bluuebunny/crossref_metadata_2025_split huggingface dataset:
EGTA induces prolonged summed depolarizations in Mytilus gill coupled ciliated epithelial cells: Implications for the control of ciliary motility
Authors: Stommel E. W. , Stephens R. E. 
Published: February 2005

AbstractAbfrontal 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.
Localization of mitotic‐apparatus‐associated 51‐kD protein in unfertilized and fertilized sea urchin eggs
Authors: Ohta Kunihiro , Toriyama Masaru , Endo Sachiko , Sakai Hikoichi 
Published: February 2005

AbstractThe 51‐kD protein, a protein component of the mitotic apparatus in sea urchin eggs, is involved in the aster‐forming activity previously shown in vitro [Toriyama et al., 1988]. Postembedding immunofluorescent labelings of eggs from fertilization through first cleavage showed that the 51‐kD protein is localized in sperm asters, centrosomal regions, spindles, basal regions of astral microtubules, and regions surrounding daughter nuclei at telophase in situ. Immunofluorescence and immunoblot analyses detected the 51‐kD protein uniformly in unfertilized eggs, but not in spermatozoa. When unfertilized eggs were treated with taxol, the 51‐kD protein was shown to be associated with taxol‐induced cytasters. Immunoblot analysis revealed that similar protein species are present in the mitotic apparatus of other species of sea urchin. It was suggested that the 51‐kD protein may be involved in microtubule nucleation and microtubule matrix in sea urchin eggs in vivo.
B‐band protein in sea urchin sperm flagella
Authors: Inaba Kazuo , Mohri Toshiko , Mohri Hideo 
Published: February 2005

AbstractA high‐molecular‐weight polypeptide, named B‐band, was partially purified from sea urchin sperm flagella using selective extraction, hydroxylapatite chromatography, and sucrose density gradient centrifugation. The molecular weight of the B‐band was 440,000 by continuous system of sodium dodecyl sulphate‐polyacrylamide gel electrophoresis (SDS‐PAGE). Sedimentation coefficient of the B‐band protein was 10.5 S, and its Stokes radius was 10 nm. When examined by low‐angle rotary shadowing electron microscopy, this molecule appeared to be composed of four globular heads and two curved linkers (“double headphone shape”), which was quite different from the shape of 21 S dynein, the outer arm dynein. Flagellar axonemes were also subjected to several chemical dissections. The B‐band was not extracted with treatments that remove both arm structures but was solubilized with treatments that extract other components such as radial spokes and nexin links. The B‐band protein in the axoneme was also more susceptible to trypsin digestion than the arm structures. These results suggest that the B‐band protein is a “double headphone‐shaped” component of the axonemal structures and makes up the elastic structure that might regulate the active sliding between adjacent doublet microtubules.
Ionic control of locomotion and shape of epithelial cells: II. Role of monovalent cations
Authors: Bereiter‐Hahn Jürgen , Vöth Monika 
Published: February 2005

AbstractThe migration of keratocytes isolated from Xenopus tadpole epidermis has been investigated in vitro. In saline the cells move with a mean speed of 5–6 μm/min. Migration is slowed down in saline with diminished sodium content and ceases in media containing not more than 4 mM sodium. Inhibition of the Na+/K+‐2Cl− cotransporter by piretanide reduces the speed of migrating cells to about one‐third of the control level, the same accounts to inhibition of the Na+/H+ antiport with amiloride at pH 7.2. At pH 6.6, however, amiloride only slightly influences locomotion. Depolarization of the plasma membrane by increased extracellular K+ concentration or by inhibition of the Na+/K+ pump by ouabain is only of minor influence during more than 1 h. Hyperpolarization of the cells using the sodium ionophore monensin impedes locomotion; this inhibition depends on an active Na+/K+ pump. Ionophore‐mediated breakdown of the K+ gradient strictly inhibits locomotion. The experiments have shown that a continuous flux of sodium ions is indispensable for the maintenance of cell locomotion. These ions may exert their action primarily by affecting cytosolic free calcium concentration and pH.
Maturation of hagfish gland thread cells: Composition and characterization of intermediate filament polypeptides
Authors: Spitzer Robert H. , Koch Elizabeth A. , Downing Stephen W. 
Published: February 2005

