied the dependence of the reactivated movements on the ATP concentration and have also examined the nucleotide specificity of the reactivation. Organelle transport was visualized in isolated lobster giant motor axons using Nomarski optics and video microscopy. The axons were permeabilized with saponin, and movement was reactivated with ATP or other nucleotides. Some slight movement was seen with ATP concentrations as low as 10 μM. The velocity and frequency of the reactivated transport increased with increasing ATP concentrations up to about 5 mM. Movement was also reactivated by deoxyadenosine triphosphate, but not by AMP-PNP (a nonhydrolyzable ATP analogue), ADP, or AMP. Although other nucleotides (CTP, GTP, UTP, ITP) could reactivate transport, movement equivalent to that produced by 0.1 mM ATP was only seen with tenfold or greater concentrations of the other nucleotides. This pattern of specificity is consistent with the hypothesis that a dynein-like ATPase, rather than a myosin, is involved in fast axonal transport.
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    Cell Motility and the Cytoskeleton 4 (1984), S. 137-149 
    ISSN: 0886-1544
    Keywords: anti-fluorescein ; fluorescent analog cytochemistry ; molecular cytochemistry ; microinjection ; actin ; acetamidofluorescein-actin ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Fluorescent analogs of cellular components are finding increasing use in the field of cell biology. The power of this technique can be augmented by the use of antibodies specific for the fluorophore to visualize selectively the fluorescent analog at the electron microscope level. Rabbit antibodies specific for fluorescein were elicited and purified according to published methods (Lopatin and Voss [1971]: Biochemistry 10:208). Immune sera and IgG formed precipitin lines with fluorescein-labeled proteins in Ouchterlony immunodiffusion assays, and significantly quenched the fluorescence of fluorescein-labeled proteins. Immune IgG and Fab fragments decorated fluorescein-labeled actin, but not unlabeled actin, in negative-stained preparations. Anti-fluorescein IgG was used for immunofluorescent localization of fluorescein-labeled actin following microinjection of the fluorescent analog into living cells. This approach was extended to the immunoelectron microscopic localization of the injected analog at the subcellular level by the use of an electron-dense marker coupled to goat anti-rabbit IgG. Many other fluorescent probes also can be used as haptens for production of antibodies. Therefore, a general method for localizing fluorescently labeled molecules at the electron microscopic level is now available. Several other applications of anti-fluorescein antibody in studies involving fluorescent analogs are also suggested.
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    Cell Motility and the Cytoskeleton 4 (1984), S. 215-226 
    ISSN: 0886-1544
    Keywords: sperm motility ; flagellum ; axoneme ; microscopy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Iontophoretic application of ATP to the flagellum of the demembranated hamster spermatozoon produced a planar pair of bends at the two ends of the stimulated site. During bend propagation, torsion appeared in the vicinity of the interbend in some responses such that the distal bend was twisted clockwise when viewed from the base of the flagellum. This pattern of propagation is consistent with the instantaneous configurations of free-swimming cells previously described. The technique used here establishes that the three dimensionality arises from propagation per se, and does not depend on forces developed during swimming. The rolling of both free-swimming intact and demembranated spermatozoa was examined by two-color darkground videomicroscopy and the direction of rotation was, as predicted, always anticlockwise. A hypothetical mechanism, involving differential speeds of propagation of active sliding within the active microtubule subset, is proposed to account for the observed waveforms.
