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  • Life and Medical Sciences  (3,596)
  • Organic Chemistry  (2,006)
  • AERODYNAMICS  (1,667)
  • 1985-1989  (7,269)
  • 1980-1984
  • 1988  (2,656)
  • 1986  (2,329)
  • 1985  (2,284)
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  • 1985-1989  (7,269)
  • 1980-1984
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  • 1
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 5 (1985), S. 251-263 
    ISSN: 0886-1544
    Keywords: Chlamydomonas ; movement ; flagellar beat ; stigma ; high-speed microcinematography ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Using high-speed microcinematography (100-500 f/s) the movement of Chlamydomonas reinhardtii mutant 622 E has been studied with frame-by-frame analysis. The stigma lies in the cell equator, displaced out of the flagellar plane anticlockwise by an angle of about 45°. During forward movement the cells rotate anticlockwise about their long axis with a frequency of 1.4-2 Hz (maximum 2.5 Hz). The rotation is caused by a lateral component of 3-dimensional beating of the flagella during the effective strokes. The helical swimming path is a result of an unequal flagellar beating. This is normally synchronous, but synchrony is interrupted from time to time by an additional beat of the outward directed flagellum, in our study one after about every twenty beats on average. These results are discussed and compared with the results published by other groups.
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  • 2
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 5 (1985) 
    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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  • 3
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 5 (1985), S. 377-392 
    ISSN: 0886-1544
    Keywords: newt ; lung ; cilia ; beat frequency ; waveform ; models ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Highly coupled newt lung ciliated cell models were used to study the effects of MgATP concentration on ciliary beat frequency and waveform. Models were prepared from ciliated lung cells of the newt Taricha granulosa by trypsin dissociation of the epithelium, demembranation with Triton X-100, and reactivation with MgATP, as described previously [Weaver and Hard, 1985]. Beat frequencies were measured stroboscopically. Ciliary waveforms of reactivated models and intact mucociliary epithelial sheets were determined by single frame analysis of high-speed movies. Waveform parameters calculated included the durations of the effective and recovery strokes, the angular velocities of the ciliary base and tip, the position of the bend along the ciliary shaft during the recovery stroke, the velocity of recovery stroke bend propagation, and the ratio of the duration of recovery stroke bend propagation to the duration of the recovery stroke itself. We found that beat frequency varied biphasically in response to MgATP at 21°C, as shown previously for isolated, individual, newt lung axonemes. Apparent Fmax (maximum beat frequency) and Km values of 25 Hz and 0.14 mM, and 35 Hz and 0.47 mM, respectively, were obtained for each linear segment of the biphasic double reciprocal plot. Demembranation did not alter either ciliary waveform or the pattern of coordination. In this system, metachrony is antilaeoplectic and ciliary waveform appears to be regulated independent of beat frequency.
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  • 4
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 5 (1985), S. 507-527 
    ISSN: 0886-1544
    Keywords: axonal transport ; microtubules ; organelles ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A model for fast axonal transport is developed in which the essential features are that organelles may interact with mechanochemical cross-bridges that in turn interact with microtubules, forming an organelle-engine-microtubule complex which is transported along the microtubules. Computer analysis of the equations derived to describe such a system show that most of the experimental observations on fast axonal transport can be simulated by the model, indicating that the model is useful for the interpretation and design of experiments aimed at clarifying the mechanism of fast axonal transport.
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  • 5
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 6 (1986), S. 1-1 
    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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  • 6
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 6 (1986), S. 25-34 
    ISSN: 0886-1544
    Keywords: respiratory cilia ; dynein ; ATPases ; porcine trachea ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Milligram amounts of mammalian ciliary axonemes were isolated from porcine tracheas. These were reactivated upon addition of ATP, indicating intact functional capability with a mean beat frequency at 37°C of 8.2 Hz. Electron microscopy showed typical ultrastructure of the isolated demembranated axonemes. Electrophoresis into polyacrylamide gradient gels containing sodium dodecyl sulfate revealed reproducible protein profiles from ten different tracheal preparations. Four major protein bands were observed in the 300-330 K molecular weight region, as well as tubulin at 51-54K. Extraction of the isolated tracheal axonemes with 0.6M KCl removed the outer dynein arms seen in electron microscopic cross-section of axonemes, preferentially solubilized two of the high molecular weight proteins at 320 and 330 K, and resulted in a three- to four-fold increase in ATPase specific activity. Sedimentation of the dialyzed salt extract on a 5-30% sucrose density gradient and subsequent fractionation yielded two peaks of ATPase activity. The faster migrating, 19S major ATPase peak correlated with the 320 and 330 K proteins, and two other proteins at 81 and 67 K. The slower sedimenting, 12S minor ATPase peak corresponded to a 308 K protein and two smaller proteins at 33 and 48 K. Thus, the outer dynein arm of tracheal cilia appeared to be associated with at least two high molecular weight proteins. These results demonstrate that adequate quantities of functionally intact axonemes can be reproducibly isolated from porcine tracheas, allowing further fractionation and analysis of mammalian cilia.
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  • 7
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 6 (1986) 
    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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  • 8
    ISSN: 0886-1544
    Keywords: contractile system ; microfilaments ; microtubules ; endoplasmic reticulum ; ciliophora ; oligotrichina ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Tontonia appendiculariformis is a marine planktonic ciliate with a long tail. The tail can contract rapidly, becoming transformed into an oval mass one-twentieth of its original length. The highly complex ulrastructure of the tail is described here in detail. A large part of the volume of the tail contains numerous more or less parallel membranous tubes. The membrane of the tubes has numerous invaginations and is probably derived from the smooth endoplasmic reticulum. This tubular material forms a continuous layer around the tail, interrupted in only one region, which contains cilia. Associated with the cilia are basal fibres with a periodically banded appearance. The tubular layer forms several folds separated by hyaloplasm containing many mitochondria. The pellicle of the tail is thrown into numerous pleats. It comprises a perilemma, a plasmalemma, and complex alveoli, but epiplasm and microtubules are absent. The alveoli appear to form septa within the folds of the layer of membranous tubes. In the region where the tail is attached to the body of the ciliate there are conspicuous bundles of microtubules and microfilaments. The membranous tubes and septa appear to be connected to small bundles of microfilaments, which presumably represent the contractile material. However, we consider the membranous tubes as potentially active in producing the change in shape. Although the structure of the tail of Tontonia is unique, there are certain similarities to the stalk of the Tintinnina and also to the motile extension of the dinoflagellate Erythropsidinium.
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  • 9
    Electronic Resource
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 6 (1986), S. 249-255 
    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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  • 10
    Electronic Resource
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 6 (1986), S. 229-236 
    ISSN: 0886-1544
    Keywords: α-helix ; filament motility ; filament contractility ; filament sliding ; microtubules ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The twisting behavior of α helices has hardly been considered hitherto with regard to the function of proteins. The well-known electrostatic repulsion between the highly charged side chains, which depends on their interaction with ions, is absolutely connected with torsional rotations of the helix as long as its hydrogen bonds hold. This means a direct transformation of chemical into mechanical energy. However, the stability of a twisted single α helix with charged side chains is low in an aqueous environment. It may easily ball up to form a globular molecule with nonhelical regions of the polypeptide chain. This corresponds to a primitive contraction that obviously occurs with spasminlike proteins that contain strongly twisted filaments as Salisbury [J. Submicrosc. Cytol. 15:105-110, 1983] has shown. Steps that increase the stability and rigidity of α helical filaments are (1) the formation of coiled-coils, (2) self-intertwining (“telephone cord phenomenon”) or intertwining with other coiled-coils as shown with the intermediate filaments, and (3) association with cytoskeletal elements (microfilaments, protofilaments of microtubules) that contain globular subunits. These coarser elements are rotated by winding and unwinding of the smaller helical molecules and thus transmit the torsion produced in the α helices to the microscopic level by the sliding (screwing) motion and the shearing effect that is connected with the waves of a rotating helix. Particles are transported if connected to the helical side arms. Since the displacement of the side arms seems to occur along the single protofilaments of a microtubule, a rotation of these protofiiaments is suggested. The bidirectional transport of particles along single microtubules may be explained by the association of left- and right-handed helices with the protofilaments. According to the models, parallel and antiparallel sliding of neurofilaments and neurotubules is suggested.
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  • 11
    Electronic Resource
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 6 (1986), S. 256-272 
    ISSN: 0886-1544
    Keywords: ciliary motility ; cAMP regulation ; swimming speed ; membrane potential ; detergent models ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The membrane potential of Paramecium controls the frequency and direction of the ciliary beat, thus determining the cell's swimming behavior. Stimuli that hyperpolarize the membrane potential increase the ciliary beat frequency and therefore increase forward swimming speed. We have observed that (1) drugs that elevate intracellular cyclic AMP increased swimming speed 2-3-fold, (2) hyperpolarizing the membrane potential by manipulation of extracellular cations (e.g., K+) induced both a transient increase in, and a higher sustained level of cyclic AMP compared to the control, and (3) the swimming speed of detergent-permeabilized cells in MgATP was stimulated 2-fold by the addition of cyclic AMP. Our results suggest that the membrane potential can regulate intracellular cAMP in Paramecium and that control of swimming speed by membrane potential may in part be mediated by cAMP.
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  • 12
    Electronic Resource
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 5 (1985), S. 529-543 
    ISSN: 0886-1544
    Keywords: actin ; regulatory protein ; cytoskeleton ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have isolated a 30,000-dalton protein from Dictyostelium which cosedimented with and affected the low shear viscosity of actin. At low concentrations, this protein increased the low shear viscosity to greater than that of the actin control, whereas higher concentrations decreased viscosity. The viscosity decrease correlated with the formation of actin filament bundles, as seen electron microscopically. This protein resembled a previously reported actin-binding protein from Dictyostelium [Fechheimer and Taylor, 84, J Biol Chem 259:4514] in electrophoretic mobility, Stokes radius, and ability to crosslink filaments, but was shown to be different by peptide mapping, lack of immunologic crossreactivity, and lack of sensitivity to calcium.
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  • 13
    Electronic Resource
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 5 (1985), S. 545-557 
    ISSN: 0886-1544
    Keywords: neutrophils ; cytoskeleton ; actin polymerization ; NBDphallacidin ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The studies presented here characterize a simple, quantitative NBDphallacidin extraction assay for determining the F-actin content of fMLP-activated neutrophils. The NBDphallacidin extraction assay is based upon the specificity of NBDphallacidin binding to F-actin and the solubility of NBDphallacidin in methanol. Cells are fixed, permeabilized, and stained with NBDphallacidin; the cells are then pelleted, the bound NBDphallacidin is extracted into methanol, and the RFI (excite 465; emit 535) of the solution is determined. Binding of NBDphallacidin to neutrophils is saturable and 90% of bound NBDphallacidin is displaced by nonfluorescent phalloidin. The extraction of bound NBDphallacidin into methanol is complete and the excitation/emission characteristics of NBDphallacidin are not altered by extraction. The assay is relatively inexpensive, applicable to the study of cells in suspension or on substratum, allows kinetic studies with 5-10s time resolution, and is not affected by the shape of the cell or the distribution of the probe. We used the NBDphallacidin extraction assay to study the kinetics of fMLP-induced change in the F-actin content of neutrophils and the effect of tBOC peptide, an inhibitor of fMLP binding, on these changes. The extraction assay reveals a rapid, sequential fMLP-induced increase followed by a decrease in F-actin content. The tBOC peptide inhibits fMLP-induced actin polymerization. Addition of tBOC during fMLP-induced polymerization or at times when F-actin content is maximal enhances F-actin depolymerization. The rate of F-actin depolymerization is ≥ fourfold faster in the presence than in the absence of tBOC. The results show that (1) The NBDphallacidin extraction assay is useful for studying the kinetics of change in F-actin content of nonmuscle cells; (2) fMLP receptor occupancy is required for fMLP-dependent polymerization but not depolymerization; and (3) both the actin polymerizing and depolymerizing processes are active in the cell within 5 s after fMLP stimulation. Implications of these observations for understanding the observed increase and, then, decrease in F-actin content of fMLP-activated cells are discussed.
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  • 14
    Electronic Resource
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 6 (1986), S. 48-55 
    ISSN: 0886-1544
    Keywords: vinculin ; actin ; adhesion plaques ; cytoskeleton ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Vinculin was purified from chicken gizzard by a modification of the method of Feramisco and Burridge [1980; J Biol Chem 255:1194]. Vinculin did not alter the viscosity of actin as measured in an Ostwald viscometer, nor did it affect actin polymerization as measured with the fluorescent NBD-actin assay. Sedimentation experiments demonstrated that vinculin did not bind to actin, and electron microscopy of negatively stained specimens indicated that vinculin did not aggregate actin filaments into bundles. These results suggest that vinculin, by itself, does not interact with actin at least under commonly used conditions to assay actin-protein interactions in vitro.
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  • 15
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 6 (1986), S. 35-47 
    ISSN: 0886-1544
    Keywords: microbeam ; microtubules ; nucleus ; cytoskeleton ; motility ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: During hyphal tip growth in the fungus Basidiobolus magnus, nuclei normally maintain a constant distance from the advancing cell apex by continuously migrating forward. It is not known whether the mechanism that produces nuclear movement also mediates nuclear positioning, or whether these two processes are under separate control. By irradiating small cytoplasmic regions with an ultraviolet microbeam, the coordination between movement and positioning could be disrupted. Regardless of the distance of the target from the nucleus, anterior irradiations (those ahead of the nucleus) caused the nucleus to stop or move backwards, whereas posterior (behind the nucleus) irradiations caused an acceleration in the nuclear velocity. The nucleus retained its ability to move following irradiation, so there was only loss of control over normal positioning. These results suggest that movement and positioning are mediated by different mechanisms. Quantitative microtubule analysis demonstrated that microtubules in the target region had been depolymerized, but in other regions of the cell they were apparently normal. We suggest that the depolymerization of microtubules affects nuclear movement by altering the tensile strength of the cytoplasm, and that cytoskeletal tension mediate nuclear positioning.We also found that accelerated nuclear movement could occur when most of the microtubules surrounding the nucleus were depolymerized. A comparison of the microtubule population surrounding the nucleus in unirradiated versus irradiated cells suggested that microtubules move with nuclei. Therefore, the nucleus does not appear to move via a direct interaction with microtubules.
