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Biomedical subjects

T Enomoto

Publications and source records attributed to T Enomoto.

At least 307 records · Page 17Linked to original sources
PubMed1984

Purification and characterization of a factor stimulating DNA polymerase alpha activity from mouse FM3A cells.

A protein factor which stimulates DNA polymerase alpha activity on heat-denatured DNA has been purified from mouse FM3A cells. The final preparation had a specific activity of 43,000 units/mg protein and lacked detectable DNA polymerase, RNA polymerase, DNA-dependent- and independent ATPase, exo- and endodeoxyribonuclease and phosphatase activities. The stimulating factor sedimented at 2.9S in a glycerol gradient. Sodium dodecyl sulfate polyacrylamide gel electrophoresis of the glycerol gradient fraction revealed the presence of a major band of 36,000 daltons, the amount of which corresponded well with the level of stimulating activity. The stimulation by the factor was specific for heat-denatured DNA, and a little or no stimulation was observed with native DNA, ribo- and deoxyribohomopolymers and single stranded circular DNA. Alkaline sucrose gradient sedimentation analysis of the reaction products revealed that newly synthesized DNA was covalently linked to the termini of heat-denatured DNA. The average chain length of the elongated span determined by the digestion with micrococcal nuclease and phosphodiesterase II, did not differ between in the presence and absence of the stimulating factor, suggesting that the stimulation by the factor was due to the increase in the initiation frequency of DNA synthesis from the 3'-hydroxyl terminus of heat-denatured DNA.

Animals↗
PubMed1984

Effect of the strandedness of cofactor DNA on the activities of DNA-dependent ATPases B and C3 from FM3A cells.

The requirements of cofactor DNA for DNA-dependent ATPases B and C3 were analyzed in detail. ATPase B and C3 required the presence of a polynucleotide for their activities. Among the DNAs tested, ATPase B showed a preference for poly(dT) as its cofactor. The other deoxyhomopolymers, except poly(dG) and heat-denatured DNA also were effective. The alternating polydeoxyribonucleotide, poly[d(A-T)] had an efficiency 23% that of heat-denatured DNA. Unlike ATPase B, ATPase C3 showed almost no activity with deoxyhomopolymers. The most effective cofactor for ATPase C3 so far tested is poly[d(A-T)]. Relatively high activity was obtained with heat-denatured DNA. The high activity of ATPase B with poly(dT) was reduced by the addition of poly(dA). The addition of noncomplementary homopolymers did not affect enzyme activity. ATPase C3 activity in the presence of 10 microM poly(dT) increased gradually with concentrations of poly(dA) up to 20 microM, after which it decreased. Almost no increase in activity was observed when noncomplementary homopolymers were added. The relatively high activity of ATPase C3 with heat-denatured DNA was suggested by its high sensitivity to ethidium bromide to be due to the double-stranded region in the heat-denatured DNA formed by self-annealing.

Adenosine Triphosphatases↗
PubMed1984

Lack of intercellular communication between chemically transformed and surrounding nontransformed BALB/c 3T3 cells.

In order to study the possible role of intercellular communication in the process of in vitro cell transformation, the communicating capacity of BALB/c 3T3 cells transformed by 20-methylcholanthrene was investigated using a dye transfer method. Morphologically transformed foci, detectable 4 to 5 weeks after treatment with 20-methylcholanthrene (1.0 microgram/ml), are clearly distinguishable under the phase-contrast microscope from surrounding nontransformed monolayer cells; therefore, a tracer dye can be injected into individual cells, and gap-junctional communication between and among transformed and nontransformed can be studied directly. When fluorescent Lucifer Yellow CH was microinjected into a cell within the transformed focus, the dye was transferred to other transformed cells but not to cells in the adjacent nontransformed area, although they were in physical contact. Similarly, dye injected into a nontransformed cell was transferred to neighboring nontransformed cells but not to cells in an adjacent transformed focus. These results indicate that when BALB/c 3T3 cells are transformed by 20-methylcholanthrene they can no longer communicate with surrounding normal cells, although they maintain the ability to communicate with other transformed cells in the focus. These results suggest that loss of the ability to communicate with surrounding nontransformed cells may be one of the important determinants of induction and expression of the final malignant transformation.

Animals↗
PubMed1983

Purification and characterization of two forms of DNA polymerase alpha from HeLa cell nuclei.