AbstractPrevious studies with the hagfish, a primitive vertebrate, have shown that the gland thread cells (GTCs) each contain a single thread (∼60 cm long in average‐sized cells) in the form of a concisely coiled cytoskeletal entity destined for export by holocrine secretion. The thread in relatively immature GTCs consists almost entirely of intermediate filaments (IFs) bundled in parallel alignment with far fewer microtubules (MTs). The three thread polypeptides described earlier (α, basic; β acidic; γ, most acidic; each with a Mr of 63–64 kD) are now further evaluated with respect to in vitro assembly, cross‐reactivity with IF polypeptides from higher vertebrates, and peptide sequence homology with known IF polypeptides. The overall results mainly suggest that the hagfish polypeptides are keratinlike substances but lamins or a new type of IF is not ruled out. However, cross‐reactivity is weak with mammalian keratins; the 8–11‐nm filaments formed from mixtures of α and γ in vitro are generally linear rather than the curvilinear structures usually formed by keratin and nonkeratin IFs; and mixtures of α and β tend to yield 9–12‐nm granules or granular strings. Polypeptide analyses on GTCs segregated on the basis of maturational stage show a progressive increase in β/γ values which correlates with cell maturation, but the α/(β+γ) ratios remain near 1. Inasmuch as β and γ have many similar properties, the documented increase in the amount of the β component in aging GTCs might in part be the result of a failure in a posttranslational modification system and may contribute to the ultrastructural changes that accompany thread maturation in preparation for holocrine secretion and subsequent modulation of the viscoelastic properties of mucus.
Electron spectroscopic imaging of the intercellular bridge
Authors: Rattner J. B. , Bazett‐Jones D. P. 
Published: February 2005

AbstractPhosphorylated proteins have been implicated in the control and regulation of many cellular events. One of the striking features of microtubule organizing centers is that they are rich in phosphoproteins [Vandre et al., 1984]. In this report we describe the application of electron spectroscopic imaging to the study of the spatial and temporal distribution of phosphorus and hence to the study of phosphorylated proteins associated with the formation and organization of one such organizing center, the intercellular bridge. Our study indicates that there are several discrete microtubule‐based domains that ultimately come to lie within the intercellular bridge and that there is a unique pattern of phosphorylation–dephosphorylation in some of these domains. These patterns correlate with the concurrent growth and sliding of antiparallel microtubules and the development of interactions among neighboring microtubules.
Functional diversity among spectrin isoforms
Authors: Coleman Thomas R. , Fishkind Douglas J. , Mooseker Mark S. , Morrow Jon S. 
Published: February 2005

AbstractThe purpose of this review on spectrin is to examine the functional properties of this ubiquitous family of membrane skeletal proteins. Major topics include spectrin‐membrane linkages, spectrin‐filament linkages, the subcellular localization of spectrins in various cell types and a discussion of major functional differences between erythroid and nonerythroid spectrins. This includes a summary of studies from our own laboratories on the functional and structural comparison of avian spectrin isoforms which are comprised of a common alpha subunit and a tissue‐specific beta subunit. Consequently, the observed differences among these spectrins can be assigned to differences in the properties of the beta subunits.
Contributions of the β‐subunit to spectrin structure and function
Authors: Coleman Thomas R. , Fishkind Douglas J. , Mooseker Mark S. , Morrow Jon S. 
Published: February 2005

