bluuebunny/crossref_metadata_2025_split huggingface dataset:
Coordinated expression of five tropomyosin isoforms and β‐actin in astrocytes treated with dibutyryl cAMP and cytochalasin D
Authors: Ferrier Richard , Had Laurence , Rabié Alain , Faivre‐Sarrailh Catherine 
Published: February 2005

AbstractCytochalasin D and dBcAMP cause cultured astrocytes to change from flat cells to retrated process‐bearing cells. F‐actin was present throughout cells stimulated with dBcAMP for 16 h, whereas cytochalasin D caused F‐actin to form massive aggregates at the tips of the cell processes. The two drugs differently regulated the expression of both β‐actin and tropomyosin genes in astrocytes cultured in the presence or absence of serum: dBcAMP caused down‐regulation and cytochalasin D caused up‐regulation. Northern blot analyses indicated that: (1) serum deprivation halved the concentration of all tropomyosin transcripts (TM‐1, TM‐2, TM‐4, TMBr‐1, TMBr‐2). Serum induced TM‐4 via transcriptional activation, independent of protein synthesis, (2) dBcAMP induced down‐regulation of β‐actin (–50%) and tropomyosin transcripts (–35 to 52%) even in the presence of serum. The concentration of profilin mRNA decreased in dBcAMP‐reactive astrocytes (‐46%). The decrease in β‐actin mRNA concentration was not blocked by cycloheximide, whereas down‐regulation of tropomyosin transcripts was completely reversed when protein synthesis was inhibited, and (3) cytochalasin D induced an increase in the concentration of tropomyosin transcripts (+ 69 to 185%) which was cumulative with serum stimulation. Cytochalasin D induction of both β‐actin and TM‐4 operated through transcriptional activation, independent of protein synthesis.The production of all tropomyosin transcripts examined here were strictly coordinated with β‐actin expression in serum‐, dBcAMP‐ and cytochalasin D‐treated astrocytes. This indicates that the differential expression of tropomyosin isoforms occurring during astrocyte maturation is due to more complex regulation than that involved in serum‐ or cAMP‐stimulated astrocytes. © 1994 Wiley‐Liss, Inc.
Dynamic instability of microtubules from cold‐living fishes
Authors: Billger Martin , Wallin Margareta , Williams Robley C. , Detrich H. William 
Published: February 2005

AbstractThe dynamic instability of microtubules free of microtubule‐associated proteins from two genera of cold‐living fishes was measured, by means of video‐enhanced differential interference‐contrast microscopy, at temperatures near those of their habitats. Brain microtubules were isolated from the boreal Atlantic cod (Gadus morhua; habitat temperature ∽ 2–15°C) and from two austral Antarctic rockcods (Notothenia gibberifrons and N. coriiceps neglecta; habitat temperature ∽ −1.8 to + 2°C). Critical concentrations for polymerization of the fish tubulins were in the neighborhood of 1 mg/ml, consistent with high interdimer affinities. Rates of elongation and frequencies of growth‐to‐shortening transitions (“catastrophes”) for fish microtubules were significantly smaller than those for mammalian microtubules. Slow dynamics is therefore an intrinsic property of these fish tubulins, presumably reflecting their adaptation to low temperatures. Two‐dimensional electrophoresis showed striking differences between the isoform compositions of the cod and the rockcod tubulins, which suggests that the cold‐adapted microtubule phenotypes of northern and southern fishes may have arisen independently. © 1994 Wiley‐Liss, Inc.
cAMP‐induced morphological changes in an immortalized schwann cell line: A prelude to differentiation?
Authors: De Deyne Patrick G. , De Vries George H. , Bigbee John W. 
Published: February 2005

AbstractSchwann cells (SC), the myelinating cells of the peripheral nervous system, show a remarkable capacity to switch from a differentiated state to a proliferative state both during development and peripheral nerve regeneration. In order to better understand the regulatory mechanisms involved with this change we are studying a Schwann cell line transfected with the SV‐40 large T gene (TSC). Serum‐free medium combined with elevating intra‐cellular cAMP levels produced a slower proliferating TSC whose morphology changed from pleiomorphic to process bearing, reminiscent of primary SC in culture. This change was abrogated by colcemid but was unaltered by cytochalasin D, indicating a major role for microtubules. Ultrastructural studies demonstrated numerous microtubules in the cellular extensions which correlated with strong immunocytochemical staining for tubulin in the processes. Analysis of cytoskeletal fractions from the treated cells revealed a greater proportion of tubulin in the polymerized state compared with untreated cells which closely resembled the distribution in primary SC. The cytoskeletal changes observed in the TSC as a result of elevating the intra‐cellular cAMP levels may reflect the earliest cellular changes in the induction of myelination. © 1994 Wiley‐Liss, Inc.
Kinesin‐related polypeptide is associated with vesicles from Corylus avellana pollen
Authors: Liu Guo‐Qin , Cai Giampiero , Casino Cecilia Del , Tiezzi Antonio , Cresti Mauro 
Published: February 2005