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  • 16
    Electronic Resource
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    Cell Motility and the Cytoskeleton 6 (1986), S. 640-648 
    ISSN: 0886-1544
    Keywords: cell model ; lamellipodia ; phosphorylation ; Ca2+-dependent contraction ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Permeabilized cell models of muscle and nonmuscle cells have proven useful for examining the regulation of actin, myosin, and other cytoskeletal proteins during cell contraction. Upon addition of Ca2+ and ATP, glycerinated chick embryonic skin fibroblasts retract their tails and lamellipodia. Ca2+-independent contractions are obtained by preincubation of cell models in Ca2+ ATPγS, followed by EGTA and ATP addition, or by addition of trypsin-treated myosin light chain kinase that no longer requires Ca2+ for reactivation. By pretreating cells before glycerination with colchicine, it is possible to study lamellipodial contraction independent of tail contraction. Similar responses to ATPγS pretreatment and unregulated myosin light chain kinase are observed in cells that only contain lamellipodia. SDS-PAGE electrophoresis of glycerinated fibroblasts incubated in ATPγ35S and Ca2+ shows that only two major proteins are thiophosphorylated, and that one of them, a band that comigrates with the 20K MW light chain of myosin, is thiophosphorylated in a Ca2+-dependent manner. Since the rate of tail contraction is several-fold faster after Ca2+ and ATPγS pretreatment or incubation in excess myosin light chain kinase, myosin light chain phosphorylation may be a rate-limiting step during contraction.
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  • 17
    Electronic Resource
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    Cell Motility and the Cytoskeleton 6 (1986), S. 105-113 
    ISSN: 0886-1544
    Keywords: video-microscopy ; colloidal gold ; immunocytochemistry ; microtubules ; receptors ; saltatory motion ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We describe a new approach to probe the molecular biology of the living cell that uses small colloidal gold particles coupled to specific ligands. They are visualized in cells by bright-field, video enhanced contrast microscopy. We describe the basic aspects of the technique and provide examples of applications to intracellular motility, cell membrane dynamics, receptor translocation, internalization, and intracellular routing. We also provide examples of the use of this approach in immunospecific labelling of cells and tissue sections.
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  • 18
    Electronic Resource
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    New York, NY : Wiley-Blackwell
    Cell Motility and the Cytoskeleton 9 (1988), S. 337-348 
    ISSN: 0886-1544
    Keywords: isoactins ; immunogold ; myofibrils ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: In vertebrate skeletal muscle, the proliferating myoblasts synthesize nonmuscle isoforms of actin, and the cells begin to express muscle-specific actin isoforms during their myogenic differentiation. To study the distributions of the actin isoforms in myogenic cells and fully differentiated skeletal muscle, we prepared a peptide antibody specific for the skeletal α isoform of actin and used this antibody along with an antibody specifically reactive with nonmuscle γ actin to stain cultured myotubes and adult skeletal myofibrils by double-indirect immunofluorescence. At this level of resolution, no differences in isoform localization were seen: Both muscle and nonmuscle actins were detected in the myotubes and in the striations of mature myofibrils. Myotubes were also double-stained using immunogold electron microscopy, and the isoform distributions were determined quantitatively by counting the two sizes of gold particles that corresponded to labeling with each antibody. A quantitative analysis of immunoreactivity revealed that, although both forms were present in all actin-containing structures, nonmuscle actin was relatively more prevalent along the edges (cortical microfilaments) of the myotubes, whereas the muscle isoform predominated in the interior regions (containing forming myofibrils). Thus, we have found evidence of a heterogeneous distribution of muscle and nonmuscle actin isoforms in differentiating myogenic cells, and we have demonstrated that a nonmuscle actin isoform is a component of the muscle contractile apparatus.
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  • 19
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    Cell Motility and the Cytoskeleton 6 (1986), S. 114-121 
    ISSN: 0886-1544
    Keywords: mitosis ; microtubules ; tubulin incorporation ; assembly polarity ; Chaetopterus ; HeLa cells ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The incorporation of tubulin into mitotic spindles in situ was studied by incubating permeabilized mitotic cells in solutions containing [3H]GTP-labeled or dichlorotriazinylamino fluorescein (DTAF)-labeled tubulin. Metaphase HeLa cells or spindle-containing “minicells” from Chaetopterus oocytes were lysed in a microtubule-assembly buffer plus 0.5% Nonidet P-40, 1 mg/ml 120,000g supernatant mammalian brain tubulin, and [3H]GTP. After different periods of incubation, mitotic spindles were isolated in 2 M-glycerol-containing assembly buffer and separated from unbound counts by centrifugation through a 4 M-glycerol cushion; 3H counts per mg protein increase linearly for 8-12 min and then reach a plateau or steady state in both Chaetopterus oocytes and HeLa cells. Addition of 4 mM CaCl2 blocks or reverses incorporation. Little or no [3H]GTP is incorporated if exogenous tubulin or lysed cells are omitted from the assembly mixture.To measure the loss rate of [3H]GTP-tubulin from mitotic spindles, cells were incubated in tubulin plus [3H]GTP for 30 min, and a 20-fold excess of cold GTP (2 mM) was added. Samples were removed after incubation for different periods, and spindles were isolated as described above and counted for 3H content. [3H]GTP is lost from spindles at a rate of about 16%/min until a new steady state is reached in about 8 min. These results are consistent with an incorporation and turnover of [3H]GTP-tubulin in spindle microtubules of these lysed-cell models.The location of this newly incorporated tubulin in the spindle was investigated by incorporating fluorescent DTAF-tubulin into mitotic spindles of these lysed cell types. A short pulse (2-5 min) appears to label microtubules (MTs) near metaphase chromosomes and longer exposures label the entire spindle.The rates of incorporation and turnover that we see by [3H]GTP and fluorescent tubulin incorporation in situ are faster than those observed with brain MTs at steady state in vitro but are in the range of the rates of spindle fiber formation in prophase, and spindle MT reassembly after cooling.
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  • 20
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    Cell Motility and the Cytoskeleton 6 (1986), S. 128-135 
    ISSN: 0886-1544
    Keywords: motion analysis ; axonal transport ; cytoplasmic transport ; Brownian motion ; AVEC-DIC microscopy ; saltatory particle motion ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A survey study of organelle movements in a variety of cell types of plant and animal origin was made with the aid of video-enhanced contrast, differential interference contrast (AVEC-DIC) microscopy followed by fine analysis of the motile behavior of the individual organelles. We found that there exists besides Brownian motion a wide spectrum of active motions in cells, i.e. motion that is directionally biased through the expenditure of metabolic energy. The types of active motion seen range from a continuous motion (sometimes appearing as streaming) in plant cells and neurons to various types of less ordered and less well directed motion. We did not see any clear-cut qualitative difference between plant and animal cells or between systems presumed to be actin- and microtubule-based. A preliminary classification of the types of active motion is presented. The ongoing research activities, which aim at a more precise definition of the different types of motion by a set of quantitative parameters, are described, and the progress made so far is reported.
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  • 21
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    Cell Motility and the Cytoskeleton 6 (1986), S. 136-145 
    ISSN: 0886-1544
    Keywords: cytoplasmic movement ; microbeam ; Ca++ ; fungi ; saltatory movement ; cytoskeleton ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have investigated the mechanisms that hyphae of the fungus Basidiobolus magnus use to accomplish bulk movement of their cytoplasm and saltatory organelle movements. When cells were irradiated with an ultraviolet microbeam, cytoplasmic contraction occurred. The posterior cytoplasm (toward the septum) always moved forward toward the irradiated area, whereas anterior cytoplasm (between the cell tip and target) never contracted back toward the site of irradiation. Thus, there is an intrinsic polarity in the cytoplasm. Irradiations also arrested saltatory movements. The effects of irradiation on both saltations and cytoplasmic movement appear to be mediated by Ca++. Chelating exogenous Ca++ before irradiation eliminated contractions and prevented the inhibition of saltations. Furthermore, the effects of irradiation could be duplicated by using the Ca++ ionophore A23187. We relate the present results to our previous report on the effects of irradiation on the cytoskeleton [McKerracher and Heath, 1986]. We conclude that two separate cytoskeletal networks exist in fungal cells, and that an actin-containing network generates bulk cytoplasmic movement.
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  • 22
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    Cell Motility and the Cytoskeleton 6 (1986), S. 159-162 
    ISSN: 0886-1544
    Keywords: superprecipitation ; actomyosin ; F-actin bundle ; unidirectional sliding ; sliding velocity ; dark field microscopy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Unidirectional sliding of myosin filaments along F-actin bundles was produced with purified muscle actin and myosin in the presence of ATP. The velocity of myosin filament sliding was independent of myosin filament length. This result supports a recent hypothesis that long distance movement of myosin cross-bridge can be induced by splitting of one ATP molecule [Yanagida, Arata, and Oosawa, 1985. Nature. 316:366-369; Higashi-Fujime. 1985. J. Cell Biol., 101:2335-2344].
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  • 23
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    Cell Motility and the Cytoskeleton 6 (1986), S. 153-158 
    ISSN: 0886-1544
    Keywords: giant smooth muscle fiber ; ctenophore ; myofilaments ; ultrastructure ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Bundles of giant smooth muscle fibers of the ctenophore Beröe have been stretched up to four times their initial length and then examined by transmission electron microscopy. Stretching induces the segregation of actin and thick (myosinlike) filaments into separate domains. The thick filaments domains are made of 20-30 filaments, with a regular spacing identical to that of nonstretched fibers. A moderate stretching permits the visualization of microtendons associating actin filament bundles to hyaluronidase-resistant condensations of the extracellular matrix. It is deduced from these observations that Beröe giant smooth muscle fibers possess myofibrils which attach at both ends upon the sarcolemmal membrane. Each myofibril is probably made of two long actin filament bundles (of approximately 150 filaments) and short (2-3 μm) myosinlike bundles (of approximately 30 filaments).
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    Cell Motility and the Cytoskeleton 6 (1986), S. 163-175 
    ISSN: 0886-1544
    Keywords: centrosomes ; fertilization ; mice ; microfilaments ; microtubules ; mitosis ; pericentriolar material ; sea urchins ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Motility and the behavior and inheritance of centrosomes are investigated during mouse and sea urchin fertilization. Sperm incorporation in sea urchins requires microfilament activity in both sperm and eggs as tested with Latrunculin A, a novel inhibitor of microfilament assembly. In contrast the mouse spermhead is incorporated in the presence of microfilament inhibitors indicating an absence of microfilament activity at this stage. Pronuclear apposition is arrested by microfilament inhibitors in fertilized mouse oocytes. The migrations of the sperm and egg nuclei during sea urchin fertilization are dependent on microtubules organized into a radial monastral array, the sperm aster. Microtubule activity is also required during pronuclear apposition in the mouse egg, but they are organized by numerous egg cytoplasmic sites. By the use of an autoimmune antibody to centrosomal material, centrosomes are detected in sea urchin sperm but not in unfertilized eggs. The sea urchin centrosome expands and duplicates during first interphase and condenses to form the mitotic poles during division. Remarkably mouse sperm do not appear to have the centrosomal antigen and instead centrosomes are found in the unfertilized oocyte. These results indicate that both microfilaments and microtubules are required for the successful completion of fertilization in both sea urchins and mice, but at different stages. Furthermore they demonstrate that centrosomes are contributed by the sperm during sea urchin fertilization, but they might be maternally inherited in mammals.
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    Cell Motility and the Cytoskeleton 6 (1986), S. 186-192 
    ISSN: 0886-1544
    Keywords: Euglena ; pellicular strip ; cell shape ; sliding ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We recently showed by videomicroscopy that adjacent pellicular strips slide relative to each other without contraction during S-shaped bending movement in Euglena fusca (Suzaki and Williamson, 1985). In order to validate this sliding strip mechanism for other species and other shape changes, a theoretical analysis and a computer simulation were carried out. Some of the commonly observed euglenoid cell shapes (rounded. S-shaped, and embryo-like shapes) were generated. Our results suggest that Euglena probably achieves a variety of cell shape changes by means of locally regulated sliding between adjacent pellicular strips of constant length and width.
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    Cell Motility and the Cytoskeleton 10 (1988), S. 172-184 
    ISSN: 0886-1544
    Keywords: chromosome movement ; spindle elongation ; micromanipulation ; mechanical properties ; mitosis ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Mechanical properties of the mitotic spindle and the effects of various operations of the mitotic apparatus on the chromosome movement and spindle elongation were investigated in fertilized eggs and blastomeres of the sand dollar, Clypeaster japonicus. On the basis of results with mechanical stretching and compression of the spindle with a pair of microneedles and the behavior of an oil drop microinjected into the spindle, it was concluded that the equatorial region of the spindle is mechanically weaker than the half-spindle region. Anaphase chromosome movement occurred in the spindle from which an aster had been removed or separated with its polar end and in the spindle in which the interzonal region had been removed. This fact indicates that chromosomes move poleward in anaphase by forces generated near the kinetochores in the half-spindle. Because of the effects of separation or removal of an aster from the spindle on the spindle elongation in anaphase and the behavior of the aster, it was concluded that the spindle elongation in anaphase is caused by pulling forces generated by asters attached to the ends of the spindle.
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    Cell Motility and the Cytoskeleton 10 (1988), S. 153-163 
    ISSN: 0886-1544
    Keywords: intracellular particle motions ; cytoplasmic streaming ; onion (Allium) epidermal cells ; video microscopy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The endoplasmic reticulum (ER) and associated organelle and particle movements in onion (Allium cepa) bulb scale epidermal cells were observed, recorded, and analyzed using computer-assisted video (AVEC-DIC, AVEC-POL and fluorescence) microscopy. The ER is composed of two interconnected sets of filamentous membrane tubules with diameters ranging from 0.1 to 0.5 μm. The first form a more stable, stationary network of intersecting polygonal membrane tubules lying closely appressed to the plasma membrane and continuous with a second very dynamic set of longer membrane tubules that often are located parallel to each other, shifting rapidly around the cytoplasm and forming dynamic knots or organization centers. The ER, mitochondria, and spherosomes fluoresced upon chlortetracycline treatment and are therefore presumed to sequester calcium. ER and mitochrondria also stain with the fluorescent dye, rhodamine 123. Mitochrondria and spherosomes are seen to move in the cytoplasm only along paths parallel to the axis of the ER tubules. Smaller particles (0.5 μm) tend to follow these same paths but may occasionally move independently. Particles and organelles move in close, but not in direct, association with the ER tubules. In optically favored cells, actin filaments were occasionally recorded located in parallel with the ER tubules and directly associated with moving particles. Streaming ceased promptly and reversibly upon treatment with cytochalasin B, which did not visibly disrupt the ER. Short-term treatment with colchicine did not inhibit streaming or disrupt the ER network, whereas long-term (hours) colchicine treatments caused the disappearance of the stationary, cortical polygonal networks and an aggregation of still slowly moving organelles and particles onto now visible actin filaments. This suggests that microtubule breakdown disrupts the three-dimensional distribution of the ER and rearranges actin filaments in the cell's cytoplasm. Actin filaments must be directly involved in generation of movement of the particles and organelles. A three-dimensional model, based on optical sectioning of the epidermal cells, is proposed to illustrate the distribution of the endoplasmic reticulum in onion epidermal cell cytoplasm.