Isolated nuclei contained two active forms of DNA polymerase alpha (form I and form II). Form II was extracted from nuclei by KCl at concentrations lower than 0.18 M. Above 0.18 M selective extraction of form I was observed. The purified two active forms differed in chromatographic and electrophoretic behaviors, in their salt requirement for optimal activity, and in preference of template-primers, although both forms exhibited properties characteristic of DNA polymerase alpha such as sensitivity of N-ethylmaleimide, 1-beta-D-arabino-furanosylcytosine triphosphate, and aphidicolin. Marked difference between the two forms was preference of template-primer that form I was more active with poly(dT).(rA)10 than poly(dA).(dT)12 whereas form II exhibited higher activity with poly(dA).(dT)12 than poly(dT).(rA)10. Possible roles of two forms of DNA polymerase alpha in the processes of DNA replication will be discussed.

Cell Nucleus↗
PubMed1983

Tumour promoter-mediated reversible inhibition of cell-cell communication (electrical coupling). Relationship with phorbol ester binding and de novo macromolecule synthesis.

A tumour promoter, 12-O-tetradecanoyl phorbol-13-acetate (TPA), reversibly inhibits the onset and maintenance of cell-cell communication measured by electrophysiological method. We have now studied the mechanism by which TPA inhibits communication of human cells (FL) in culture. Using [3H]phorbol-12,13-dibutyrate [( 3H]PDBu), we found a class of specific, high-affinity, saturable binding sites in intact FL cells; they have a dissociation constant of 15.4 nM, and at saturation about 3 X 10(5) PDBu molecules were bound to each cell. The binding of [3H]PDBu to FL cells was inhibited by TPA, phorbol-12-13-didecanoate and mezerein, whereas phorbol and 4 alpha-phorbol-12-13-didecanoate had no effect. There is a close correlation between the ability of the former compounds to inhibit [3H]PDBu binding and their capacity to inhibit cell-cell communication. When FL cells are dispersed with EDTA and plated onto a culture dish, they start to couple electrically within 2 h; such cell coupling was not affected by the presence of cycloheximide or actinomycin D. TPA inhibits the formation of electrical cell coupling as well as its maintenance, even in the presence of cycloheximide; the recovery of cell-cell communication after the removal of TPA was not significantly affected by the addition of cycloheximide or actinomycin D. Taken together, these results suggest that TPA-mediated reversible inhibition of intercellular communication is mediated by specific binding of TPA to cellular receptors and that macromolecular synthesis is not necessary.

Binding Sites↗
PubMed1983

Independent induction of sister-chromatid exchanges by 3-aminobenzamide and ultraviolet radiation in HeLa cells.

The effect of 3-aminobenzamide in HeLa cells was examined in several aspects. 10(-3) M 3-aminobenzamide inhibited poly(ADP-ribose) polymerase activity to more than 95% in disrupted nuclei of HeLa cells. The mode of inhibition of the enzyme was competitive inhibition with NAD at a Ki value of 2.5 microM 3-aminobenzamide. The combined treatment of HeLa cells with 3-aminobenzamide and ultraviolet radiation revealed the independent induction of SCEs by these 2 agents.

Benzamides↗
PubMed1983

Pustulosis palmaris et plantaris. With reference to the cross-reactivity between tonsillar epithelium antigen and skin antigen.

Though it is well known that pustulosis palmaris et plantaris (PPP) is one of skin diseases caused by focal infection of the tonsils, its etiology remains unknown. The purpose of this study was to elucidate the role of the tonsils in the pathogenesis of PPP, by using histological and immunological techniques. Our findings were as follows: The common histological findings in tonsillar lacunae of patients with PPP were pus plugs, pronounced infiltration of polymorphonuclear cells and mononuclear cells, desquamation of keratinized squamous cells, uneven thickness of the epithelium, and inflammation of the subepithelial area. By applying Arthur's method, skin extracts of sole (S) and epithelial extracts of tonsillar lacunae (T) were prepared as antigens for rabbit immunization. Cross-reactivity between skin extract of sole and epithelial extract of tonsils was tested using the Ouchterlony technique. Anti-T antisera reacted to both T and S. Anti-S antisera reacted similarly to T and S. Direct and indirect immunofluorescence studies using anti-T and anti-S antisera revealed positive staining either in the lacunal epithelium of tonsil or in the corneous, granular and papillary layers of sole. As regards the pathogenesis, it is suggested that the antigenic modification of tonsillar epithelium caused by various stimuli produces cells and/or antibodies so as to challenge against skins.