AbstractA 100‐kDa polypeptide with microtubule‐interacting properties was identified in a Golgi vesicle‐enriched fraction from Corylus avellana pollen. The k71s23 antibody (directed to the kinesin heavy chain from bovine brain) [Tiezzi et al., 1992: Cell Motil. Cytoskeleton 21:132–137] localized the polypeptide on the external surface of membrane‐bounded organelles. Some 100‐kDa‐containing vesicles co‐pelleted with microtubules (polymerized from purified bovine brain tubulin) either in presence or absence of 5 mM AMPPNP, but they could be released by 10 mM ATP or 0.5 M KCl. The pollen microtubule‐interacting protein, salt‐extracted from membranes and partially purified by gel filtration, exhibited an ATPase activity (16.2 nmolPi/mg/min) which could be stimulated about 2‐fold (32.5 nmolPi/mg/min) by addition of bovine brain microtubules. We suppose that the 100‐kDa polypeptide is part of a molecular complex showing properties of the kinesin class. © 1994 Wiley‐Liss, Inc.
Localization of NuMA protein isoforms in the nuclear matrix of mammalian cells
Authors: Zeng Changqing , He Dacheng , Brinkley B. R. 
Published: February 2005

AbstractUsing a monoclonal antibody 2D3 generated against a kinetochore‐enriched human chromosome preparation, we identified a high molecular mass protein with nuclear staining in interphase and polar staining of the pericentriolar region in the mitotic spindle. Initially termed centrophilin, this protein associates with the minus‐ends of spindle microtubules (MT) and appears to be important in spindle organization [Tousson et al., 1991: J. Cell Biol. 112:427–440]. Comparison of a partial cDNA sequence obtained for centrophilin with the full length cDNA sequence of nuclear mitotic apparatus protein (NuMA) [Compton et al., 1992: J. Cell Biol. 116:1395–1408; Yang et al., 1992: J. Cell Biol. 116:1303–1317] has indicated that NuMA and centrophilin are the same protein. Using a polyclonal NuMA antibody, we have provided further evidence that NuMA exists as iso‐forms as shown by peptide mapping and immunoblots. Sequential fractionation experiments along with immunofluorescence, immunoblotting, and EM immunogold labeling have demonstrated that NuMA isoforms are novel components of nuclear core filaments. Thus, NuMA, a long coiled‐coil protein, appears to have dual functions in interphase and mitosis during the cell cycle. In interphase, NuMA likely plays a structural role in the nucleoskeleton that may be important in nuclear organization and functions, whereas in mitosis, NuMA appears to be associated with spindle MT organization and chromosome positioning. © 1994 Wiley‐Liss, Inc.
Substructure of cytoplasmic dense bodies and changes in distribution of desmin and α‐actinin in developing smooth muscle cells
Authors: Chou Rong‐Ghi R. , Stromer Marvin H. , Robson Richard M. , Huiatt Ted W. 
Published: February 2005

AbstractThe substructure of assembling cytoplasmic dense bodies (CDBs) and changes in the distribution of desmin and α‐actinin during development of smooth muscle were studied in gizzard samples from 10‐ and 16‐day embryos and from 1‐ and 7‐day post‐hatch chickens. CDBs in these cells lack the density of CDBs in mature or adult smooth muscle cells and, thus, allow observations of the changes inside CDBs. The random filament orientation seen in younger embryonic cells is first modified to include relatively small patches of IFs that are somewhat straighter and are approaching a side‐by‐side arrangement. As development proceeds, the IFs in these arrays become straighter, are parallel over longer lengths of the IFs and later acquire the density characteristic of mature CDBs. Anti‐desmin labeling in embryonic 10‐ and 16‐day cells showed that desmin intermediate filaments (IFs) were located in the myofilament compartment but were concentrated in or near assembling CDBs. Anti‐desmin labeling shifted to the perimeter of CDBs after hatching. Cross sections, longitudinal sections, and stereo pairs all show that IF profiles are present inside unlabeled assembling CDBs. Anti‐α‐actinin labeling was directly on CDBs and was often associated with the cross‐connecting filaments (CCFs) (average diameter of 2–3nm) inside CDBs. We propose, based on these data, that desmin IFs, α‐actinin‐containing CCFs, and actin filaments are the principal components of the substructure of assembling CDBs. We also present a proposed model for CDB assembly. © 1994 Wiley‐Liss, Inc.
Gain setting in Chlamydomonas reinhardtii: Mechanism of phototaxis and the role of the photophobic response
Authors: Zacks David N. , Spudich John L. 
Published: February 2005