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    Cell Motility and the Cytoskeleton 10 (1988), S. 185-196 
    ISSN: 0886-1544
    Keywords: mitosis ; kinetochore ; video microscopy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We describe preliminary results from two studies exploring the dynamics of microtubule assembly and organization within chromosomal spindle fibers. In the first study, we microinjected fluorescently labeled tubulin into mitotic PtK1 cells and measured fluorescence redistribution after photobleaching (FRAP) to determine the assembly dynamics of the microtubules within the chromosomal fibers in metaphase cells depleted of nonkinetochore microtubules by cooling to 23-24°C. FRAP measurements showed that the tubulin throughout at least 72% of the microtubules within the chromosomal fibers exchanges with the cellular tubulin pool with a half-time of 77 sec. There was no observable poleward flux of subunits. If the assembly of the kinetochore microtubules is governed by dynamic instability, our results indicate that the half-life of microtubule attachment to the kinetochore is less than several min at 23-24°C.In the second study, we used high-resolution polarization microscopy to observe microtubule dynamics during mitosis in newt lung epithelial cells. We obtained evidence from 150-nm-thick optical sections that microtubules throughout the spindle laterally associate for several sec into “rods” composed of a few microtubules. These transient lateral associations between microtubules appeared to produce the clustering of nonkinetochore and kinetochore microtubules into the chromosomal fibers. Our results indicate that the chromosomal fiber is a dynamic structure, because microtubule assembly is transient, lateral interactions between microtubules are transient, and the attachment of the kinetochores to microtubules may also be transient.
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    Cell Motility and the Cytoskeleton 10 (1988), S. 237-245 
    ISSN: 0886-1544
    Keywords: video microscopy ; digital image processing ; fluorescence photobleaching ; microtubule dynamics ; living cell dynamics ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The ability to tag biological molecules fluorescently and to detect their distribution in living cells has promoted the study of cytoplasmic organization in general and microtubule dynamics in particular. The techniques that we have selected and developed allowed the determination of spatial and temporal changes of the microtubule network in living fibroblasts at the level of individual microtubules. We have employed two general approaches for determining pattern changes: direct video microscopy and photobleaching and subsequent observation. Direct observation of fluorescent microtubules by high-definition video microscopy provided good spatial resolution at several time points, but was limited to the less congested and thinner periphery of the cell. This approach was made possible by a relatively bright, photostable reporter, xrhodamine-tubulin, and showed that microtubules underwent rounds of assembly and disassembly from their ends. Bleaching and subsequent observation of lysed cells improved the signal to noise ratio by extracting soluble chromophore and permitted observations in congested areas, but was limited to a single time interval. This approach demonstrated that microtubule domains were replaced one by one and that turnover was most rapid at the cell periphery. Antibodies specific for nonbleached chromophore can be used to enhance the signal to noise ratio further or to extend spatial resolution by the use of immunoelectron microscopy. Direct video microscopy and photo-bleaching are two approaches to the study of dynamics that have complementary strengths and wide application to the biology of living cells.
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    ISSN: 0886-1544
    Keywords: axoplasmic transport ; motility ; microtubules ; MAPs ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Vesikin, a protein that can associate with squid axoplasmic vesicles or optic lobe microtubuies, has been implicated as a force-generating molecule involved in microtubule-dependent vesicle transport [Gilbert and Sloboda, 1986, 1988]. Because vesikin crossreacts with an antibody to porcine brain microtubule associated protein 2 (MAP 2), studies were conducted to compare squid vesikin and brain MAPs. When taxol stabilized microtubules containing vesikin as a microtubule associated protein were incubated in the presence of ATP, vesikin dissociated from the microtubule subunit lattice. This behavior would be expected for an ATP-dependent, force generating molecule that serves as a crossbridge between vesicles and microtubules. When chick brain microtubules were treated under the same conditions, MAP 2 remained bound to the microtubules while MAP 1 dissociated in a manner similar to vesikin. One dimensional peptide mapping procedures revealed that, although digestion of vesikin and MAP 2 generated several peptides common to both proteins, vesikin and MAP 2 are clearly not identical. Furthermore, the addition of vesikin or MAPS 1 and 2 to purified tubulin stimulated microtubule assembly in a manner dependent on the concentration of added protein. These findings demonstrate that brain MAPs share characteristics common to squid vesikin and support the suggestion that brain MAPs 1 and 2 might act as a force generating complex for vesicle transport in higher organisms.
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    Cell Motility and the Cytoskeleton 10 (1988), S. 255-262 
    ISSN: 0886-1544
    Keywords: regulation of organelle transport ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Allen Video-enhanced constrast/differential interference constrast (AVEC-DIC) microscopy was used in conjunction with video intensification immunofluorescence microscopy to demonstrate that organelles and vesicle (particles) can move in either direction along microtubular linear elements in fibroblasts [Hayden et al., 1983]. Since it is not possible to determine the number of microtubules making up a linear element with light microscopy alone, AVEC-DIC microscopy was used in conjunction with whole-mount electron microscopy to show bidirectional transport along a single microtubule [Hayden and Allen, 1984]. These studies demonstrate that the structural polarity of the microtubule does not determine the direction of particle motion, and since dynein is an asymetric molecule, a simple microtubule-dynein-particle hypothesis cannot explain bidirectional transport along a single microtubule.Very little is known about regulation of particle transport in most cell types. Human embryonic lung fibroblasts grown on glass coverslips were serum-deprived for 24 hours and re-fed with serumless medium; the particle translocations/5 minutes were then determined The cells were then re-fed with either serumless medium, serum-containing medium, or serumless medium containing some bioactive factor, and the particle translocations/5 minutes were again determined for the same cells. Medium containing 10% fetal bovine serum inhibited particle translocation by 51.8%. Of the bioactive factors tested, only vasopressin produced a significant reduction in particle translocations (38%). This suggests that protein kinase C or calcium/calmodulin kinase could be involved in regulating particle transport.
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    Cell Motility and the Cytoskeleton 10 (1988), S. 263-270 
    ISSN: 0886-1544
    Keywords: cilia ; axoneme ; ATP-induced microtubule sliding ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The dynein arms of ciliary doublet microtubules cause adjacent axonemal doublets to slide apart with fixed polarity. This suggests that there is a unique mechanochemistry to the dynein arm with unidirectional force generation in all active arms and also that not all arms are active at once during a ciliary beat. Negative stain and thin-section images of arms in axonemes treated with β, γ methylene adenosine triphosphate (AMP-PCP) show a consistent subunit construction where the globular head of the arm interacts with subfiber B of doublet N + 1. This interpretation differs from that provided by freeze etch and STEM interpretations of in situ arm construction and has implications for the mechanochemical cycle of the arm. A computer model of the arms in relation to other axonemal structures has been constructed to test these interpretations. Attachment of the head of the arm to subfiber B is directly demonstrable in splayed axonemes in AMP-PCP. About half of the doublets in an axoneme show such attachments, while half do not. This might imply that about half the doublets in an axoneme are active at any given instant and can be identified as such. This information may be useful in probing questions of how active arms differ biochemically from inactive arms and of how microtubule translocators in general become active.
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    Cell Motility and the Cytoskeleton 9 (1988), S. 191-204 
    ISSN: 0886-1544
    Keywords: turbidity ; ciliary doublet ; biphasic ; extrusion ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Several characteristics of the sliding disintegration of Tetrahymena ciliary axonemes were found by turbidimetric assay, the ATP-regenerating system, and quantitative determination of the ATP concentration. At ATP concentrations exceeding 40 μM, the response in terms of turbidity was biphasic and could be divided into three phases. The dependence of each phase on ATP concentration was examined. The time duration of phase 2 increased with ATP concentration. When the ATP concentration was kept constant by the ATP-regenerating system, consisting of pyruvate kinase and phosphoenol pyruvate, the time duration of phase 2 increased with the concentration of phosphoenol pyruvate. On examining the change in turbidity with decreasing ATP concentration, the transition from phase 2 to phase 3 was found to occur at an ATP concentration of 40 μM.Dark-field and electron microscopy indicated the sliding disintegration to be closely correlated with the degree of tubidity. At phase 1, one or two doublets extruded from most of the axonemes, and disintegration failed to progress during phase 2. At the transition point from phase 2 to 3, at about 40 μM, ATP, other doublets were noted to extrude from the axonemes one after the other, causing turbidity to be minimal by the end of phase 3.The ATP concentration dependence of stepwise sliding disintegration suggests that each axoneme may possess the ability to regulate doublet microtubule sliding at lower or higher concentrations of ATP. In response to local differences or gradients of ATP concentration along the axoneme, the axonemes may cause localized sliding of doublets, thus subsequently generating active bending movement.
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    Cell Motility and the Cytoskeleton 9 (1988), S. 205-218 
    ISSN: 0886-1544
    Keywords: F-actin ; actin bundling protein ; cytoimmunofluorescence ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The Ca+2-sensitive actin-binding protein isolated from Dictyostelium discoideum, 30,000-D protein (Fechheimer and Taylor: J. Biol. Chem. 259:4514-4520, 1984;) has recently been localized in filipodia of substrate-adhered amoebae (Fechheimer: J. Cell Biol. 104:1539-1551, 1987). We have determined that this protein has a Mr of 34,000 daltons and is strictly colocalized with actin filaments in both substrate-attached Dictyostelium amoebae and cultured fibroblasts. 3T3 fibroblasts, as well as normal and virally transformed rat kidney fibroblasts (NRK) contain a 34-kilodalton (kD) protein that cross-reacts specifically with antibody to the Dictyostelium bundling protein. Mammalian 34-kD protein is colocalized with F-actin in stress fibers and the cortical cytoskeleton in substratadhered fibroblasts. In substrate-adhered vegetative Dictyostelium, F-actin and 34-kD protein are concentrated in regions of the cell cortex exhibiting filipodia and membrane ridges. Multiple filipodia formed after exposure to the chemoattractant folic acid stain intensely for 34-kD protein, implying participation in the assembly of actin bundles during filipod formation. The cortex of pseudopodia also contained high concentrations of bundling protein, but pseudopod interiors did not. In contrast to vegetative Dictyostelium, F-actin and 34-kD protein were not colocalized in cells that had progressed through the development cycle. In fruiting bodies, 34-kD protein was detected by immunofluorescence microscopy only in prespore cells, while F-actin appeared in stalk cells and spores.
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    Cell Motility and the Cytoskeleton 9 (1988), S. 349-360 
    ISSN: 0886-1544
    Keywords: neurofilaments ; intermediate filaments ; neuronal cytoskeleton ; neurofilament heterogeneity ; neurofilament composition ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Triton X-100 insoluble neurofilament (NF) fractions were obtained from two parts of the stellate ganglion and the main giant axon. These were analyzed by one- and two-dimensional gradient polyacrylamide gel electrophoresis, cyclic assembly and disassembly, and electron microscopy. The NF fractions from the ganglion cell bodies (GCB) and from the part of the ganglion mainly consisting of axon initial segments (GIS) were of similar composition; neither contained detectable amounts of the 220 kda and high molecular weight ( 400 kda) NF subunits that were prominent in the axonal NF fraction. However, the GCB and GIS did contain large quantities of a set of 65 kda polypeptides that were minor constituents of the axonal NF fraction. The 65 kda-containing NF fraction from the ganglion could be cyclically disassembled and reassembled, but only under low salt conditions, in contrast to the high salt conditions used to cycle axonal NFs. A comparison of the peptide map of the 65 kda polypeptides with that of the 60 kda axonal NF subunit showed them to be different. These biochemical differences between the ganglionic and axonal NF fractions correlated with morphologic distinctions: ganglionic NFs were relatively smooth surfaced, whereas axonal NFs had long sidearms. Such observations support the hypothesis that the NF cytoskeleton of the neuron soma is different from that of the axon. Furthermore, the change from the somal form to the axonal form of NFs appears to occur in the region where the axon initial segment increases in diameter to become the axon proper.
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    Cell Motility and the Cytoskeleton 10 (1988), S. 363-373 
    ISSN: 0886-1544
    Keywords: nuclear rotation ; karyoplasmic streaming ; nucleus ; nucleolus ; 3-D motion ; time-lapse photography ; NGF ; GABA ; calcium ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Nuclear rotation (NR) is typically measured as motion of nucleoli within nuclei of cells in vitro. This occurs in cycling cells. However, its observation in neurons arrested in interphase indicates that mechanisms related to mitosis are not a prerequisite. We have recently shown that NR occurs in three dimensions within the nuclear space, that it occurs within the space delineated by the outer nuclear membrane and that it includes chromatin domains in addition to nucleoli and have postulated that this motion of chromatin domains is related to changes in gene expression. We now show that exposure of dorsal root, sensory neurons in vitro to nerve growth factor (NGF) or to γ-aminobutyric acid (GABA), agents which alter gene expression, and to agents causing redistribution of calcium, such as EGTA and the calcium ionophore A23187, significantly alters NR. The NGF increased the mean rate of NR and did so at a time after exposure when activity of RNA polymerases have been shown to rise. Exposure to GABA resulted, within minutes, in shifts of the nucleolus within the three-dimensional space of the nucleus, associated in some neurons with significant, sigmoidal increases in the rate of NR. The calcium ionophore A23187 as well as chelation of extracellular calcium with EGTA similarly increased rates. Importantly, excess calcium, with EGTA remaining present, returned NR of all nucleoli to rates not different from controls. This indicates that the increase in NR seen with EGTA is specific to the chelation of calcium and not a nonspecific response to EGTA. It is difficult to link the action of agents which alter gene expression or transmembrane ion balance with changes in NR. Nevertheless, in support of our hypothesis, the results presented here show that agents known to alter gene expression, alter NR in a temporally coincident manner and that they do so, possibly, by calcium-dependent mechanisms.