AbstractThe three avian spectrins that have been characterized consist of a common α‐subunit (240 kD) paired with an isoform‐specific β‐subunit from either erythrocyte (220 or 230 kD), brain (235 kD), or intestinal brush border (260 kD). Analysis of avian spectrins, with their naturally occurring “subunit replacement” has proved useful in assessing the relative contribution of each subunit to spectrin function. In this study we have completed a survey of avian spectrin binding properties and present morphometric analysis of the relative flexibility and linearity of various avian and human spectrin isoforms. Evidence is presented that, like its mammalian counterpart, avian brain spectrin binds human erythroid ankyrin with low affinity. Cosedimentation analysis demonstrates that (1) avian erythroid protein 4.1 stimulates spectrin‐actin binding of both mammalian and avian erythrocyte and brain spectrins, but not the TW 260/240 isoform, (2) calpactin I does not potentiate actin binding of either TW 260/240 or brain spectrin, and (3) erythrocyte adducin does not stimulate the interaction of TW 260/240 with actin.In addition, a morphometric analysis of rotary‐shadow images of spectrin isoforms, individual subunits, and reconstituted complexes from isolated subunits was performed. This analysis revealed that the overall flexibility and linearity of a given spectrin heterodimer and tetramer is largely determined by the intrinsic rigidity and linearity of its β‐spectrin subunit. No additional rigidity appears to be imparted by noncovalent associations between the subunits. The scaled flexural rigidity of the most rigid spectrin analyzed (human brain) is similar to that reported for F‐actin.
cAMP‐independent and cAMP‐dependent protein phosphorylations by isolated goldfish xanthophore cytoskeletons: Evidence for the association of cytoskeleton with a carotenoid droplet protein
Authors: Palazzo Robert E. , Lynch Thomas J. , Taylor John D. , Tchen T. T. 
Published: February 2005

AbstractTriton‐insoluble cytoskeleton of nonpigment cells has bound protein kinase that phosphorylates, with or without added cAMP, tubulins and the intermediate filament proteins p60, p56, p53, and p45a to give multiple charge variants. In the absence of 8‐Br‐cAMP, Triton‐insoluble cytoskeletons from xanthophores also phosphorylate p60, p56, and p45a, but not p53; tubulin phosphorylation may also be reduced. In the presence of 8‐Br‐cAMP, p53, as well as several other peptides, are phosphorylated. One of these latter peptides was identified as the carotenoid droplet (pigment organelle) protein p57, whose phosphorylation and dephosphorylation precede pigment dispersion and aggregation respectively (Lynch et al.: J. Biol. Chem. 261:4204–4211, 1986). The amount of pp57 produced depends on the state of pigment distribution in the xanthophores used to prepare the cytoskeletons for labeling. With cytoskeletons from xanthophores with aggregated pigment, pp57 is a major labeled phosphoprotein seen in two‐dimensional gels. With cytoskeletons prepared from xanthophores with dispersed pigment, the yield of labeled pp57 is greatly reduced (by at least 90%). Together with earlier results, we propose that, in the aggregated state, p57 serves to bind carotenoid droplets to the cytoskeletons, most likely the microtubules. The significance of other cAMP‐dependent phosphorylation reactions is unknown but may be related to cAMP‐induced cytoskeleton rearrangement in intact xanthophores.
Evidence that intermediate‐like filaments are found in the micronucleus of Paramecium bursaria during prophase
Authors: Lewis Larry M. 
Published: February 2005

AbstractThe micronuclear spindle apparatus in Paramecium bursaria was studied by electron microscopy during prophase, metaphase, and anaphase of the first meiotic division. During prophase, the spindle apparatus consists mostly of intermediate‐like filaments, relatively few spindle microtubules, and unique cone‐shaped structures termed microlamellae. Microlamellae join the ends of chromosomes to the fibrous elements of the spindle. The capacity to preserve the intermediate‐like filaments is largely dependent upon the use of collidine buffer during fixation. In contrast, during metaphase and anaphase, microtubules are the dominant fibrous element of the spindle. The microtubules interact with chromosomes during these phases by joining to true kinetochores. Neither treatment with cytochalasin B or fixation with a low concentration of osmium tetroxide affects the development of intermediate filaments during prophase. Because intermediate‐like filaments are abundant during prophase and microtubules are more common during metaphase and anaphase, the structural differences may reflect differences in the mechanisms for chromosome movement.
Long‐term observation of cultured cells by interference‐reflection microscopy: Near‐infrared illumination and Y‐contrast image processing
Authors: Zand Martin S. , Albrecht‐Buehler Guenter 
Published: February 2005

