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Volume 10 (1985)
- Issue 4 Pages 315-
- Issue 3 Pages 195-
- Issue 2 Pages 95-
- Issue 1 Pages 1-
Volume 10, Issue 4
Displaying 1-12 of 12 articles from this issue
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Yoshito Tominaga, Shoshi Muto, Teruo Shimmen, Masashi Tazawa1985 Volume 10 Issue 4 Pages 315-325
Published: 1985
Released on J-STAGE: April 18, 2008
JOURNAL FREE ACCESSThe presence of calmodulin (CaM) was demonstrated in the homogenate of Chara internodal cells by the radioimmunoassay method. It amounted to 400 ng per ml homogenate.
The possible involvement of CaM in cytoplasmic streaming was examined by applying CaM-binding drugs to intact cells and plasmalemma-perme-abilized cells. Thirty μM of trifluoperazine (TFP) applied to intact Chara cells did not interfere with the cessation of streaming accompanying membrane excitation, which causes a transient increase in cytoplasmic Ca2+ concentra-tion. After a long exposure to TFP, the action potential resulted in an irreversible depolarization and inhibited the recovery of streaming due to loss of the semipermeability of the plasma membrane.
In permeabilized cells, neither the motile system nor the Ca2+-induced cessation was inhibited by TFP up to 100 μM, by fluphenazine (FPH) at 1 μM, or by N-(6-aminohexyl)-5-chloro-1-naphthalene sulfonamide (W-7) at 10 μM. However, the recovery of streaming induced by washing away the Ca2+ was completely inhibited by these drugs. On the other hand, these drugs did not affect the streaming of tonoplast-free cells, irrespective of the presence or absence of 10 μM Ca2+. Thus cytoplasmic streaming is controlled by two Ca2+-sensitizing components, one related to the streaming cessation and the other, probably CaM, related to recovery.
FPH at 50 μM or W-7 at 100 μM, even in the absence of Ca2+, stopped the cytoplasmic streaming of permeabilized cells. Washing away the drugs did not lead to streaming recovery after W-7 treatment, but sometimes did after FPH treatment.View full abstractDownload PDF (1216K) -
Daisuke Takahashil, Hiromu Murofushi, Koichi Ishiguro, Jun Ikeda, Hiko ...1985 Volume 10 Issue 4 Pages 327-337
Published: 1985
Released on J-STAGE: April 18, 2008
JOURNAL FREE ACCESSPhosphoprotein phosphatase prepared from bovine cardiac muscle was used to study the roles of axonemal phosphoproteins in the flagellar motility of sea urchin spermatozoa. When isolated axonemes were incubated with cyclic AMP-dependent protein kinase, γ[32P]ATP and cyclic AMP, more than 15 polypeptides were phosphorylated. Most were dephosphorylated by treatment with phosphoprotein phosphatase.
When Triton models of sea urchin spermatozoa were treated with phospho-protein phosphatase followed by an addition of ATP, the flagellar motility of the models was drastically reduced in comparison with that of the untreated models. The motility of the phosphatase-treated Triton models was partially restored by an addition of cyclic AMP and cyclic AMP-dependent protein kinase. These data give strong support to the idea that the motility of eukaryo-tic flagella is controlled by a protein phosphorylation-dephosphorylation system.View full abstractDownload PDF (3269K) -
Kozo Utsumi, Koichi Nobori, Shigeo Terada, Masanobu Miyahara, Toshihik ...1985 Volume 10 Issue 4 Pages 339-348
Published: 1985
Released on J-STAGE: April 18, 2008
JOURNAL FREE ACCESSThe amount of free calcium in the cytoplasm is important in stimulation coupled with a number of cellular functions. The putative iono-phoretic action of membrane lipid metabolites on Ca2+ offers convenient explanation of the stimulation-coupled mobilization of cytoplasmic Ca2+. To analyze the ionophoretic action of the lipid metabolites, we devised a sensitive method to study Ca2+ transport that uses liposome-entrapped Quin 2. A cal-cium ionophore, A23187, increased the fluorescence intensity of the Ca2+-Quin 2 complex as a function of Ca2+ transport into liposomes.