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    Cell Motility and the Cytoskeleton 10 (1988), S. 420-431 
    ISSN: 0886-1544
    Keywords: sperm motility ; hyperactivation ; vanadate ; nickel ; cadmium ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Free Ca2+ changes the curvature of epididymal rat sperm flagella in demembranated sperm models. The radius of curvature of the flagellar midpiece region was measured and found to be a continuous function of the free Ca2+ concentration. Below 10-7 M free Ca2+, the sperm flagella assumed a pronounced curvature in the same direction as the sperm head. The curvature reversed direction at 2.5 × 10-6 M Ca2+ to assume a tight, hook-like bend at concentrations of 10-5 to 10-4 M free Ca2+. Sodium vanadate at 2 × 10-6 M blocked flagellar motility, but did not inhibit the Ca2+-mediated change in curvature. Nickel ion at 0.2 mM and cadmium ion at 1 μM interfered with the transition and induced the low Ca2+ configuration of the flagellum. The forces that maintain the Ca2+-dependent curvature are locally produced, as dissection of the flagella into segments did not significantly alter the curvature of the excised portions. Irrespective of the induced pattern of curvature, the sperm exhibited coordinated, repetitive flagellar beating in the presence of ATP and cAMP. At 0.3 mM ATP the flagellar waves propagated along the principal piece while the level of free Ca2+ controlled the overall curvature. When Ca2+-treated sperm models with hooked midpieces were subjected to higher concentrations of ATP (1-5 mM), some cells exhibited a pattern of movement similar to hyperactivated motility in capacitated live sperm. This type of motility involved repetitive reversals of the Ca2+-induced bend in the midpiece, as well as waves propagated along the principal piece. The free Ca2+ available to the flagellum therefore appeared to modify both the pattern of motility and the flagellar curvature.
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    Cell Motility and the Cytoskeleton 10 (1988), S. 471-481 
    ISSN: 0886-1544
    Keywords: Dictyostelium ; limited proteolysis ; thick filaments ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Dictyostelium myosin was associated into dimers and small oligomers at very low ionic strength, filamentous at intermediate ionic strength, and monomeric in solution conditions of high ionic strength. These different associations were probed by fragmenting myosin with chymotrypsin, trypsin, or V-8 protease. All three proteases digested monomeric myosin giving rise to multiple fragments with a wide range of molecular weights. Filamentous myosin was not digested by the V-8 protease, was preferentially cleaved at a single site in the middle of the heavy chain by chymotrypsin, and was cleaved at several sites by trypsin. If the reaction was carried out in very low ionic strength, however, two of these proteases generated stable fragments of high molecular weight. Electron microscopic analysis of these stable fragments showed that tails were shorter than in intact myosin, indicating that the cleavage sites were in the rod portion of the molecule. Under the same conditions of enzymatic digestion, myosin that had been radio labeled in vivo with 32P was analyzed by SDS-PAGE and autoradiography. By comparing the state of phosphorylation and the size of the stable fragments, it was determined that the heavy chain phosphorylation site was located between 55 and 70 kD from the tip of the myosin tail, near a region where the tail displayed sharp bends.
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    Cell Motility and the Cytoskeleton 10 (1988), S. 464-470 
    ISSN: 0886-1544
    Keywords: ciliary beat ; cell coupling ; calcium dependency ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Abfrontal ciliated cells of Mytilus edulis gill beat when mechanically stimulated, a consequence of a Ca++-based generator potential and regenerative response. In contrast, the lateral ciliated epithelial cells arrest when stimulated, a consequence of a Ca++-based generator potential and a Na+/Ca++-based regenerative response. Iontophoretic injection of EGTA in abfrontal cells, followed by mechanical stimulation, results in a large, prolonged depolarization that returns to the resting level stepwise. It has been hypothesized that this phenomenon is caused by successive Ca++-dependent repolarizations in coupled cells, first in adjacent cells and then in the injected cell, in accord with relative EGTA loading. We have now demonstrated this same stepwise repolarization phenomenon in the Na+/Ca++-dependent lateral ciliated cells. In this case, each repolarization step is often preceded by a small spike. With either cell type, using two-electrode recording techniques, we can detect the stepwise repolarization in distant cells, proportionately decremented when the second (KCl) electrode is some distance from the injection (EGTA) electrode and stimulus. When force is applied between the electrodes and nearest the KCl electrode, a greater initial response is recorded from this electrode, presumably resulting from depolarization of its impaled cell, prolonged by EGTA diffusion through the intervening cell junctions. The subsequent repolarization steps are of approximately the same size, suggesting repolarization of cells between the two electrodes. These observations are consistent with the cell coupling/EGTA loading hypothesis and indicate that both cell types mediate repolarization through Ca++ and propagate ciliary beat or arrest through intracellular coupling.
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    Cell Motility and the Cytoskeleton 11 (1988), S. 235-247 
    ISSN: 0886-1544
    Keywords: video-enhanced contrast microscopy ; transverse fibers ; transport ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Wound healing in Swiss 3T3 cultures was investigated with video-enhanced contrast (VEC) microscopy. The formation of protrusions at the leading edge of cells along wounds was investigated in detail during the spreading stage, which usually lasted from 1 to 4 hr postwounding. Lamellipodia exhibited a continuous rearward, or centripetal, transport of a variety of cellular constituents at rates of ∼0.26 μm/sec from the leading edge. The lamellipodia were also the sites of lateral migration as well as extension and retraction of actin microspikes. Actin fibers oriented transversely to the direction of movement were also observed to transport centripetally at similar rates. These fibers may in part give rise to large actin fibers forming at the interface between the base of the lamellipodia and the lamellae. Beads 0.5 μm in diameter attached to the dorsal surfaces of lamellipodia also transported centripetally at rates of ∼0.21 μm/sec. Thus there is an apparent correlation between transport of a variety of structures within lamellipodia and with surface movements of lamellipodia.
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  • 41
    ISSN: 0886-1544
    Keywords: actin ; rhodamine-phalloidin ; cell shape and movement ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The amoeboflagellate transformation in Physarum polycephalum involves a series of dramatic changes in cell shape and motile behavior. This report describes the morphological and behavioral changes through which a synchronously transforming population of cells passes, stressing that, although there are a series of distinguishable stages, cells at all stages display striking plasticity. Our previous studies showed that amoeboflagellates transiently display a flattened motile extension - the ridge - that projects from a specific location on the cell surface and contains a laminar core densely packed with a series of crisscrossing arrays of actin microfilaments. Details are presented here concerning the movements of the ridge as well as the dynamics of ridge formation and disassembly in relation to other morphogenetic events of the transformation. The ridge forms at about the same time as transforming cells begin to elongate, propagates undulations parallel to the long axis of the cell as the transformation proceeds, and disassembles late in the transformation. Staining of fixed cells with the fluorescent probe rhodaminephalloidin shows that the actin of amoeboid cells is strikingly redistributed as the transformation proceeds. Amoeboflagellates contain most of the stainable actin in the ridge and in a ventral-posterior spot that may be a site of cell-substratum adhesion. These results provide additional insights into the possible functions of the ridge and the roles of actin during the amoeboflagellate transformation.
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  • 42
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    Cell Motility and the Cytoskeleton 11 (1988), S. 260-274 
    ISSN: 0886-1544
    Keywords: cytoskeleton ; atheromatosis ; wound healing ; fibromatosis ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The distribution of smooth muscle (SM) and non muscle myosins was compared with that of α-SM actin in various normal and pathological tissues and in cultured cells by means of indirect immunofluorescence using a monoclonal antibody specific for α-SM actin [anti-αsm-1, Skalli et al., 1986b] and two polyclonal antibodies raised against bovine aortic myosin (ABAM) and human platelet myosin (AHPM), respectively.In normal tissues ABAM stained vascular and parenchymal smooth muscle cells (SMC), myoepithelial cells and myoid cells of the testis in a pattern similar to that reported by other authors with antisera raised against non vascular SM myosin. Cells stained with ABAM were always positive for anti-αsm-1. In human and experimental atheromatous plaques, most cells were positive for AHPM; a variable proportion was also stained for ABAM plus anti-αsm-1. Myofibroblasts from rat granulation tissue, Dupuytren's nodule and stroma from breast carcinoma were constantly positive for AHPM and negative for ABAM; however, myofibroblasts from Dupuytren's nodule and breast carcinoma were anti-αsm-1 positive. Early primary cultures of rat aortic SMC were positive for ABAM and anti-αsm-1 and became negative for ABAM and positive for AHPM after a few days in culture. They remained positive for AHPM and anti-αsm-1 after passages; the staining of AHPM and anti-αsm-1 appeared to be colocalized along the same stress fibers.These results may be relevant for the understanding of SMC function and adaptation, and show that in non malignant SMC proliferation, α-SM actin represents a more general marker of SM origin than SM myosin.
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  • 43
    ISSN: 0886-1544
    Keywords: griseofulvin ; microtubule organizing center ; cell division ; β-mercaptoethanol ; pronuclei ; cytochalasin ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Centrosomes undergo cell cycle-dependent changes in shape and separations, changes that govern the organization of the cytoskeleton. The cytoskeleton is largely organized by the centrosome; however, this investigation explores the importance of cytoskeletal elements in directing centrosome shape. Since the sea urchin egg during fertilization and mitosis displays dramatic and synchronous changes in centrosome shape, the effects of cytoskeletal inhibitors on centrosome compaction, expansion, and separation were explored by the use of anticentrosome immunofluorescence microscopy. Centrosome expansion and separation was studied during two phases: the transition after sperm incorporation, when the compact sperm centrosome enlarges and the sperm aster develops, and from prometaphase to telophase, when the compact spindle poles enlarge. Compaction was investigated when the dispersed centrosome at interphase condenses into the two spindle poles at prometaphase. Although centrosome expansion and separation typically occur concurrently, β-mercaptoethanol results in centrosome separation independent of expansion. Microtubule inhibitors prevent centrosome expansion and separation, and expanded centrosomes collapse. Since pronuclear union is arrested by microtubule inhibitors, this treatment also affords the opportunity to explore the relative attractiveness of the male and female pronuclei for these centrosomal antigens. Both pronuclei acquire centrosomal material; though only the male centrosome is capable of organizing a functional bipolar mitotic apparatus at first division, the female centrosome nucleates a monaster. Microfilament inhibition (cytochalasin D) prevents centrosome separation but not expansion or compaction. These results demonstrate that as the centrosome shapes the cytoskeleton, the cytoskeleton alters centrosome shape.
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  • 44
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    Cell Motility and the Cytoskeleton 11 (1988), S. 275-290 
    ISSN: 0886-1544
    Keywords: ciliary motility ; inclination ; polarity of beating ; sliding velocity ; sliding translocation rate ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Motor responses of the frontal cirri of the ciliate Stylonychia were recorded at the axial view of the ciliary base with high-speed cinematography. Voltage-clamp applying sustained hyperpolarizing voltage steps was used to explore the properties of the ciliary cycle modulated by the membrane potential. Upon hyperpolarization between - 1 and - 13 mV, a previously inactive frontal cirrus reoriented from a neutral posture and started beating so that the axis of the beating cone of a proximal cirral segment assumed an orientation near 100° (proceeding counterclockwise from posterior = 0°) and inclination near 60° (0° = perpendicular to the cell surface). The major beating amplitude was limited to about 150°. Increasing hyperpolarization increased the spatial polarity of the cycle (ratio of major over minor amplitude, from 2 to 2.4). Rates of the power stroke increased with hyperpolarizations up to - 4 mV but were consistently smaller than those of the return stroke during the ciliary cycle (ratio: 0.4 to 0.6; = temporal polarity). Comparison of different hypothetical beat forms (0-shape, D-shape, and egg-shape) showed that the orientation-time data are the major determinants of the angular velocity and rate of reorientation of the cilium during the cycle. Geometric transformation of these data led to descriptions of the cycle of a proximal ciliary segment in terms of active sliding velocities and rates of unidirectional sliding translocation between identified doublets. Three voltage-sensitive functional parameters of the cilium - the inclination (which is noncyclic) and the rates of active sliding and sliding translocation (both of which are cyclic in nature) - are discussed as generating the spatial and temporal properties of the ciliary beat.
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  • 45
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    Cell Motility and the Cytoskeleton 11 (1988), S. 291-302 
    ISSN: 0886-1544
    Keywords: mitosis ; mitotic apparatus ; microtubules ; kinetochores ; metabolic inhibitors ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Hyperosmotic sucrose treatment of metaphase PtK-1 cells has been shown to produce a reversible concentration-dependent effect on spindle elongation linked to a functional alteration in the connection of the chromosome to the spindle (Pover et al.: European Journal of Cell Biology 39:366-372, 1985). Spindle elongation, similar to that which occurs at anaphase B, is thought to be driven by the compression stored in the form of microtubule curvature in the nonkinetochore (nkMT) population of microtubules at metaphase (Snyder et al.: European Journal of Cell Biology 35:62-69, 1984 and 39:373-379, 1985). Addition of metabolic inhibitors to Ham's F-12 salts with deoxyglucose (D/F-12 medium) containing 0.4 M sucrose and 1 mM DNP does not within statistical error affect the rate and extent of sucrose-induced spindle elongation; rates and extents are 60-75% of normal anaphase B motions. Electron microscopic analysis of metaphase cells treated with D/F-12 medium and 0.4 M sucrose with 1 mM DNP demonstrates that spindle microtubules lose curvature and become straight in appearance, typical of microtubule organization in untreated anaphase cells. Sucrose-treated cells released into D/F-12 medium show a rapid reduction in spindle length; however, cells treated with either 0.4 M sucrose or 0.4 M sucrose and 1 mM DNP-containing D/F-12 medium and released into DNP-containing D/F-12 medium do not exhibit a significant reduction in spindle length. Electron microscopic analysis links changes in spindle length with microtubule/kinetochore associations. These data suggest that energy required for the initial phases of spindle elongation during anaphase is preloaded into the mitotic spindle by metaphase and does not require additional energy to be expressed as examined by sucrose-induced spindle elongation in the presence of metabolic inhibitors. Second, energy is required to make or maintain (or both) functional chromosome associations with the spindle as measured by reduction in spindle length following sucrose removal.
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  • 46
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    Journal of Physical Organic Chemistry 1 (1988), S. 115-117 
    ISSN: 0894-3230
    Keywords: Organic Chemistry ; Physical Chemistry
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology , Physics
    Notes: A novel oxygen induced reduction of α,β-unsaturated carbonyl compounds is discovered. The reduction of the carbon-carbon double bond of α,β-unsaturated carbonyl compounds by benzeneselenol was caused by an introduction of molecular oxygen into the reaction system. This reduction is likely to proceed via a radical chain pathway involving an SH2 type reaction between a phenylseleno radical and a 1,2-adduct of benzeneselenol to the carbonyl group of the α,β-unsaturated carbonyl compound to give an allylic radical which absracts a hydrogen atom from benzeneselenol to form the reduction product.