AbstractThe unicellular green alga Chlamydomonas reinhardtii maintains sensitivity of its phototaxis response (alignment of swimming direction along the axis of a light beam) over several orders of magnitude of light intensities. It is widely accepted that the rotation of the swimming cell provides temporal comparisons of light intensities via periodic contrast generated by its asymmetrically positioned refractile eyespot organelle. The cells also exhibit a second behavioral response to light called the photophobic (or stop) response, which is a brief cessation of swimming caused by a temporal change in light intensity. The cells are desensitized to photophobic stimuli by light exposure. Through comparative measurements of both responses, we explain the behavioral basis of the large dynamic range of phototaxis in terms of precise desensitization of the photophobic response. The basis of the explanation is that the flagellar beat changes which cause phototactic orientation are the residual of the photophobic response after desensitization (i.e., “mini‐photophobic” reactions which cause brief reorienting motions without a full stop). This interpretation predicts quantitatively the dependence of the extent of desensitization on light intensity and the dependence of onset and maintenance of phototaxis on extent of desensitization. These predictions are tested and confirmed in this report. © 1994 Wiley‐Liss, Inc.
Different reactivity with monoclonal anti‐tubulin antibodies between native and fixed mitotic microtubules in sea urchin eggs
Authors: Oka Mikako T. , Arai Takao , Hamaguchi Yukihisa 
Published: February 2005

AbstractThe effect on fixation on the reactivities of mitotic microtubules with monoclonal anti‐tubulin antibodies was investigated by the indirect immunofluorescence procedure. All of the seven antibodies used intensely stained mitotic microtubules in sea urchin eggs lysed and fixed with methanol at −20°C, whereas only two of them stained the stabilized microtubules in the lysed eggs before the fixation. The other five did not stain the mitotic microtubules even after microtubule components other than tubulin were removed by treating the lysed eggs with 0.4 M KCl solution containing taxol. These results exclude the possibility that the fixation affects proteins, which interact with microtubules including microtubule‐associated proteins (MAPs) and interfere with the binding of monoclonal antibodies with tubulin, and strongly suggest that the fixation directly affects the three‐dimensional conformation of tubulin Furthermore, microinjection of these antibodies indicated the results as follows [combining the results reported previously; Oka et al., 1990: Cell Struct. Funct. 15: 373–378]: The antibodies which stained mitotic microtubules stabilized in the lysed eggs induced disassembly of native mitotic microtubules in the living eggs, but those which did not stain the stabilized microtubules did not disassemble the native microtubules. From these results, it is suggested that the monoclonal antibodies which stain microtubules in the eggs lysed but not fixed are useful for microinjection experiments. © 1994 Wiley‐Liss, Inc.
Switching of the dominant calcium sequestering protein during skeletal muscle differentiation
Authors: Koyabu Sukenari , Imanaka‐Yoshida Kyoko , Ioshii Sérgio O. , Nakano Takeshi , Yoshida Toshimichi 
Published: February 2005