AbstractInterference‐reflection microscopy (IRM) is the only method presently available with which to visualize cell‐substratum adhesions in living tissue culture cells continuously for long periods of time without the use of fluorescent markers (Curtis: J. Cell Biol. 20:199–215, 1964; Izzard and Lochner: J. Cell Sci. 21:129–159, 1976). This method utilizes approximately 1% of the incident illumination to produce the IRM image (Verschueren: J. Cell Sci. 75:279–301, 1985) and so far has required the use of high‐intensity light sources in the visible spectral range (400–800 nm). Unfortunately, visible light of this intensity and spectral range induces marked changes in the behavior and morphology of motile fibroblasts, including cessation of locomotion. In contrast, the present paper reports that continuous observations of live cells in IRM for periods of up to 8 hours are possible if the illuminating light is in the red to near‐infrared range (650–950 nm) and without any observable change in normal cell morphology or behavior. In addition, we describe how the technique of Y‐contrast image processing can be applied to IRM images to create a three‐dimensional image of the ventral cell surface topography.
Mitoskelin: A mitochondrial protein found in cytoskeletal preparations
Authors: Price Maureen G. , Gomer Richard H. 
Published: February 2005

AbstractA 70 kD protein, which we have named mitoskelin, is highly enriched in cytoskeletal preparations from bovine cardiac muscle. Mitoskelin has three main variants with isoelectric points between 5.6 and 5.8. Immunoblotting with polyclonal antibodies directed against mitoskelin shows that, like intermediate filament proteins, the majority of mitoskelin resists solubilization from a myocardial homogenate by a series of extraction solutions ranging from very low salt to 0.6 M KI buffers and by 0.1–1% Nonidet P‐40 detergent. By double‐label immunofluorescence on cells and tissues, mitoskelin is colocalized with the mitochondrial marker cytochrome c oxidase. Mitoskelin is associated with the inner membranes of mitochondria as shown by immunoelectron microscopy and immunoblotting Immunological cross‐reactivity and similarities of molecular weight, pI, distribution, and chromatographic properties indicate that mitoskelin is the 70 kD component of complex I (NADH: ubiquinone oxidoreductase), a portion of the mitochondrial oxidative phosphorylation system. No function or activity has yet been demonstrated for the 70 kD component of the 25‐polypeptide complex I. Dialysis against physiological buffers allows purified, urea‐solubilized mitoskelin to form 10 nm wide filamentous structures that do not closely resemble intermediate filaments. These results suggest the exciting possibility that mitochondria may contain a membrane‐associated filamentous skeleton.
Rapid growth cone translocation on laminin is supported by lamellipodial not filopodial structures
Authors: Kleitman N. , Johnson M. I. 
Published: February 2005

AbstractTo determine the relationship between growth cone structure and motility, we compared the neurite extension rate, the form of individual growth cones, and the organization of f‐actin in embryonic (E21) and postnatal (P30) sympathetic neurons in culture. Neurites extended faster on laminin than on collagen, but the P30 neurites were less than half as long as E21 neurites on both substrata. Growth cone shape was classified into one of five categories, ranging from fully lamellipodial to blunt endings. The leading margins of lamellipodia advanced smoothly across the substratum ahead of any filopodial activity and contained meshworks of actin filaments with no linear f‐actin bundles, indicating that filopodia need not undirlie lamellipodia. Rapid translocation (averaging 0.9–1.4 μm/min) was correlated with the presence of lamellipodia; translocation associated with filopodia averaged only 0.3–0.5 μm/min. This relationship extended to growth cones on a branched neurite where the translocation of each growth cone was dependent on its shape. Growth cones with both filopodial and lamellipodial components moved at intermediate rates. The prevalence of lamellipodial growth cones depended on age of the neurites; early in culture, 70% of E21 growth cones were primarily lamellipodial compared to 38% of P30 growth cones. A high percentage of E21 lamellipodial growth cones were associated with rapid neurite elongation (1.2 mm/day), whereas a week later, only 16% were lamellipodial, and neurites extended at 0.5 mm/day. Age‐related differences in neurite extension thus reflected the proportion of lamellipodial growth cones present rather than disparties in basic structure or in the rates at which growth cones of a given type moved at different ages. Filopodia and lamellipodia are each sufficient to advance the neurite margin; however, rapid extension of superior cervical ganglion neurites was supported by lamellipodia independent of filopodial activity.
Selective reduction of anaphase B in quinacrine‐treated PtK1 cells
Authors: Armstrong L. , Snyder Judith A. 
Published: February 2005