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  • 47
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    Journal of Physical Organic Chemistry 1 (1988), S. 191-195 
    ISSN: 0894-3230
    Keywords: Organic Chemistry ; Physical Chemistry
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology , Physics
    Notes: The photochemical reaction of MAQO with various aromatic amines were studied by ESR. The results show that nitroxide radicals are stable productrs of the photooxidation of both diphenylamines and phenylamines. The photolyzed phenothiazine does not yield nitroxide as the final product, instead it gives the neutral radical as the stable final product.
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  • 48
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    Journal of Physical Organic Chemistry 1 (1988), S. 333-349 
    ISSN: 0894-3230
    Keywords: Organic Chemistry ; Physical Chemistry
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology , Physics
    Notes: New series of platinum complexes of cyclopropenylidenes of the types of PtX2(CP)2 and trans-PtX(PBu3)2(CP) have been synthesized, where CP is di-t-butylcyclopropenylidene (BCP) or bis(diisopropylamino)cyclopropenylidene (ACP). The 13C-NMR chemical shifts, and 13C-195Pt coupling constants (1JPtC) for the complexes are discussed in comparison with those values derived from closely related series of compounds, trans-PtCl(PR3)2L; L — —CH3, —C6H5 and —C≡CBu-t. An excellent linear relationship through the origin was obtained between 1JPtC and the formal ‘s’ % character of the carbon directly bonded to Pt for the series trans-PtCl(PR3)2L in which the Pt—C bond is regarded as a pure σ-linkage, whereas 1JPtC deviates largely from this relationship when pπ—dπ bonding interaction possibly exists in the Pt—C bond. The NMR data suggest the strong nmr trans-influence of the cyclopropenylidenes and that in the Pt—CP bond the σ-interaction is appreciable but the π-interaction is negligible.
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  • 49
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    Cell Motility and the Cytoskeleton 5 (1985) 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 50
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    Cell Motility and the Cytoskeleton 5 (1985) 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 51
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    Cell Motility and the Cytoskeleton 5 (1985), S. 447-461 
    ISSN: 0886-1544
    Keywords: chemokinesis ; orthokinesis ; klinokinesis ; polymorphonuclear leucocytes ; locomotion ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Evidence is presented to show that klinokinesis, which was previously demonstrated in bacteria and amoeba only, may also occur in metazoan cells. The chemotactic peptide formyl-Met-Leu-Phe (fMLP) elicited orthokinetic and klinokinetic responses of human blood-borne polymorphonuclear leucocytes (PMNs) under the test conditions used. Increased speed (orthokinesis) was due to an increase in the proportion of migrating cells as well as in the speed of the locomoting subset. The klinokinetic effect was manifested by a decrease in the klinolocomotion index, the mean angle of changes in direction ≥ 90°, and the frequency of turns ≥ 90°. The klinolocomotion index was inversely related to speed. This explains the synergistic effect of klinokinesis and orthokinesis in this system. Colchicine alone had and orthokinetic effect which was exclusively due to alterations in the proportion of migrating cells and it altered the turning behaviour without exerting a klinokinetic effect. However, colchicine had marginal orthokinetic and klinokinetic effects on fMLP-stimulated cells resulting in reduced translocation. The relationship between klinokinesis and mean angle or frequency of turns has been analysed. Klinokinesis was a substantial though not the major element of the chemokinetic response to fMLP under the conditions used. No other metazoan cells have been shown to possess such a complete pattern of responses, including orthokinesis, klinokinesis, and chemotaxis, which regulate locomotion.
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  • 52
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    Cell Motility and the Cytoskeleton 5 (1985), S. 355-375 
    ISSN: 0886-1544
    Keywords: Newt ; lung ; cilia ; cell models ; ciliary coordination ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Demembranated ciliated cell models are useful for studying mechanisms responsible for the regulation of ciliary coordination and waveform. This paper describes procedures for isolating ciliated cells from the newt, Taricha granulosa, by trypsin dissociation, their subsequent demembranation by Triton X-100, and their reactivation with MgATP to produce highly motile, coordinated, ciliated cell models. Reactivation of cell models with a high degree of mechanochemical coupling depended on avoiding mechanical damage and maintaining optimal conditions during all stages of isolation and reactivation. Highly motile models were prepared from cells incubated in trypsin, treated briefly with EDTA, separated by gentle agitation, and concentrated by centrifugation at low gravitational forces. Optimal demembranation and reactivation conditions were similar to those described previously for isolated newt lung axonemes. Under these conditions, nearly 100% of the models were reactivated when provided with MgATP and 90-95% beat with coordinated waves. The ciliary tufts beat at frequencies within the range measured in living cells and their reactivated motility was stable for at least 30 min at constant MgATP. These highly coupled models were used to show (1) that development of coordination in the ciliary tuft occurs at a higher substrate concentration range (10-25 μM) than that required to initiate motility per se (2-10 μM); (2) that outer dynein arms may not contribute to beat frequency at substrate concentrations below 35 μM; and (3) that vanadate has effects both on beat frequency and coordination of the tufts.
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  • 53
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    Cell Motility and the Cytoskeleton 6 (1986), S. 305-313 
    ISSN: 0886-1544
    Keywords: cytoplasmic streaming ; Setcreasea purpurea ; intracellular particle movements ; intercellular transport ; azide ; low temperature ; calcium ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Cytoplasmic streaming and its response to azide and low temperature were examined by using high-resolution video-enhanced light microscopy in Setcreasea purpurea staminal hair cells of immature flowers. Particles and organelles examined moved along well-defined pathways, in repeated and unequal saltatory steps, at different rates and sometimes against the main direction of flow (bidirectionally) in both transvacuolar strand and peripheral cytoplasm. Particle movements were reversibly inhibited with azide. Low temperatures caused transvacuolar strands to shift or break. This cytoplasm accumulated in areas outside of the vacuole where spherosomes continued to saltate, but not along well-defined pathways. In the peripheral cytoplasm, however, the spherosomes continued to move normally, amyloplasts became swollen, and they plus the other organelles (except spherosomes) were stationary. Normal particle movements were obtained when chilled cells were rewarmed to 27°C for ca 15 min.
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  • 54
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    Cell Motility and the Cytoskeleton 5 (1985), S. 475-489 
    ISSN: 0886-1544
    Keywords: digital image processing ; flagella ; cilia ; bends ; Hemicentrotus ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A novel method of digital image analysis of the bends of eukaryotic flagella and cilia was devised. In the analysis system, all image pixels were systematically extracted and processed to measure angular direction and curvature. Simulation experiments on theoretical model pictures of flagella with sine-generated or arcstraight line bending waves demonstrated that the method can be used with considerable high accuracy. This method then revealed abrupt changes in slope of the curvature in sperm flagella and embryo cilia of the sea urchin, Hemicentrotus pulcherrimus. This indicates that the digital image processing used may be helpful in the study of flagellar and ciliary movements.
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  • 55
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    Cell Motility and the Cytoskeleton 6 (1986) 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 56
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    Cell Motility and the Cytoskeleton 6 (1986), S. 225-228 
    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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  • 57
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    Cell Motility and the Cytoskeleton 6 (1986), S. 347-353 
    ISSN: 0886-1544
    Keywords: axoplasmic transport ; flagella ; microtubule ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The generation of bending waves by microtubules in squid nerve axoplasm has been modelled using appropriately modified versions of computer programs developed previously for simulation of flagellar bending waves. The results confirm that a constant longitudinal force directed along the axis of the microtubule is sufficient to cause the generation of regular oscillations and propagated bending waves when the forward gliding movement of the microtubule is obstructed. No control mechanism is required to modulate the active force-generating system. In order to obtain bending waves similar to those observed experimentally, it was necessary to use a model for the force-generating system in which the active force decreases with increasing sliding velocity. If the elastic bending resistance of axoplasmic microtubules is similar to that of microtubules in sperm terminal filaments, the longitudinal force per unit length generated by the axoplasmic microtubules must be of the same order of magnitude as the force generated by dynein arms along the doublet microtubules of eukaryotic flagella.
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  • 58
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    Cell Motility and the Cytoskeleton 6 (1986), S. 339-346 
    ISSN: 0886-1544
    Keywords: axonal transport ; human nerve ; video-enhancement ; digital image processing ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The mechanism by which organelles are transported bidirectionally in axoplasm is still unknown; however, evidence of a key role for microtubules in many nonmammalian models has been established. We have observed common or shared tracks within the axoplasm of human nerves along which multiple organelles of varying size and shape are bidirectionally transported. Organelles traveling anterogradely and retrogradely were visualized by video-enhanced differential interference contrast optics and analyzed with the aid of computer-image-processing techniques.Speeds of translocating organelles were determined at eight to 16 translocation points along a path or “track.” Each translocation speed was plotted against its corresponding position on the track to develop a “speed/position diagram.” Regardless of mean organelle speed or direction of motion, organelles sharing a common track exhibited similar patterns of “speeding up” and “slowing down” relative to position along the track. Speed position data for organelles translocating the local axonal region of a common track showed no unique patterns (not different from a uniform distribution, p 〈 0.05). The unique speed/position patterns exhibited by common tracks were not necessarily related to the patterns of other tracks in the immediate vicinity (distance between tracks of 〈 0.50 μm). These findings suggest that (1) there are “common tracks” shared by organelles moving retrogradely and anterogradely; (2) both the organelles and the “track” associated with its translocation play a role in the resultant motion of that organelle; (3) the influence exerted by a common track on the motion of an organelle results in a pattern of speed changes related to position along the track.
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  • 59
    ISSN: 0886-1544
    Keywords: Allogromia ; cytoplasmic transport ; microtubules ; reticulopod withdrawal ; tubulin-containing paracrystal ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Bundles of microtubules (MTs) are readily visualized in vivo by videomicroscopy in highly flattened reticulopodia of the foraminiferan protozoan Allogromia sp. strain NF. In this report we use videomicroscopy, immunocytochemistry, and high-voltage electron microscopy to characterize the dynamic changes that occur in this extensive MT cytoskeleton, and in the associated cytoplasmic transport, during induced withdrawal and subsequent reextension of reticulopodia. Within seconds after application of the withdrawal stimulus (seawater substitute made hypertonic with MgCl2) intracellular bidirectional transport along linear MT-containing fibrils ceases and is replaced by an inward, constant-velocity flow of cytoplasm along the fibrils. As withdrawal continues, most fibrils become wavy and coalesce to form phase-dense pools. These wavy fibrils and phase-dense pools contain a paracrystalline material and few if any MTs. Same-section correlative immunofluorescence and high-voltage electron microscopy reveal that the paracrystalline material contains tubulin. During recovery linear fibrils (MTs) rapidly extend from the phase-dense pools (paracrystals), which concurrently shrink in size, thus reestablishing normal network morphology and motility. We conclude that the MT cytoskeleton in Allogromia reticulopodia is transfonned during withdrawal into a tubulin-containing paracrystal, which serves as a temporary reservoir of MT protein and an initiation site for MT regrowth.
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  • 60
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    Cell Motility and the Cytoskeleton 6 (1986), S. 355-362 
    ISSN: 0886-1544
    Keywords: centriole ; DNA synthesis ; cell cycle ; Chinese hamster ovary cells ; taxol ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The relationship between centriole formation and DNA synthesis was investigated by examining the effect of taxol on the centriole cycle and the initiation of DNA synthesis in synchronized cells. The centriole cycle was monitored by electron microscopy of whole-mount preparations [Kuriyama and Borisy, J. Cell Biol., 1981, 91:814-821]. A short daughter centriole appeared in perpendicular orientation to each parent during late G1 or early S and elongated slowly during S to G2. Addition of 5-20 μg/ml taxol to a synchronous population of cells in S phase did not inhibit centriole elongation; rather, elongation was accelerated. In contrast, when taxol was added to M phase or early G1 cells, centriole duplication was completely inhibited. The taxol block was reversible since nucleation and elongation of centrioles resumed as soon as the drug was removed. Cells exposed to taxol progressed through the cell cycle and became blocked in mitosis, as indicated by an increase in the mitotic index, but eventually the mitotic arrest was overcome, resulting in formation of multinucleated cells. A peak in mitotic index was seen in the following generation, indicating that chromosomes duplicated in the presence of taxol. Incorporation of 3H-thymidine followed by autoradiography confirmed that DNA synthesis was initiated in the presence of taxol even though formation of daughter centrioles was inhibited. It seems, therefore, that centriole duplication is not a prerequisite for entry into S phase. Since DNA synthesis has already been demonstrated not to be necessary for centriole duplication, these two events, normally coordinated in time, appear to be independent of each other.
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    Cell Motility and the Cytoskeleton 6 (1986), S. 389-405 
    ISSN: 0886-1544
    Keywords: cell membrane complex ; extracellular matrix ; fibronectin ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Intermediate filaments (IF) were found in close proximity to the plasma membrane in substrate attached baby hamster kidney cells (BHK-21) and chick embryo fibroblasts (CEF) as well as cells removed from their substrate in the absence of trypsin. However, in cells removed with trypsin, it appeared that IF had retracted away from the membrane. In cells with abundant extracellular matrix (ECM), colchicine induced massive cables of IF, which appeared to interact with specialized areas of the inner plasma membrane. In cells lysed to extract most microfilaments and cytoplasmic constituents, the intact IF network which remained was closely associated with the ECM. From these ultrastructural observations it was concluded that IF interact in some way with a “cell membrane complex” defined as comprising the plasma membrane and molecules attached to its inner and outer surfaces.In order to investigate the possibility that components of the membrane complex may co-isolate with IF, native intermediate filaments (NIF) were prepared. In addition to the structural subunits and other associated polypeptides, a ∼220 kd species which reacted specifically with antibodies directed against the ECM protein fibronectin (FN) was observed; 220 kd was still present after NIF were isolated under pH conditions where FN is more soluble, suggesting that its presence was not simply due to the coprecipitation of two insoluble proteins. Immunofluorescence and immunogold localization confirmed that FN is a component of the cell membrane complex with which IF appeared to interact.