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    Cell Motility and the Cytoskeleton 4 (1984) 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 115-130 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The relationship between the cytoskeleton, stress fiber formation, and cell shape has been difficult to determine in fibroblasts grown in tissue culture. Vagaries in cell shape are complicated, as well, by stochastic cell movements. We dictated the attachment sites and shape of fibroblasts by growing them on square adhesive substrates surrounded by nonadhesive substrates. Cytoskeletal models were made by treating the cells with buffered Triton X-100 and glycerol. The residues were then examined by scanning electron microscopy followed by light microscopy of the same cells. The cytoskeletons of randomly moving cells were examined with whole mount transmission microscopy to confirm images seen with scanning microscopy. The cells thus examined demonstrated definite relationships between ruffling activity and stress fiber terminations, which were limited to the more adhesive, palladium substrate. No stress fibers were seen to end on the lesser adhesive substrate, agarose, and ruffling did not occur across the agarose. Cells too small to fill an entire square tended to extend across one diagonal of the square, and the stress fibers ran parallel to the longest axis of these cells. Larger cells were able to completely fill their squares. The cytoskeletons of these cells were organized in a spatial relation to the square shape of the cells. The cortical meshwork was aligned circularly and diagonally within the cells. Stress fibers appeared to form from the microfilaments of the meshwork and were aligned diagonally across the cells. We conclude that the diagonal arrangement of the stress fibers and cortical meshwork is caused by the same mechanism by which smaller cells spread over the longest axis of a square. Regions of cells where the meshwork was absent or where stress fibers were tightly bundled were occupied by more randomly arranged cytoskeletal components. Regions of tighyly bundled stress fibers did not seem to coincide with regions of cortical meshwork as seen by either whole mount transmission or scanning electron microscopy. Stress fibers were revealed in the light microscope to course beneath more randomly oriented cytoskeletal elements. These “lacework-like” elements were found frequently in square cells. Conspicuous structures in this random lacework were focal points of radially arranged filaments. Our observations suggest a continuity between stress fibers and the cortical microfilaments. The orientation of fibers and filaments was, in turn, dependent on cell shape for organization within the cell.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 163-171 
    ISSN: 0886-1544
    Keywords: concanavalin A ; platelet ; lamella ; pseudopod ; lectin receptor ; cytochalasins ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Ligand-induced redistribution of concanavalin A (Con A) receptors on the surfaces of substrate-spread rabbit platelets was studied. These receptors were diffusely distributed on the surface of prefixed cells. Incubation of living cells with Con A and anti-Con A antiserum led to redistribution of corresponding receptors: These receptors were patched, removed from the upper surface of substrate-attached pseudopodia and lamellipodia, and accumulated on the surface of central cell parts. Capping of receptors was inhibited by cytochalasins B and D and by cold. Thus, capping of surface receptors can take place not only in large nucleated cells but also in platelets, that is, in small anucleate cell fragments. It is suggested that the cells of various types in the course of their attachment to the substratum form pseudopodia and lamellipodia that have a number of common characteristics, including the ability to move the cross-linked surface receptors centripetally.
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    Cell Motility and the Cytoskeleton 4 (1984), S. 351-370 
    ISSN: 0886-1544
    Keywords: axon ; rate ; nervous system ; tissue culture ; cell growth ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A new formula calculates rates of directed axonal growth (elongation or retraction) using measurements of growth cone movements. By explicitly separating changes in axonal length from other nonelongational growth cone movements, the calculated rates reflect the detailed cellular growth mechanisms more directly than previous growth measures. In addition, the formula produces three distinct parameters of axonal elongation: n, a growth step rate; s, a growth step size; and P, a probability that a growth step leads to axonal elongation. For normal and regenerating individual chick and frog axons in culture, the formula has quantitated the following differences: the axon itself can elongate more rapidly in the chick, and the axon elongates in smaller steps in the chick. The underlying dynamics of growth of regenerating axons are quite similar to normal axons, but, in the short term, regenerating axons elongate in larger steps and at a slower rate. The distribution of these new rate measurements suggests that the elongation of axons can be usefully modelled as a one-dimensional stochastic walk.
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    Cell Motility and the Cytoskeleton 4 (1984), S. 371-385 
    ISSN: 0886-1544
    Keywords: microtubules ; dynein ; tubulin ; cilia and flagella ; microtubule associated proteins ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Dynein, obtained from axonemes of Chlamydomonas, binds by both its A and B ends to microtubules assembled from twice cycled (2 ×) and purified (6S) brain tubulin as well as to microtubules in native spindles, thereby inducing microtubule crossbridging. The two ends of the dynein arm exhibit distinct binding characteristics for the different microtubule preparations. Greater than 99% of the dynein arms are bound exclusively by their B ends to microtubules assembled from 6S tubulin in the presence of dynein and decorated to saturation. In contrast, greater than 80% of the dynein arms are bound by both their A and B ends to and, therefore, crossbridge 6S microtubules that are only partially dynein decorated. Binding of the A end of the dynein arm to saturated 6S microtubules can be enhanced by destabilizing the binding of the B end upon addition of ATP and vanadate. These observations suggest that Chlamydomonas dynein arms can bind by their A ends to microtubules assembled from 6S tubulin only when the B ends of the arms either are not bound or are bound but do not occupy all available dynein binding sites. Dynein exhibits a slight preference for binding by its A end to microtubules assembled from 2 × tubulin and containing microtubule associated proteins (MAPs). Approximately 90% of the dynein arms crossbridge adjacent 2 × microtubles that are only partially decorated. But as saturation of these microtubules with dynein is approached, the majority of the arms are bound solely by their A ends, while a smaller percentage are bound by their B ends or by both their A and B ends. These studies indicate that the type of microtubule as well as the degree of saturation of the microtubule with dynein can determine whether microtubule crossbridging occurs.