A similar Ca2+ flux into the liposomes was induced by phospholipase A2 (PLA2) and by various long-chain fatty acids in liposomes that consist of phospholipids containing unsaturated fatty acids. The potencies of the fatty acids for Ca2+ transport is inversely correlated with their melting points. The oxidized products of the unsaturated fatty acids increased the Ca2+ and nons-pecific permeability of the biological membranes. These results suggest that stimulation-coupled PLA2 activation might mediates the mobilization of cytoplasmic Ca2+.View full abstractDownload PDF (927K) -
Tsuneo Takadera, Tetsuro Mohri1985 Volume 10 Issue 4 Pages 349-359
Published: 1985
Released on J-STAGE: April 18, 2008
JOURNAL FREE ACCESSAddition of Ca2+ ionophore (A23187) to the medium stimulated the Na+-independent leucine transport in Chang liver cells, increasing the cytoplasmic free Ca2+ concentration, irrespective of the presence or absence of extracellular Ca2+ Anticalmodulin drugs, such as chlorpromazine, trifluoperazine, and W-7, significantly inhibited the leucine transport in the cells. The stimulatory effect of A23187 on leucine transport was completely blocked in the presence of the anticalmodulin drug. Two microtubule dis-rupting drugs, colchicine and colcemid, significantly stimulated leucine transport. On the other hand, taxol, a microtubule stabilizing agent, decreased the stimulatory effect of colchicine on the leucine transport. These results strongly suggest the involvement of Ca2+ and calmodulin in regulation of Na+-independent leucine transport, possibly through control of assembly and disassembly of the microtubule network.View full abstractDownload PDF (1142K) -
Takahiro Tsuchiya, Kazuoand Tobe, Hideo Nagashima1985 Volume 10 Issue 4 Pages 361-372
Published: 1985
Released on J-STAGE: April 18, 2008
JOURNAL FREE ACCESSNon-parenchymal fat-storing cells (FSCs) were isolated from rat liver by Metrizamide density centrifugation after liver perfusion with media containing pronase and/or collagenase. These cells were cultured in Williams' E medium with 20 % fetal calf serum under the usual conditions. FSCs, identi-fied by vitamin A-specific fluorescence (VAF), were viewed under conventional light microscopy 24, 48 and 72 h after seeding. At 24 h, some FSCs remained hemispheric; at 48 h all FSCs were well spread out, and at 72 h the number of FSCs had increased and the VAF intensity had become weaker. Scanning electron microscopy revealed that the features of the FSCs clearly differed from those of the other non-parenchymal cells. The in vitro FSC features closely resembled the in vivo features. FSCs cultured for 48 h had flattened, triangular or oval shapes with fairly smooth surfaces that showed scattered microvilli 0.1 μm long. Two types of processes, long slender (130 μm maximum length) and short broad ones, protruded from the cell body. These processes had smaller projections 0.2 μm wide that looked like fern leaves. Many lipid droplets could be seen under the cell surface.
IgG-coated sheep erythrocytes (EAs) and IgM-C3-coated ox erythrocytes (EACs) were used to investigate whether there are Fc and C3b receptors on the FSCs. No rosette-formation characteristic of EAs or EACs was present on the FSCs; proof that FSCs have neither receptor.View full abstractDownload PDF (1137K) -
Yukio Ishimi, Wakao Sato, Masatoyo Kojima, Kaoru Sugasawa, Fumio Hanao ...1985 Volume 10 Issue 4 Pages 373-382
Published: 1985
Released on J-STAGE: April 18, 2008
JOURNAL FREE ACCESSA nucleosome assembly protein (AP-I) was purified approxi-mately 50 % from the cytosol of HeLa S3 cells by three purification steps. Using this protein fraction as an antigen, we established three stable hybridomas that secrete monoclonal antibodies specific for AP-I by the conventional method of cell fusion. Immunoblotting of the HeLa S3 cytosol, proved AP-I exists as a 58-kDa peptide in vivo, not as the 53-kDa peptide previously identified as active in nucleosome assembly (Ishimi, Y., et al., Eur. J. Biochem., 142, 431-439, 1984). An immunocytochemical study using the monoclonal antibody with the highest specificity against AP-I pin pointed the intranuclear localization of AP-I in HeLa S3 cells.View full abstractDownload PDF (1205K) -
An Improved Method for Isolation of Mutator Mutants from Mouse FM3A Cells and Their CharacterizationMasao Hyodo, Narushi Ito, Midori Yamashita, Kenshi Suzuki1985 Volume 10 Issue 4 Pages 383-390
Published: 1985
Released on J-STAGE: April 18, 2008
JOURNAL FREE ACCESSAn improved method to select mutator mutants was de-veloped. By this new method, mutator mutants were isolated efficiently, and 7 mutants were obtained from cultured mouse FM3A cells. These mutator mutants have an elevated rate of spontaneous mutation at 3 genetic loci (resistance to ouabain, blasticidin S, and tunicamycin). The sensitivity of these mutants to aphidicolin and arabinofuranosylcytosine was the same as in the wild-type cells. Determination of the size of the cellular dNTP pool revealed that there was no large imbalance in the precursor pool in the mutator mutants. These results suggested that the mutator character may be due to alteration in some factor(s) correlated directly to DNA replication. Also, there was no change in the sensitivity of all these mutator mutants to DNA damaging agents.View full abstractDownload PDF (806K) -