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  • 62
    ISSN: 0886-1544
    Keywords: marginal band ; spectrin ; vimentin ; surface-associated cytoskeleton ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Platinum-carbon (Pt-C) replicas of freeze-dried erythrocyte cytoskeletons of the toad, Bufo marinus, were prepared using a modified Balzers 300 system. Examination in stereo of replicas of the microtubule-containing marginal band revealed filaments projecting from the microtubule walls to form links between adjacent microtubules. These cross-bridging proteins may bundle the microtubules into the configuration of the marginal band (MB) and may also serve to stabilize the structure. The MB appears to have linkages to components of the surface-associated cytoskeleton (SAC). The SAC forms a continuous matrix that spreads across the upper and lower surfaces of the cell adjacent to the plasma membrane and extends around the outer perimeter of the MB. Thus, the SAC encapsulates the MB and the central nucleus. After lysis, the elements of the cytoskeleton remain in a configuration similar to that found in the whole cell. Spectrin (fodrin) and actin were identified by immunofluorescence in the region of the SAC. When labeled with antibodies specific for vimentin and synemin, a network of intermediate filaments can be detected in the region between the nucleus and the MB. These vimentin filaments are also enclosed within the SAC and appear in Pt-C replicas to emerge from the area of the nuclear envelope. As the filaments extend toward the periphery of the cell, they form attachments to the SAC. Attachments of intermediate filaments to both the nucleus and the SAC thus appear to anchor the nucleus in its central position within the cytoskeleton.
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    Cell Motility and the Cytoskeleton 6 (1986) 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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    Cell Motility and the Cytoskeleton 6 (1986), S. 419-427 
    ISSN: 0886-1544
    Keywords: colcemid ; nocodazole ; kinetochores ; microtubules ; spermatocytes ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Reversal of meiotic arrest in crane-fly spermatocytes by U.V. irradiation of Colcemid-arrested cells or by rinsing Nocodazole-arrested cells in fresh buffer results in the induction of chromosome malorientation. Malorientations observed among Colcemid-recovering and Nocodazole-recovering spermatocytes at frequencies higher than normally observed in untreated cells included associations of sister kinetochores of half-bivalents with both spindle poles (amphitely), in contrast with associations of sisters with only one pole (syntely) as is usually found during the first meiotic division. In several cases, prior to anaphase onset, maloriented bivalents appeared unusually tilted with respect to the spindle axis, and during anaphase they gave rise to laggard half-bivalents that did not segregate during anaphase along with half-bivalents having proper syntelic orientation. The results parallel previous findings obtained during cold recovery, and the properties of the drugs used here suggest that their action on microtubules, although reversible, induces malorientation during recovery from meiotic arrest.
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    Cell Motility and the Cytoskeleton 6 (1986), S. 428-438 
    ISSN: 0886-1544
    Keywords: kinetochores ; spindle apparatus ; anaphase ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We investigated the involvement of kinetochore microtubules (kMTs) in mediating chromosome-to-pole connections in crane-fly (Nephrotoma suturalis and Nephrotoma ferruginea) spermatocytes. Two experimental treatments were used to yield spindles with reduced numbers of nonkinetochore microtubules (nkMTs). Short-term (10-15 min) exposure of spermatocytes to 2°C caused depolymerization of the majority of nkMTs, resulting in a kMT:(kMT + nkMT) ratio of 0.76. Long-term (24h) exposure to 2°C followed by recovery at 6°C resulted in a kMT:(kMT + nkMT) ratio of 0.55, the spindle having more nkMTs than a 2°C-treated spindle but fewer than an untreated spindle, in which the kMT:(kMT + nkMT) ratio was 0.27. The numbers and lengths of kMTs in 6°C-grown spindles were similar to those in untreated cells, suggesting that the overall inhibition of MT assembly at 6°C apparently did not affect the mechanism by which kMTs are formed. We observed most kMTs of early anaphase spindles to be long (〉3 μm), and many extended to the polar regions of the spindle. Thus, the crane-fly spindle appears not to be as atypical as it was previously suggested to be.
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    Cell Motility and the Cytoskeleton 6 (1986), S. 406-418 
    ISSN: 0886-1544
    Keywords: Intermediate filaments ; microfilaments fibroblast cell spreading ; focal center ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Spreading and fully spread chick embryo fibroblasts (CEF) were examined by double-label fluorescence microscopy using the actin-specific probe rhodamine-phalloidin and an antibody directed against CEF intermediate filaments (IF). During midspreading, a striking relationship became discernible: statistical analysis showed that approximately half of the cell population exhibited one or more phase-dense, phalloidin-binding nodules that appeared to act as foci from which IF diverged. Coincidence between actin-containing structures and IF was not limited to these centers; IF could also frequently be seen running in close parallel arrays with stress fibers.Ultrastructural analysis confirmed the presence of non-membrane-bound out-pocketings along the length of stress fibers from which 10-nm IF diverged. These structures varied in size and shape, and displayed a dense, fine fibrillar appearance. IF and microfilaments (MF) were distinguished by size and by decoration of MF with myosin subfragment-1. Other IF-MF interactions were seen in cells of all stages: IF were observed to loop through stress fibers, most frequently at the cell margins. In colchicine-treated cells, IF became redistributed into cables that often ran parallel and appeared to merge with stress fibers. Cytochalasin D-treated CEF exhibited loose aggregates of actin-containing material that appeared to be associated with IF.These results suggest the possibility of an interaction between actin-containing structures and IF, particularly during cell spreading in cultured fibroblasts.
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    Cell Motility and the Cytoskeleton 6 (1986), S. 439-447 
    ISSN: 0886-1544
    Keywords: dynamics of actomyosin fibril ; microfilament bundle ; NBD-phallacidin ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Dynamic changes in the spatial organizations of cytoplasmic fibrils (microfilament bundles) related to the contraction-relaxation cycle in thin-spread plasmodia of Physarum polycephalum were investigated by fluorescence microscopy, where NBD-phallacidin was used to stain the fibrils, combined with polarizing light microscopy.The fibrillar organization in the anterior region, which consists of a fanlike spreading plasmodial sheet, strikingly changed according to the phase of the cycle. In the early stage of the contraction, as the endoplasm began to stream backward, the fibrils developed into a number of slender and flabby fibrils emanating from the inside of the cell membrane and the nodes. They became thicker and more straightforward fibrils running parallel to each other at the middle stage, and finally formed a thick framework consisting of a “polygonal network” near the tip of the migrating front and a “parallel array” in the inner part. In the relaxation phase, as the endoplasm streamed forward, the fibrillar framework disintegrated gradually and finally disappeared almost completely, remaining only around the nodes in some cases.The fibrillar patterns in the posterior region, which consists of ramified strands, showed no conspicuous rhythmic change with alternation of the streaming direction.
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  • 68
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    Keywords: cytoplasmic fibril ; birefringence ; microfilament ; contraction-relaxation cycle ; Life and Medical Sciences ; Cell & Developmental Biology
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    Notes: The contractility of Physarum plasmodium was investigated using cell models that were prepared by treating thin-spread plasmodia with ice-cold 0.2% Triton X-100. Cell models obtained from the anterior regions of the thin-spread plasmodia in the contraction phase retained many birefringent cytoplasmic fibrils. The fibrils vigorously contracted on addition of ATP, inducing simultaneous contraction of the whole cell models. In contrast, cell models prepared from the anterior regions in the relaxation phase scarcely contained the birefringent fibrils and exhibited only weak contractility on addition of ATP. The posterior regions of the thinspread plasmodia, which were composed of ramified plasmodial strands, always retained many fibrils when treated with the Triton solution and showed intensive contraction on addition of ATP.SDS-polyacrylamide gel electrophoresis showed that the model was enriched for actin and myosin. About 40% of the actin was extracted from the plasmodium by the Triton treatment, while scarcely any myosin was extracted.Fragmin, a F-actin-fragmenting factor, caused the birefringent fibrils to diminish in the presence of Ca2+, but more than 30 minutes was required for their complete disappearance. The birefringent fibrils weakened by 30-minute fragmin treatment disappeared immediately on addition of ATP or AMP-PNP.
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    Cell Motility and the Cytoskeleton 6 (1986), S. 469-478 
    ISSN: 0886-1544
    Keywords: plant microtubules ; mitosis ; cytokinesis ; plant cell culture ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Treatment with 10 μm taxol disrupted mitotic and cytoplasmic arrays of microtubules (MT) in cultured cells of two higher plants, Vicia hajastana (vetch) and Zinnia elegans. When treated for 1, 24, and 48 h, cells in both cultures showed similar effects. After 1 h, multipolar arrays of MT were noted in prophase, large aster-like arrays of MT appeared in metaphase, and extra MT shared poles with otherwise normal-appearing metaphase and anaphase configurations. After 24 and 48 h, some phragmoplasts were multipartite or misplaced. In interphase cells, micronuclei and multinucleate cells were evidence of irregular mitosis and cytokinesis. Cytoplasmic MT in elongated cells were oriented parallel to, instead of at right angles to the long axis of the cell. Some interphase cells lost asymmetry while maintaining organized arrays of MT. Taxol appears to disrupt mitotic and cytoplasmic arrays of MT, seemingly overriding the mechanism(s) regulating MT polymerization and orientation.
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    Cell Motility and the Cytoskeleton 6 (1986), S. 458-468 
    ISSN: 0886-1544
    Keywords: cytoskeleton ; axoplasm ; fast flow ; quick-freeze ; deep-etch ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Bidirectional organelle movements were observed in fresh and permeabilizedreactivated (0.02% saponin, 5 mM Mg++ ATP) walking leg axons of crayfish with video-enhanced contrast, differential interference contrast (AVEC-DIC) microscopy; and the cytoskeletal organization of those axons was studied with quickfreeze, deep-etch electron microscopy (QF,DE) to understand the structure of the microtubule (MT) domain and to determine the basic cytoskeletal structures necessary for organelle transport in vivo. Vesicles and mitochondria moved bidirectionally in the central parts of fresh or permeabilized-reactivated axons. Although the axoplasm of the fresh axon was composed of longitudinally oriented microtubules and granular materials in which membrane organelles were embedded, a network of fine strands existed in the core of the granular materials. Crossbridges between membrane organelles and microtubules were present. In the central part of reactivated axons, the cytoskeleton consisted of microtubules, highly anastomosing networks of fine strands (6.6 ± 1.4 nm in width) that crosslinked the microtubules with each other, and relatively short, straight crossbridges (25 ± 3.9 nm in length, 5.5 ± 2.1 nm in width) crosslinking membrane organelles with microtubules. It has been shown that a 270KD microtubule associated protein (MAP) could be a main component of crossbridges between MTs [Hirokawa, 1986]. Hence the dynamic conformational change of crossbridges between membrane organelles and microtubules could play an important role when membrane organelles are transported.
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    Cell Motility and the Cytoskeleton 6 (1986), S. 479-484 
    ISSN: 0886-1544
    Keywords: motility ; chemotaxis ; chemoattractant ; cytoskeleton ; folic acid ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Living vegetative amoebae of NC-4H Dictyostelium discoideum were studied to determine if a variety of pteridines had any effect on the filopodia. We observed that production, elongation, and branching of these filopodia were stimulated by pteridines that are chemoattractants for cells of this strain. This stimulation occurs at chemotactically effective concentrations and is observed before motility is evident. A relationship between filopodia and chemoattractant signal processing is discussed.
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    Cell Motility and the Cytoskeleton 6 (1986), S. 485-491 
    ISSN: 0886-1544
    Keywords: PMN chemotaxis ; PMN storage ; PMN locomotion ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Previous studies of the storage of polymorphonuclear leukocytes (PMNs) have used an empirical approach to define “optimal” conditions. To date, no storage conditions have been described which satisfactorily preserve the chemotactic function of PMNs beyond 24 h. In an effort to define the precise nature of the storage lesion, we studied the chemotactic locomotion of freshly isolated PMNs and PMNs which had been suspended in citrate-phosphate-dextrose-adenine (CPD-Al) plasma and stored in PVC bags, at 20-22°C for 24 h. We used time-lapse video recording and computer image analysis to quantitate the motion of PMNs migrating under agarose. The positions of individual motile cells were traced at 1-min intervals for 5 min. The following parameters were used to quantitate migration: (1) speed (distance/min), (2)) persistence of locomotion index (velocity/speed), (3) orientation angle (the angle of the vector describing the next displacement of a cell relative lo a direct line toward the chemoattractant), and (4) chemotropic index (cosine of the orientation angle). After 24 h of storage, the following changes were observed: (1) fewer cells migrated, (2) (he speed of migrating cells was reduced by 25%, (3) the persistence of locomotion index decreased by 7%, which indicates that migrating cells made slightly more/wider turns, and (4) the chemotropic index was decreased by 30%, which indicates that migrating cells were less accurate in their orientation toward the chemoattractant. Apparently, the storage of PMNs selectively impairs the ability of some cells to orient accurately in a chemotactic gradient and changes the distribution of these locomotor parameters within the population.
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    Cell Motility and the Cytoskeleton 6 (1986), S. 492-501 
    ISSN: 0886-1544
    Keywords: chromosome orientation ; prometaphase ; meiosis ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: At metaphase in crane-fly primary spermatocytes, the two sister kinetochores at the centromere of each homologue in a bivalent normally are adjacent and face the same pole; one homologue has all its kinetochore microtubules (kMTs) extending toward one pole and its partner has all its kMTs extending toward the opposite pole. In contrast, during recovery from exposure to 2°C, one or both homologues in many metaphase bivalents had bipolar malorientations: all kMTs of one kinetochore extended toward one pole and some or all those of its sister extended toward the other. Metaphase sister kinetochores that had most of their kMTs extending toward the same pole were adjacent, and those with most extending toward opposite poles were separated from each other. Distances between homologous centromeres were similar to those in properly oriented bivalents. Maloriented bivalents were tilted relative to the spindle axis, and analysis of living cells showed that tilted configurations were rare during prometaphase in untreated cells but frequently arose in cold-recovering cells as initial configurations, then persisted through metaphase. This was in contrast to unipolar configurations of bivalents (configurations suggesting orientation of both homologous centromeres toward the same pole), which always reoriented shortly after the configuration arose. We conclude that in cold-recovering cells, bipolar malorientations are more stable than unipolar malorientations, and the orientation process is affected such that bipolar malorientations arise in bivalents upon initial interaction with the spindle and persist through metaphase.
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    Cell Motility and the Cytoskeleton 6 (1986), S. 502-509 
    ISSN: 0886-1544
    Keywords: respiratory cilia ; dynein ; ATPase ; cystic fibrosis ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Isolated ciliary axonemes from pig trachea were exposed to increasing concentrations of purified Pseudomonas aeruginosa rhamnolipid. This is a defined ciliary system allowing observation of direct impairment of functional axonemes. Axonemal motility and ATPase activity were decreased in proportion to rhamnolipid concentrations. ATPase-associated proteins observed in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and dynein arms seen in ultrastructural cross sections progressively disappeared from axonemes with exposure to rhamnolipid. These four independent measures establish that the rhamnolipid removes the ATPase-containing outer dynein arms from the ciliary axoneme, thereby rendering the axoneme immotile.