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    Cell Motility and the Cytoskeleton 4 (1984), S. 405-416 
    ISSN: 0886-1544
    Keywords: cardiac muscle ; myofibril ; cell spreading ; Z bands ; alpha-actinin ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Cardiac myocytes were isolated from 5-6-day-old chick embryos and allowed to spread in culture. The distribution of alpha-actinin in the cells was followed for five days in culture by exposing permeabilized cells to rhodamine-labeled alpha-actinin and also by injecting the labeled alpha-actinin into living myocytes. In addition to labeling the Z bands of sarcomeres, the added alpha-actinin also labeled small particles that were usually arranged periodically in linear arrays with a spacing between particles of 0.3-2.0 μm. Actin was localized between the particles of alpha-actinin by means of fluorescein-labeled heavy meromyosin. The punctate localization of alpha-actinin was prominent in pseudopods, behind ruffles, and at the periphery of spreading cells. Long rows of particles of alpha-actinin were often parallel to one another with the alpha-actinin particles in register. These linear arrays appeared to merge laterally to form strands with broader concentrations of alpha-actinin. Other linear arrays were parallel to myofibrils in the cell and some extended outward from the ends of myofibrils. We conclude that during spreading of cardiac myocytes, myofibrils form at the cell periphery behind the extending margins of the cell, and that the aggregates of alpha-actinin found in these areas are nascent Z bands in the forming myofibrils.
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    Cell Motility and the Cytoskeleton 4 (1984), S. 469-503 
    ISSN: 0886-1544
    Keywords: cytogel ; actomyosin ; Physarum ; oscillations ; mechanics ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The contractility of actomyosin gels is the basis for a variety of cellular motility phenomena. We present here a mechanical analysis of contractile gels. By making certain hypotheses on the chemical regulation of cytogel contraction we formulate a model for the rhythmic contractions of plasmodia in the slime mold Physarum polycephalum which is in accord with a number of experimental observations.
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    Cell Motility and the Cytoskeleton 2 (1982), S. 127-132 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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    Cell Motility and the Cytoskeleton 2 (1982), S. 153-158 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 2 (1982), S. 143-147 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
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    Cell Motility and the Cytoskeleton 4 (1984) 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 4 (1984), S. 7-23 
    ISSN: 0886-1544
    Keywords: axoplasm ; elastic modulus ; viscosity ; motility ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A magnetic sphere viscoelastometer has been developed to peform rheological experiments in living axoplasm of Loligo pealei. The technique includes the use of a calibrated magnetic sphere viscoelastometer on surgically implanted ferro-magnetic spheres in intact squid giant axons. The axoplasm was discerned to be “living” by the biological criterion of tubulovesicular organelle motility, which was observed before and after experimentation. From these in vivo experiments, new structural characteristics of the axoplasm have been identified. First, analysis of magnetic sphere trajectories has shown the axoplasm to be a complex viscoelastic fluid. Directional experimentation showed that this material is structurally anisotropic, with a greater elastic modulus in the direction parallel to the axon long axis. Second, both magnetic sphere and in vivo capillary experiments suggested that the axoplasm is tenaciously anchored to the axolemma. Third, it was found that axoplasm could be modelled as a linear viscoelastic material in the low shear rate range of 0.0001 to 0.004 s-1. The simplest mechanical model incorporating the discovered properties of the material in this range is Burger's model.
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