AbstractA major Ca2+‐storing protein in endoplasmic reticulum (ER) of non‐muscle cells is calreticulin (CR), which is considered to be functionally homologous to calsequestrin. Calsequestrin is a Ca2+‐binding protein in sarcoplasmic reticulum (SR) of striated muscle, which stores Ca2+ during muscle relaxation. In order to investigate the expression and distribution of calsequestrin and calreticulin during skeletal muscle differentiation, cultured chick embryonic skeletal muscles were observed by immunofluorescence using anti‐calsequestrin, anti‐calreticulin, antidesmin, and anti‐sarcomeric myosin antibodies and rhodamine‐phalloidin. Within 6 hours in culture, myoblasts started to express desmin. Desmin‐positive cells demonstrated the reticular staining of calreticulin, as did desmin‐negative cells. Around fusion, calsequestrin and sarcomeric myosin started to appear in desmin‐positive cells. The expression of calsequestrin slightly preceded that of sarcomeric myosin. As the myotubes matured, the fluorescent dots of calsequestrin increased and spread to the cell periphery along the myofibrils, while the reticular pattern of calreticulin gradually disappeared. Double labeling showed that calsequestrin colocalized with calreticulin. In mature myotubes, anti‐calsequestrin staining demonstrated many dots along myofibrils, whereas calreticulin was barely seen except at the perinuclear region. These results suggest that the expression of calsequestrin and calreticulin are switched during skeletal muscle differentiation. © 1994 Wiley‐Liss, Inc.
Antibodies that can discriminate between dynein heavy chains and their HUV1 photoproducts
Authors: Sperling Linda , Houari Abdellah , Moudjou Mohammed , Mazarguil Honoré , Wright Michel 
Published: February 2005

AbstractDyneins are multi‐subunit enzymes that transduce chemical energy into the mechanical energy that makes cilia and flagella beat and moves organelles towards the minus end of microtubules. The ATPase activity is borne by heavy chains, and recent molecular analysis indicates that dynein heavy chain genes form an ancient multigene family: the similarity between the same isoform of two distantly related species is greater than that between different isoforms of the same species. We have exploited sequence identities between a Paramecium axonemal dynein heavy chain gene cloned in our laboratory and sequences of dynein heavy chains from other species to prepare antibodies against active‐site peptides capable of recognizing dynein heavy chains regardless of species or isoform. One of the antibodies is perfectly specific for the larger product of V1 photolysis (HUV1) and thus incorporates a unique property of the hydrolytic ATP binding site of all known dynein heavy chains, the capacity for photocleavage in the presence of micromolar vanadate. Our characterization of these reagents suggests that they will be useful for biochemical and in situ studies of known dyneins as well as identification of potential new members of the family. © 1994 Wiley‐Liss, Inc.
Phosphoprotein phosphatase 1 (PP1) is a component of the isolated sea urchin mitotic apparatus
Authors: Johnston Jennifer A. , Sloboda Roger D. , Silver Robert B. 
Published: February 2005

AbstractA protein component of isolated mitotic apparatus having a relative molecular mass of 62,000 (p62) is a substrate of a calcium/calmodulin dependent protein kinase, and the phosphorylation of p62 in vitro correlates directly with microtubule disassembly. In vivo experiments have determined the phosphorylation of p62 increases after fertilization; maximum incorporation of phosphate occurs during late metaphase/early anaphase and decreases thereafter. Because the level of p62 is constant throughout the cell cycle [Johnston and Sloboda, 1992: J. Cell Biol. 119:843–54] the decrease in phosphorylation of p62 observed after anaphase onset is most likely due to the action of a phosphatase. By examination of the relative amount of phosphorylated p62 which remained radiolabeled as a function of time using a standard in vitro phosphorylation assay, the activity of a phosphoprotein phosphatase capable of dephosphorylating p62 in the isolated mitotic apparatus was observed. To characterize the p62 phosphatase, okadaic acid and calyculin A were used to inhibit the dephosphorylation of p62 in vitro. It was found that specific concentrations of okadaic acid (50–500 nM) and of calyculin A (10–100 nM) were effective at inhibiting the dephosphorylation of p62 in vitro. Lower concentrations of either inhibitor had a negligible effect on dephosphorylation of p62. These data indicate the presence of phosphoprotein phosphatase type 1 activity associated with mitotic apparatus isolated from sea urchin embryos using the procedures described here. The implications of these findings relative to our understanding of the regulation of mitosis and cytokinesis are discussed. © 1994 Wiley‐Liss, Inc.
Characterization of a monoclonal antibody prepared against plant actin
Authors: Andersland John M. , Fisher Deborah D. , Wymer Carol L. , Cyr Richard J. , Parthasarathy M. V. 
Published: February 2005

AbstractAnti‐actin monoclonal antibodies were prepared using phalloidin‐stabilized actin that was purified from pea roots by DNase I affinity chromatography. One monoclonal antibody, designated mAb3H11, bound plant actin in preliminary screenings and was further analyzed. Immunoblot analysis showed that this antibody had a high affinity for plant actin in crude and purified preparations but a low affinity for rabbit muscle actin. In immunoblots of plant extracts separated on two‐dimensional gels it appeared to bind all actin isoforms recognized by the JLA20 anti‐chicken actin antibody. Using immunofluorescent cytochemistry, the antibody was used to observe actin filaments in aldehyde‐fixed and methanol‐treated tobacco protoplasts. These results indicate that mAb3H11 should be a useful reagent for the study of plant actins. © 1994 Wiley‐Liss, Inc.
Myosin reorganization in activated RBL cells correlates temporally with stimulated secretion
Authors: Spudich Annamma 
Published: February 2005