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    Cell Motility and the Cytoskeleton 6 (1986), S. i 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 6 (1986) 
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    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
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    Cell Motility and the Cytoskeleton 6 (1986), S. 521-533 
    ISSN: 0886-1544
    Keywords: intracellular organelle transport ; microtubules ; microfilaments ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Reticulomyxa is a large, multinucleated freshwater protozoan with striking intracellular transport. Cyloplasmic streaming and saltatory movements of individual organelles (at rates of up to 25 μm/sec) are observed within the naked cell body and the extensive reticulate peripheral network of fine cytoplasmic strands. As demonstrated by video-enhanced light microscopy, individual organelles move only when associated with cytoskeletal linear elements. The linear elements are composed of mixed colinear bundles of microtubules and actin filaments, which form the backbone of the reticulopodial network. The constant branching, sprouting, and fusion of network stands suggest unique membrane properties and an unusually dynamic cytoskeleton. The electrophoretic mobility of Reticulomyxa tubulins and the lack of crossreactivity with several antibodies known to react with many plant and animal tubulins suggest that they may differ from other tubulins more widely than might be expected. Reticulomyxa's large size, the rapidity and pervasiveness of the two forms of transport, and the simple and ordered cytoskeleton make the organism well suited for future studies on the mechanisms of intracellular transport.
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    Cell Motility and the Cytoskeleton 6 (1986), S. 537-548 
    ISSN: 0886-1544
    Keywords: microtubules ; sea urchins ; kinesin ; mitosis ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: In this report, we describe an in vitro system for analyzing microtubule-based movements in supernatants of sea urchin egg and embryo homogenates. Using video enhanced DIC microscopy, we have observed bidirectional saltatory particle movements on native taxol-stabilized microtubules assembled in low speed supernatants of Lytechinus egg homogenates, and gliding of these microtubules across a glass surface. A high speed supernatant of soluble proteins, depleted of organelles, microtubules, and their associated proteins supports the gliding of exogenous microtubules and translocation of polystyrene beads along these microtubules. The direction of microtubule gliding has been determined directly by observation of the gliding of flagellar axonemes in which the (+) and (-) ends could be distinguished by biased polar growth of microtubules off the ends. Microtubule gliding is toward the (-) end of the microtubule, is ATP sensitive, and inhibited only by high concentrations of vanadate. These characteristics suggest that the transport complex responsible for microtubule gliding in S2 is kinesin-like. The implications of these molecular interactions for mitosis and other motile events are discussed.
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    Keywords: microinjection ; mitosis ; microtubule-associated proteins ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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    Notes: Microtubule-associated protein 2 (MAP2) derivatized with iodoacetamidotetramethylrhodamine or with iodoacetamidofluorescein binds to microtubules after injection into living interphase cells [Scherson et al, 1984]. The binding of derivatized MAP2 stabilized microtubules in vitro; it was therefore important to check if the binding of MAP2 in vivo perturbed the dynamics and organization of the microtubule network. We have addressed these questions by studying the effect of the injection of derivatized MAP2 on mitosis in PtK 1 cells and on the recovery of the microtubule network from low temperature incubation in interphase cells. We found that the presence of derivatized MAP2 did not change the duration of any mitotic stage and that the injected cell normally completed mitosis. We subsequently showed that the injected MAP2 bound to the microtubules within 5 minutes after injection and remained bound throughout the course of mitosis. The reorganization of the microtubule network upon cooling and rewarming was studied in the cytoplasm of human foreskin fibroblasts (356 cells). During the recovery, the distribution of the fluorescent MAP2 in living cells was identical with the microtubule pattern visualized by immunofluorescence in lysed and fixed cells.In these experiments, the fluorescent MAP2 bound to microtubules can be considered as a nonperturbing reporter of the microtubule network. This result is discussed in terms of the role of MAPs in the dynamics and organization of microtubules in living cells.
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    Cell Motility and the Cytoskeleton 6 (1986), S. 580-585 
    ISSN: 0886-1544
    Keywords: flagella ; microtubules ; Chlamydomonas ; bending movement ; oscillation ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: When detergent-extracted cell models of Chlamydomonas reinhardtii were left in the presence of 1 mM Mg-ATP for more than 30 minutes flagellar axonemes tended to become frayed into fine bundles of microtubules. Under such conditions, bundles made up of a pair of outer-doublet microtubules displayed oscillatory bending movements of low (〈 2 Hz) frequencies. The two doublet microtubules underwent association-dissociation cycles coupled with gross bending movement. A model is presented to explain this phenomenon by unidirectional sliding interaction between the two microtubules.
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    Cell Motility and the Cytoskeleton 6 (1986), S. 620-627 
    ISSN: 0886-1544
    Keywords: radiolabeled organelle profile ; retrograde transport system ; anterograde transport system ; turnover ; nodes of Ranvier ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: In a preceding study [Blum, J.J., and Reed, M.C. (1985): Cell Motil. 5:507-527], factors responsible for the shape and velocity of the leading edge of the radiolabeled organelle profile were analyzed, but processes that might influence the shape of the plateau-like region behind the advancing wave were ignored. It is now shown that deposition of material from the fast transport system into membrane-associated structures, degradation of such deposited material and its return to the soma by the retrograde transport system, or leakage of radiolabeled material from the axon can account for the shape of the plateau. Furthermore, these processes are compatible with the maintenance of such structural inhomogeneities as the nodes of Ranvier.
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    Cell Motility and the Cytoskeleton 9 (1988), S. 1-8 
    ISSN: 0886-1544
    Keywords: substrate attached materials (SAM) ; chemotaxis ; leukocytes ; adherence ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We describe a technique to visualize substrate-attached materials (SAM) of polymorphonuclear leukocytes (PMN) using the fluorescent lipid analog 1, 1′-dioctadecyl-3,3,3′,3′,-tetramethylindocarbocyanine-perchlorate (DiC18Icc). DiC18Icc was incorporated into the membranes of living cells or SAMs. Since cell preparation does not require fixation, SAMs can be rapidly visualized by fluorescence microscopy. SAMs are generated by subjecting attached cells to a shearing force by rinsing with phosphate-buffered saline (PBS). The SAM-labeling protocol identified a membrane compartment as shown by detergent extraction. The SAMs of PMN leukocytes observed with this technique display complex patterns of interconnecting filaments, foci with radiating filaments, and smooth membranous areas with interconnecting filaments. The sensitivity and nondestructive nature of the DiC18Icc-labeling procedure have allowed us to observe filopodia of motile cells. The results are consistent with the hypothesis that locomotion involves a series of attachment and detachment steps. After 60 minutes of locomotion, these trailing filopodia have been measured at lengths up to 100 μm. The amount of membrane associated with these filopodia accounts for roughly 10% of the total membrane are of resting cells. These data set limits for models of membrane flow during chemotaxis.
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    Cell Motility and the Cytoskeleton 9 (1988), S. 60-72 
    ISSN: 0886-1544
    Keywords: villin ; actin ; rat brush border ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The biochemical properties of villin purified from the brush borders of chicken and rat small intestines were compared, with emphasis on their physical properties and their Ca++-dependent interaction with actin. Like chicken villin, rat villin exists as two isoforms present in equimolar concentrations; the rat isoforms are slightly more acidic than those of chicken villin (6.08 and 6.11 versus 6.26 and 6.34). Rabbit antisera raised against either villin crossreacted with the other one. Like the avian protein, rat villin bundled F-actin at calcium concentrations below 0.1 μM. Above ∼1 μM calcium, it accelerated the rate of actin assembly and restricted filament lengths of F-actin formed either during coassembly with villin or by addition of villin to preformed filaments. The threshold calcium concentration required for effective severing of preformed filaments was approximately tenfold higher than that required for restricting lengths during coassembly. The extent of filament shortening was proportional to the amount of villin present. At a fixed villin concentration, filament length decreased with increasing [Ca++] over a broad range from 10-7-10-4 M. In general, the mean filament lengths and the dispersion about the mean value were lower in samples where filaments were coassembled with villin than when villin was added to preformed filaments.
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    Cell Motility and the Cytoskeleton 9 (1988), S. 48-59 
    ISSN: 0886-1544
    Keywords: axon ; growth cone ; retraction ; taxol ; slow transport ; axonal transport ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Axons in tissue culture retract and shorten if their tips are detached from the substrate. The shortening reaction of the axon involves contractile forces that also arise during normal axonal motility, elongation, and retraction. We studied shortening in axonal segments isolated from their parent axons by transecting the axon between the growth cone and the most distal point of adhesion to the substrate. Within 15-20 minutes after transection, an isolated axonal segment shortened and pulled its tail end toward the growth cone. During the shortening process, long sinusoidal bends arose along the axon. The identical shortening reaction occurs without transection, when the axon tip is detached from the substrate. Pharmacological studies with inhibitors of glycolysis indicate that the shortening mechanisms utilize metabolic energy, presumably ATP. The rate of sinusoidal shortening is similar to both the rate of polymer translocation in the axon by slow axonal transport and the rate of normal axonal elongation. Taxol inhibits the shortening reaction with a similar dose dependence to its inhibition of axonal growth. Together, all these observations suggest that the same basic intracellular motility mechanisms are involved in normal axonal growth, in slow axonal transport, and in the shortening reaction: the intracellular dynamic system that utilizes ATP to generate longitudinal movements of polymers within the axon may be the same mechanism underlying both the retraction and the elongation of the axon.
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  • 85
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    Cell Motility and the Cytoskeleton 6 (1986), S. 96-98 
    ISSN: 0886-1544
    Keywords: microtubules ; evolution ; eukaryotes ; phagotrophy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Earlier hypotheses of the origin of flagella appear untenable in the light of recent evidence on the ancestry of eukaryotes. It is suggested that microtubules and flagella evolved early in eukaryote evolution to enhance phagotrophy.
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  • 86
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    Cell Motility and the Cytoskeleton 6 (1986), S. 89-95 
    ISSN: 0886-1544
    Keywords: Ciliary inclination ; bending reorientation ; power stroke ; ciliary amplitude ; angular velocity ; unipolar sliding transfer ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Axial views of depolarization- and hyperpolarization-dependent activation of the frontal cirri of Stylonychia were cinematically recorded at high rate (250 frames/s) under voltage-clamp. Images of a cirrus performing the cycle were processed by using computer assistance. In responding to the polarity and amplitude of the voltage signal, a cirrus inclines proximally with a particular angle and orientation. The ciliary cycle-always counterclockwise-is superimposing upon steady inclination. Correction for inclination allowed the assessment of the directional change rate and, after inclusion of the amplitude data, the determination of the ciliary angular velocity during the cycle. The method serves to isolate a new ciliary parameter: inclination, and to register precisely parameters of the cycle which may be meaningful for the understanding of the sliding mechanism.
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  • 87
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    Cell Motility and the Cytoskeleton 6 (1986), S. 83-88 
    ISSN: 0886-1544
    Keywords: sea urchin ; spermatozoa ; Triton model ; protein kinase ; cyclic AMP ; phosphoprotein phosphatase ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Flagellar motility of Triton models of sea urchin spermatozoa was reactivated by cyclic AMP-dependent protein kinase and a protein factor, termed motility activator, both of which were prepared from the detergent-extract of sea urchin spermatozoa. It was shown that phosphorylation of the motility activator by the protein kinase is necessary for the reactivation of flagellar motility [Ishiguro et al, J. Cell Biol. 92:777-782, 1982; Murofushi et al, in “Biological Functions of Microtubules and Related Structures,” Academic Press, 1982]. Reactivating factor was also detected in a KCI-extract of the axoneme fraction devoid of the detergent-extractable materials. The activity of this factor was also cyclic AMP- and protein kinase-dependent. Furthermore, when freshly prepared Triton models were treated with phosphoprotein phosphatase prepared from bovine cardiac muscle, the flagellar motility was drastically suppressed. This inhibition of the motility was partially recovered by the addition of cyclic AMP and protein kinase to the phosphatase-treated models.
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  • 88
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    Cell Motility and the Cytoskeleton 6 (1986), S. 99-104 
    ISSN: 0886-1544
    Keywords: flagella ; wave shapes ; motility ; calcium ; adaptation ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: In its normal culturing environment, the trypanosomid flagellate Crithidia oncopelti propagates basally-directed planar waves, but may under certain conditions exhibit base-to-tip wave propagation in what is regarded as an avoidance response. The beat frequency and wave shape in both modes of beating are dependent on the viscosity of the swimming medium; viscosity may also influence the direction of wave propagation. If Crithidia experience a sudden increase in viscosity, there is a marked increase in the proportion of the population that is seen to exhibit wave propagation from base to tip; this proportion gradually decreases with time until the whole sample has reverted to “normal” beating. In a single organism, the resumption of normal beating is not accomplished in a single transition but by a series of switches between the forward and reverse modes. The interval of time between successive switches appears to be random, while the length of time spent in base-to-tip wave propagation gradually decreases. Despite the randomness of the switching process, its rate when averaged over successive time intervals is found to be constant at a particular viscosity and also dependent on it. The precise manner by which this organism is able to control its direction of wave propagation is unclear. However, the switching behavior it exhibits during the period of adaptation to an increased mechanical loading of the flagellum may reflect a process that characterizes a facet of this controlling mechanism.
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  • 89
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    Keywords: actinogelin ; α-actinin ; reconstituted actin-gel ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We studied the properties of actinogelin, a Ca2+-regulated actin cross-linking protein isolated from Ehrlich tumor cells or rat liver. Chicken gizzard α-actinin was used as a Ca2+-insensitive control. Actinogelin, which has very high gelation activity under low Ca2+ conditions, was found using electron microscopic or fluorescence studies to induce formation of a characteristic structure in which actin filaments and bundles radiate to (or converge from) all directions from spot-like core structures. A similar structure was induced with actinogelin, even in the presence of 0 7 saturation of tropomyosin. No such structure was detected with actinogelin under high Ca2+ conditions, and only a few were found with gizzard α-actinin. Because reconstituted structures are similar to those observed intracellularly, actinogelin may be important in the formation of similar microfilament organization in the cells. It seems also important that these structures are reconstituted with only two purified protein components, i.e., actinogelin and actin.Immunocompetition studies showed that actinogelin and gizzard α-actinin partially shared antigenicity, and their molecular shape and peptide maps were similar. Their amino acid compositions [Kuo et al., 1982], subunit and domain structures, and binding sites on actin [Mimura and Asano, 1987] are also very similar. Therefore, it is concluded that actinogelin belongs to α-actinin superfamily proteins. Furthermore, the presence of functionally different subfamilies concerned with Ca2+ sensitivy, gelation-efficiency, and others is discussed. Actinogelin, which induces networks of actin filaments, may be classified as high gelation type.