Antibodies↗
PubMed1983

Membrane interaction and modulation of gene expression by tumor promoters.

Phorbol ester tumor promoters bind to specific cellular receptors (probably protein kinase C) and modulate membrane structure and function and gene expression of target cells. Using cell culture systems, we are studying the interaction of phorbol esters with the cellular membrane and subsequent modulation of gene expression. Our recent results can be summarized as follows: 1) specific binding of phorbol esters to mammalian cells can be inhibited by a human placental factor, which we have partially purified and characterized. 2) Phorbol ester tumor promoters reversibly inhibit intercellular communication, as measured by electrical coupling and dye transfer between cultured cells, suggesting that they inhibit both ionic and molecular transfer between cells. 3) In vitro transformation of Balb/c 3T3 cells results in blockage of intercellular communication between normal and transformed cells, indicating that blocked intercellular communication may play a role in cell transformation. 4) 12-O-Tetradecanoylphorbol-13-acetate (TPA) can continuously inhibit differentiation and globin gene expression in Friend erythroleukemia cells, without affecting their growth rate, for about 3 years. Both globin gene expression and terminal differentiation of Friend cells occur again upon removal of TPA from culture medium during such long-term culture.

Animals↗
PubMed1983

Antiteratogenic effects of tumor inhibitors, caffeine, antipain, and retinoic acid in mice.

To learn the effects of tumor inhibitors on chemically induced malformations, caffeine, antipain, and 13-trans-retinoic acid were given to pregnant ICR/Jcl mice after a single dose of urethan, N-hydroxyurethan, N-methyl-N-nitrosourea, N-ethyl-N-nitrosourea, or 4-nitroquinoline 1-oxide, which induces about 50% of the malformed fetuses. When caffeine was given immediately after carcinogen treatment on Day 10, urethan- and N-ethyl-N-nitrosourea-induced malformations were significantly suppressed by caffeine posttreatment, while N-hydroxyurethan- and N-methyl-N-nitrosourea-induced malformations were not suppressed by caffeine. 4-Nitroquinoline 1-oxide-initiated teratogenesis was also suppressed, but not significantly so (p not equal to 0.07). The results were very similar to those of the effects of caffeine on tumors induced by these carcinogens. Malformations of genetic origin (cleft palates and cleft lips) in CL/Fr mice were also suppressed significantly by caffeine treatment on Days 8 to 11, although the level of inhibition was less than that in chemically induced malformations. A protease inhibitor (antipromotor), antipain, also suppressed urethan-induced malformations. The antiteratogenic effects of antipain were most effective when it was given during the period of 24 to 48 hr after urethan treatment, while those of caffeine were most effective when it was given immediately after urethan. The promoting process might be involved in chemically induced teratogenesis, as it was in carcinogenesis. A natural retinoid (13-trans-retinoic acid) also suppressed urethan-induced malformations. Thus, tumors and malformations induced by chemical carcinogens were suppressed by tumor inhibitors, suggesting the similarity of both processes in the subcellular level, in spite of their morphological differences.

4-Nitroquinoline-1-oxide↗
PubMed1983

Estrogen receptor in the "non-lymphocytes" in the thymus of the ovariectomized rat.

Estrogen binding components have been characterized in cytosols from lymphocyte fractions and non-lymphocyte fractions of the thymus and mesenteric lymph node of ovariectomized rats. The cytosol from the non-lymphocyte fraction of the thymus contained high affinity-specific binding estrogen receptor (equilibrium dissociation constant .=. 3 X 10(-10) M, number of binding sites .=. 10 fmol/mg protein) with a sedimentation coefficient of 8 S. The receptor showed specific binding for estradiol-17 beta, estrone and estriol, but not for corticosterone, progesterone or testosterone. Only lower estrogen binding was observed in the cytosols from the lymphocyte fraction of both tissues and the non-lymphocyte fraction of the lymph node. Morphological changes of the thymus in ovariectomized rats after administration of estradiol benzoate were also observed. Clustered vacuoles with incomplete microvilli in the epithelial cells increased in number and in size after estrogen treatment. These findings suggest that the effect of estrogen on the thymus is mediated via the cytoplasmic estrogen receptor in the non-lymphocytes, probably thymic epithelial cells.

Animals↗