AbstractRat basophilic leukemia cells secrete histamine and serotonin in response to cross‐linking of the IgE receptor by multivalent antigen [Metzger et al., 1986: Ann. Rev. Immunol. 4:419–470]. Receptor crosslinking also induces phosphorylation of the light and heavy chains of myosin II with kinetics similar to that of secretion [Ludowyke et al., 1989: J. Biol. Chem. 264:12492–12501]. Here we show that myosin II localization changes after activation with similar kinetics. Furthermore, these changes are coincident with changes in cell shape and increase in motile activity induced by activation. Within 2 min, activated cells begin to flatten, spread on their substratum, and extend lamellipodia which show active ruffling. Quantitation of the extent of cell spreading from video micrographs shows that 48% of the cells increase significantly in surface area by 5 min and 71% by 15 min. Myosin II is uniformly distributed in unactivated cells but is deficient in newly formed lamellipodia that start to appear at 2 min after activation. In contrast these lamellipodia show strong staining for actin. Further changes in myosin organization are detected by 15 min after activation when myosin reappears in the cell periphery, is concentrated in the perinuclear area, and is also organized in punctate linear arrays that extend from the nucleus to the cell periphery. The kinetics of the early cell shape changes and formation of the myosin‐deficient lamellipodia correlate well with, and may relate to, the increase in the level of myosin II phosphorylation reported by Ludowyke et al. [1989: J. Biol. Chem. 264:12492–12501]. Changes in the distribution of cell surface‐bound IgE also occur upon antigen activation, and they correlate with the myosin distribution in a manner that suggests that they may be driven by myosin II. © 1994 Wiley‐Liss, Inc.
Protein tyrosine phosphorylation during sea urchin fertilization: Microtubule dynamics require tyrosine kinase activity
Authors: Wright Shirley J. , Schatten Gerald 
Published: February 2005

AbstractProtein tyrosine phosphorylation plays an important role in cell growth, mitosis, and tumorigenesis. It has also been implicated in meiotic maturation and fertilization. We have used anti‐phosphotyrosine immunofluorescence and immunoblotting to identify sperm and egg proteins which are phosphorylated on tyrosine residues prior to and during sea urchin fertilization. On immunoblots of sperm proteins, the monoclonal anti‐phosphotyrosine antibody detected three major proteins with molecular weights of 44, 82, and 100 kD, and six minor bands at 46, 48, 70, 76, 95, and 150 kD. These phosphotyrosyl proteins were localized to the sperm acrosomal and centriolar fossae. In contrast, staining was found globally in unfertilized eggs, and the antibody recognized two major egg phosphotyrosyl proteins of molecular weights 42 and 50 kD, and five minor bands at 40, 90, 116, 130, and 150 kD. While immunofluorescent staining remained throughout the fertilized egg cytoplasm, there were dynamic changes in the staining intensity of single bands. The 90 kD immunoreactive band increased in intensity, and the 40 and 42 kD bands disappeared by 15 min after fertilization. Loss of the 40 and 42 kD bands was due to dephosphorylation by okadaic acid‐sensitive phosphatase(s). The 50 kD immunoreactive protein was unchanged up to the 8‐cell stage and was still present in blastulae, indicating its importance throughout fertilization and early development. Alterations in the pattern of phosphotyrosine‐containing proteins during fertilization did not depend on nascent proteins and could not be completely mimicked by increasing intracellular calcium, pH, and protein kinase C activity alone. Since changes in the fertilization pattern of phosphotyrosyl proteins occurred during formation of the sperm aster and mitotic spindle, we analyzed the role of protein tyrosine kinase activity in these processes using the tyrosine kinase specific inhibitor, erbstatin. Both the sperm aster and mitotic spindle were disrupted, indicating an involvement of tyrosine phosphorylation in these processes during interphase and mitosis. We conclude that the changes in phosphotyrosyl proteins play an important role in fertilization and early development of sea urchin eggs. Control of microtubule assembly into the sperm aster and mitotic spindle of the first cell cycle are examples of such roles. © 1995 Wiley‐Liss, Inc.