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  • 90
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    Cell Motility and the Cytoskeleton 10 (1988), S. 349-362 
    ISSN: 0886-1544
    Keywords: immunofluorescence ; cytoplasmic actins ; muscle actins ; epitope ; isoactins ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Two IgG1, κ monoclonal antibodies (Mab) against actin have been obtained from a fusion in which chicken gizzard actin was used as the immunogen. One Mab, designated B4, shows a preferential reactivity toward enteric smooth muscle actin but also cross-reacts with skeletal, cardiac, and aorta actins on the basis of immunoblots, ELISA assays, and indirect immunofluorescence. However, this antibody does not react with either cytoplasmic actin in any of these assay systems. A second Mab, designated C4, reacts with all six known vertebrate isoactins as well as Dictyostelium discoideum and Physarum polycephalum actins. Thus B4 Mab appears to react with an epitope that is at least partially shared among the muscle actins but not found in cytoplasmic actins, while C4 Mab binds to an antigenic determinant that has been highly conserved among the actins. The binding sites of both Mabs on skeletal actin overlap that of pancreatic DNase I. Both antibodies bind a SV8 proteolytic product comprising the amino-terminal two-thirds of the actin molecule, and their epitopes appear to overlap since C4 can compete for the binding of B4 to skeletal actin. Neither antibody is able to prevent actin polymerization.
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  • 91
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    Cell Motility and the Cytoskeleton 9 (1988) 
    ISSN: 0886-1544
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
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  • 92
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    Cell Motility and the Cytoskeleton 9 (1988), S. 9-16 
    ISSN: 0886-1544
    Keywords: adaptation ; cAMP ; cell motility ; chemotaxis ; Dictyostelium discoideum ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: When developing amebae of Dictyostelium discoideum are treated with constant concentrations of cAMP above 10-8 M, the average rate of motility is depressed, with maximum inhibition at roughly 10-6 M. It is demonstrated that shifting the concentration of cAMP from 0 M to concentrations ranging from 10-8 to 10-6 M in a perfusion chamber results in the immediate inhibition of motility. After shifting from 0 M to 10-8 or 10-7 M, the rate of cell motility remains low, then rebounds to a higher level, exhibiting a standard adaptation response. No adaptation is exhibited after a shift from 0 M to 10-6 M, a concentration resulting in maximum inhibition. It is demonstrated that the level of inhibition and the extent of the adaptation period are dependent upon the concentration of cAMP after the shift, and that submaximal inhibition is additive. The characteristics of adaptation in this motility response are very similar to the characteristics of adaptation for the relay system and phosphorylation of the putative cAMP receptor.
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  • 93
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    Cell Motility and the Cytoskeleton 9 (1988), S. 17-29 
    ISSN: 0886-1544
    Keywords: sequestered actin bundles ; polygonal arrays ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Using mainly fluorescence microscopy after rhodamine-phalloidin staining, the F-actin distribution in the mouse lens epithelium was studied with regard to the effects of age, genetic strain, and mechanical injury.These studies have revealed that aside from its association with the plasma membrane the structural organization of F-actin in the mouse lens epithelium in situ is characterized by two major configurations: (1) a filamentous arrangement in such patterns as stress fibers, polygonal arrays (PAs), and meshworks, and (2) a highly concentrated structure called a sequestered actin bundle (SAB).The aging study indicated that the SAB is a consistent character in C57BL/6 mice from the age of 5 wk on, but not in CF1 mice. The size and shape of the SAB change gradually with age as inferred from two-dimensional measurements. The genetic study on the SAB character using hybrids and congenic strains showed that it is inherited as a Mendelian dominant, probably multigenic mode. Finally, the injury study revealed a structural modification in cells around the wound, including flattening of cells at the edge and extension of processes into the wound space. In the rest of the epithelium, injury amplified membrane infolding and fluorescence of polygonal arrays but diminished the size and fluorescence intensity of SABs. These changes are thought to be correlated with wound repair involving cell division and migration.These studies illustrate the variability in F-actin expression in situ in lens epithelial cells that can be induced by intrinsic and extrinsic factors.
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  • 94
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    Cell Motility and the Cytoskeleton 10 (1988), S. 217-228 
    ISSN: 0886-1544
    Keywords: immunogold ; microtubules ; optical sectioning ; video microscopy ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The distribution of F-actin cables in dividing endosperm cells of a higher plant, Haemanthus, was visualized with the immunogold-silver-enhanced method and compared with the arrangement of immunogold-stained microtubules in the same cells. The three-dimensional distribution of F-actin cables and microtubules during mitosis and cell plate formation was analyzed using ultrathin optical sectioning of whole mounts in polarized light video microscopy. F-actin cables form a loose irregular network in the interphase cytoplasm. Much of this network remains outside of the spindle during mitosis. A few F-actin cables were detected within the spindle. Their pronounced rearrangement during mitosis appears to be related to the presence and growth of microtubule arrays. During prometaphase, actin cables located on the spindle surface and those present within the spindle tend to arrange parallel to the long axis of the spindle. Cables outside the spindle do not reorient, except those at the polar region, where they appear to be compressed by the elongating spindle. Beginning with mid-anaphase, shorter actin cables oriented in various directions accumulate at the equator. Some of them are incorporated into the phragmoplast and cell plate and are gradually fragmented as the cell plate is formed and ages. Actin cables adjacent to microtubule arrays often show a regular punctate staining pattern. Such a pattern is seldom observed in the peripheral cytoplasm, which contains few microtubules. The rearrangement of F-actin cables mimicks the behavior of spindle inclusions, such as starch grains, mitochondria, etc., implying that F-actin is redistributed passively by microtubule growth or microtubule-related transport. Thus F-actin or actomyosin-based motility does not appear to be directly involved in mitosis and cytokinesis in higher plants.
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  • 95
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    Cell Motility and the Cytoskeleton 9 (1988), S. 73-84 
    ISSN: 0886-1544
    Keywords: cilia ; metachronal waves ; electron microscopy ; calcium ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Structural and behavioral features of intact and permeabilized Paramecium tetraurelia have been defined as a basis for study of Ca2+ control of ciliary reversal. Motion analysis of living paramecia shows that all the cells in a population swim forward with gently curving spirals at speeds averaging 369 ± 19 μm/second. Ciliary reversal occurs in 10% of the cell population per second. Living paramecia, quick-fixed for scanning electron microscopy (SEM), show metachronal waves and an effective stroke obliquely toward the posterior end of the cell. Upon treatment with Triton X-100, swimming ceases and both scanning and transmission electron microscopy reveal cilia that uniformly project perpendicularly from the cell surface. Thin sections of these cells indicate that the ciliary, cell, and outer alveolar membranes are greatly disrupted or entirely missing and that the cytoplasm is also disrupted. These permeabilized paramecia can be reactivated and are capable of motility and regulation of motility. Motion analysis of cells reactivated with Mg2+ and ATP in low Ca2+ buffer (pCa7) shows that 71% swim forward in straight or curved paths at speeds averaging 221 ± 20 μm/second. When these cells are quick-fixed for SEM the metachronal wave patterns of living, forward swimming cells reappear. Motion analysis of permeabilized cells reactivated in high Ca2+ buffers (pCa 5.5) shows that 94% swim backward in tight spirals at a velocity averaging 156 ± 7 μm/second. SEM reveals a metachronal wave pattern with an effective stroke toward the anterior region. Although the permeabilized cells do not reverse spontaneously, the pCa response is preserved and the Ca2+ switch remains intact. The ciliary axonemes are largely exposed to the external environment. Therefore, the behavioral responses of these permeabilized cells depend on interaction of Ca2+ with molecules that remain bound to the axonemes throughout the extraction and reactivation procedures.
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  • 96
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    Cell Motility and the Cytoskeleton 9 (1988), S. 101-110 
    ISSN: 0886-1544
    Keywords: prokaryotic motility ; periplasmic flagella ; hydrodynamics ; model ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Spirochetes are a group of bacteria with a unique ultrastructure and a fascinating swimming behavior. This article reviews the hydrodynamics of spirochete motility, and examines the motility of the spirochete Leptospira in detail. Models of Leptospira motility are discussed, and future experiments are proposed.The outermost structure of Leptospira is a membrane sheath, and within this sheath are a helically shaped cell cylinder and two periplasmic flagella. One periplasmic flagellum is attached subterminally at either end of the cell cylinder and extends partway down the length of the cell. In swimming cells, each end of the cell may assume either a spiral or a hook shape. Translational cells have the anterior end spiral shaped, and the posterior end hook shaped. In the model of Berg et al., the periplasmic flagella are believed to rotate between the sheath and the cell cylinder. Rotation of the anterior periplasmic flagellum causes the generation of a gyrating spiral-shaped wave. This wave is believed sufficient to propel the cells forward in a low-viscosity medium. The cell cylinder concomitantly rolls around the periplasmic flagella in the opposite direction - which allows the cell to literally screw through a gel-like viscous medium without slippage. This model is presented, and it is contrasted to previous models of Leptospira motility.
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  • 97
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    Cell Motility and the Cytoskeleton 9 (1988), S. 117-128 
    ISSN: 0886-1544
    Keywords: centrosome ; aster-forming activity ; tubulin polymerization ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Mitotic apparatuses (MAs) isolated from sea urchin metaphase eggs were chilled on ice to depolymerize microtubules, homogenized, and incubated with tubulin. This caused formation of many small asters with microtubules focusing on granules which were probably fragments of the centrosome. The aster-forming protein components of the granules in the homogenized MAs were solubilized in 0.5 M KCl containing 50% glycerol. After dialysis against low-ionic-strength buffer solution, proteins congregated to form granular assembly capable of initiating aster formation. Phosphocellulose column chromatography enabled the separation of the aster-forming protein fraction which contained a 51,000 molecular weight protein (51-kd protein) as a major component. The protein fraction possessing the aster-forming activity was also prepared from methaphase whole egg homogenate, and the elution profile of the 51-kd protein on phosphocellulose column also coincided with that of the aster-forming activity. The granular assembly reconstituted from the phosphocellulose fraction formed asters whose microtubules show the same growth rate and length distribution as those of asters reconstructed from the granules in the homogenized MAs. Anti-51-kd protein antibody that was raised in rabbit and affinity-purified stained the center of asters which were reconstructed either from the granules in the homogenized MAs or from the granular assembly reconstituted from the phosphocellulose fraction. These results suggest that the 51-kd protein is a component in the aster-forming activity of the centrosomal component in vitro.
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  • 98
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    Cell Motility and the Cytoskeleton 9 (1988), S. 312-324 
    ISSN: 0886-1544
    Keywords: flagella ; sea urchin spermatozoa ; waveform analysis ; Ciona spermatozoa ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Data obtained by manual digitization of photographs of flagellar bending waves have been analyzed by determining size parameters for the bends by least-squares fitting of a model waveform. These parameters were then used to normalize the data so that the average shape of the bends could be determined. Best fits were obtained with a model waveform derived from the constant curvature waveforms used previously but with provision for a linear change in curvature across the central region of the bend-the gradient curvature model (GCM). The central regions of the GCM bending waves are separated by transition regions with length determined by a parameter called the truncation factor (FT). Fitting the GCM to sine-generated bending waves give optimal fit when FT = 0.34. Fitting the GCM to four different samples of flagellar bending waves gave best fits with values of FT ranging from 0.17 for ATP-reactivated Lytechinus spermatozoa beating at approximately 10 Hz to 0.32 for live spermatozoa of Arbacia. The difference between the Arbacia waveforms and a sine-generated waveform is therefore very small, but a sine-generated waveform lacks the degree of freedom represented by FT that is required to fit other waveforms optimally.The residual differences between the waveform data and optimal GCM waveforms were averaged and found to be small. In most cases, the curvature in the central region of the optimal GCM decreased in magnitude towards the tip of the flagellum; however, this slope was highly variable and sometimes positive. Significant variations in both this slope and FT were found in individual bends as they propagated along a flagellum.
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  • 99
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    Cell Motility and the Cytoskeleton 9 (1988), S. 325-336 
    ISSN: 0886-1544
    Keywords: anaphase ; aster ; mitosis ; motility ; spindle ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: An earlier, laser microbeam study produced evidence that, in Fusarium solani, extranuclear polar forces function at anaphase B of mitosis to pull apart the incipient daughter nuclei, whereas the central spindle functions primarily to limit the rate at which they separate. To elucidate further the various dynamics of mitotic anaphase, 8-14 mitoses in hyphae of F. solani were analyzed at 0.5-2.0-sec intervals using high-resolution, digitally processed, videotaped sequences. The spindle growth rate, although fluctuating frequently, averaged 0.6 μm/min during metaphase, increased to 3.6 μm/min during anaphase A and was maximal at 6.1 μm/min during anaphase B. Commonly, chromosomes migrated poleward during anaphase A at fluctuating rates, the average rate being an unprecedented 7.5 μm/min. During anaphase the mitotic apparatus migrated to and fro in the hyphae at rates of 3-15 μm/min, an apparent effect of opposing, fluctuating and typically unequal cytoplasmic forces applied to the two spindle poles. Thus, the molecular mechanisms underlying the various anaphase movements in F. solani do not operate entirely smoothly and uniformly. Accelerated growth of the central spindle is temporally associated with anaphase A and the development of asters. Thus, chromosome disjunction may allow the polar forces to increase the rate of spindle elongation. Microtubule dynamics and motor molecules appear to be adequate to account for the observed rates of mitotic movements.
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  • 100
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    Cell Motility and the Cytoskeleton 9 (1988), S. 361-374 
    ISSN: 0886-1544
    Keywords: microtubular cytoskeleton ; Dinoflagellates ; immunofluorescence ; Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The cytoskeletal microtubule system has been studied in six species of unarmoured Dinoflagellates using immunofluorescence and electron microscopy. Several structures have been detected and described: (1) a subpellicular layer of microtubules, constituting the microtubular cytoskeleton, running singly or in bundles from the anterior part of the cell to the posterior; (2) a feeding apparatus, containing a ribbon of microtubules, which corresponds to a small peduncle in some species and is simply represented by a cytostome in some other species; and (3) the longitudinal flagellum that runs in a long intracytoplasmic pocket before becoming free at the extremity of the sulcus. A thorough study of the organization of the microtubular structures in a wide spectrum of Dinoflagellates is a prerequisite for understanding the evolutionary history of the group.
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