Akihiko Yoshimura1985 Volume 10 Issue 4 Pages 391-404
Published: 1985
Released on J-STAGE: April 18, 2008
JOURNAL FREE ACCESSThe early interaction between KB cells and adenovirus was studied by examining the uptake of an extracellular fluorescent macromolecule, FITC (fluorescein isothiocyanate)-labeled dextran (FD). When cells in sus-pension were incubated with both adenovirus and FD, cell-associated FD increased 2-to 3-fold the value obtained without adenovirus. Under fluores-cence microscopy, cells incubated with adenovirus showed bright, whole-cellular fluorescence; whereas, those incubated without adenovirus, or with heat-inactivated virus, showed weaker fluorescence, mainly of the pinocytic vesicles. The increased uptake of the FD by adenovirus was inhibited by treating KB cells with the drugs chloroquine, ammonium chloride and monensin that raise the pH of the acidic compartment. Entry of adenovirus into the KB cell's nucleus also was inhibited by these drugs. The conclusion is that entry of adenovirus into the cell involves its passage of an acidic compartment (probably the endocytic vesicle) and that co-endocytosed macromolecules are released into the cytosol on entry.View full abstractDownload PDF (1261K) -
Ryoji Ishida, Masahiro Kozaki, Taijo Takahashi1985 Volume 10 Issue 4 Pages 405-409
Published: 1985
Released on J-STAGE: April 18, 2008
JOURNAL FREE ACCESSCaffeine has been shown to enhance the lethal effect of DNA-damaging agents in mammalian cells, and the potentiation by caffeine of this effect is generally interpreted as the result of inhibition by caffeine of the repair of damaged DNA. However, the mechanism by which caffeine enhances the lethal effect of DNA-damaging agents has not yet been elucidated. During studies on the effect of caffeine on DNA repair, we found by alkaline elution analysis that caffeine alone produced DNA strand breaks or alkali labile sites in Chinese hamster ovary cells. The amount of DNA breakage or alkali labile sites depended on the concentration of caffeine. We propose that DNA breakage induced by caffeine may be involved in the enhancement of the lethal effect of DNA-damaging agents.View full abstractDownload PDF (468K) -
Yukiko Kunugi-Uehara, Junji Miyakoshi, Wasako Oda, Chiyoko Inagaki1985 Volume 10 Issue 4 Pages 411-416
Published: 1985
Released on J-STAGE: April 18, 2008
JOURNAL FREE ACCESSChanges in microsomal Na+, K+-, Mg2+- and Ca2+-ATPase activities during cell proliferation were examined in Chinese hamster V-79 (V-79) cells (normal cells) and human HeLaS-3 (HeLaS-3) cells (malignant cells). For V-79 cells, the Mg2+-ATPase activity per cell (pmol Pi/h/cell) in the confluent phase was higher than that in the logarithmically growing (log) phase. The amount of microsomal protein per cell was also high in the confluent phase. Specific activities (μmol Pi/h/mg protein) of Na+, K+-, Mg2+- and Ca2+-ATPases were significantly lower in the confluent phase than in the log phase. For HeLaS-3 cells, an increase in Ca2+-ATPase activity per cell was observed. The amount of microsomal protein per cell did not change between the log and confluent phase. The specific activity of Ca2+-ATPase in the confluent phase was also markedly higher than in the log phase. The relation between changes in ATPase activities and cell proliferation is discussed.View full abstractDownload PDF (630K) -
Ryoji Ishida, Taijo Takahashi, Takeharu Nishimoto1985 Volume 10 Issue 4 Pages 417-420
Published: 1985
Released on J-STAGE: April 18, 2008
JOURNAL FREE ACCESSThe tsBN2 cell line, a temperature sensitive (ts) mutant of the BHK21/13 cell line (Nishimoto, T. et al. Somatic Cell Genet. 4, 323-340, 1978) has a ts defect in its regulatory mechanism for the initiation of chromosome condensation, the so-called, premature chromosome condensation (PCC) being induced at a nonpermissive temperature (Nishimoto, T. et al. Cell 15, 475-483, 1978, Nishimoto, T. et al. J. Cell Physiol. 109, 299-308, 1981). Using the 'tsBN2' mutation, we analyzed chromosomes of tsBN2 cells arrested in the G2 phase with neocarzinostatin (NCS) and found considerable aber-rations, such as gaps, breaks and double minutes-like fragmentation, in addi-tion to a typical G2-chromosome (a long filamental chromosome made up of two chromatids). Our results provide evidences that the tsBN2 cell line can be used for examinations of the chromosomes of cells arrested in the G2 phase.View full abstractDownload PDF (438K) -
Chikara Sato, Shosuke Ito, Takuji Takeuchi1985 Volume 10 Issue 4 Pages 421-425
Published: 1985
Released on J-STAGE: April 18, 2008
JOURNAL FREE ACCESSWe isolated cultured melanocyte population from dorsal skin of C57BL/6J (genotype, a/a; C/C) new-born mouse and established several cell lines. One of the melanocyte clones, TM 10, produces both eumelanin and pheomelanin. This cell line seems to be suitable for study of the regulation mechanism of production of these pigments in vitro.View full abstractDownload PDF (517K)
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