AbstractQuinacrine, an acridine derivative which competitively binds to ATP binding sites, has previously been shown to cause the reorganization of metaphase spindle microtubules (MTs) due to changes in interactions of non‐kinetochore microtubules (nkMTs) of opposite polarity (Armstrong and Snyder: Cell Motil. Cytoskeleton 7:10–19, 1987). In the study presented here, mitotic PtK1 cells were treated in early anaphase with concentrations of quinacrine ranging from 2 to 12 μM to determine energy requirements for chromosome motion. The rate and extent of chromosome‐to‐pole movements (anaphase A) were not affected by these quinacrine treatments. The extent of anaphase B (kinetochore‐kinetochore separation) was reduced with increasing concentrations of quinacrine. Five micromolar quinacrine reduced the extent of kinetochore‐kinetochore separation by 20%, and addition of 12 μM quinacrine reduced the kinetochore‐kinetochore separation by 40%. To determine the role of nkMTs in anaphase spindle elongationquinacrine‐treated metaphase cells were treated with hyperosmotic sucrose concentrations, and spindle elongation was measured (Snyder et al.: Eur J. Cell Biol. 39:373–379, 1985). Metaphase cells treated with 2–10 μM concentrations of quinacrine for 2–5 min reduced spindle lengths by 10–50% prior to 0.5 M sucrose treatment for 5 min. This treatment showed a significant reduction in the ability of sucrose to induce spindle elongation in cells pretreated with quinacrine. As spindle length and birefringence was reduced by quinacrine treatment, sucrose‐induced elongation was concomitantly diminished. These data suggest that quinacrine‐sensitive linkages are necessary for anaphase B motions. Reduction in these linkages and/or MT length in the nkMT continuum may reduce the ability of the nkMTs to hold compression at metaphase. This form of energy is thought to drive a significant proportion of normal anaphase B in PtK1 cells and sucrose‐induced metaphase spindle elongation.
Nonuniform behavior of multiple isoactins in the same cell is a cell‐dependent phenomenon
Authors: Shires Ann K. , Rubenstein Peter A. 
Published: February 2005

AbstractThe functional significance of multiple isoactins in the same cell is still not understood. To address this question, we examined the response of smooth muscle and cardiac muscle α‐isoactins to a serial extraction procedure applied to both muscle and nonmuscle cell types. We compared these extraction results with results obtained with the β‐ and γ‐nonmuscle actin isoforms from the same cells. In differentiated BC3H1 nonfusing muscle cells (smooth muscle α‐isoactin), in human rhabdomyosarcoma cells (cardiac α‐isoactin), and in chick skeletal muscle cells (cardiac α‐isoactin), different fractions were found selectively enriched in either the nonmuscle or the muscle‐specific actin isoforms compared with their relative abundance in whole cell extracts. Conversely, when these same isoactins were examined either in undifferentiated BC3H1 cells or in mouse nonmuscle cells stably transfected with a cardiac α‐isoactin gene, no enrichment of these isoforms above their relative abundance in whole cell extracts was observed. These results indicate that within the muscle or muscle‐like cells examined, the different actin isoforms were either selectively utilized or localized. These results further show that isoactin‐specific responses observed were apparently related to the cell type in which they were found and not to differences in inherent physical properties such as solubility of the different isoactins examined.