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E Wang

Publications and source records attributed to E Wang.

At least 433 records · Page 24Linked to original sources
PubMed1983

Amplified C lambda and c-abl genes are on the same marker chromosome in K562 leukemia cells.

The human leukemia cell line K562, derived from a patient with Philadelphia chromosome-positive chronic myelogenous leukemia, contains amplified c-abl oncogenes and unrearranged C lambda genes. Using in situ hybridization techniques, we have determined that the amplified c-abl and C lambda DNA sequences of K562 cells are both located on the same abnormal acrocentric marker chromosome, which may represent an altered Philadelphia chromosome.

Cell Line↗
PubMed1983

An intermediate filament-associated protein, p50, recognized by monoclonal antibodies.

Monoclonal antibodies (mAB) were raised to be used as probes to identify cytoplasmic components associated with intermediate filaments (IF). Four hybridomas (B27, B76, B78, and B100) secreting mAB were generated by fusing mouse myeloma cells with the spleen cells of mice immunized intraperitoneally with Triton-high salt insoluble materials from BHK-21 cells. This insoluble material consists mostly of IF, a small number of microfilaments, and some polyribosomes. Biochemical studies show that the Triton-insoluble materials contain many proteins, including vimentin (decamin) and desmin. Immunofluorescence microscopy of BHK-21 cells stained with the four mAB showed that these mAB decorate the IF in a dotted pattern. Double staining with polyclonal antibody to vimentin confirmed the reactivity of the mAB with the IF. These mAB also stained the vimentin-containing filament system in a variety of other cells including epithelial cells (PTK1 and HeLa) and cells of astroglial origin. Histological studies showed that mAB-B100 stained many types of tissue including epidermis, smooth muscle, and subdermis pericytes, but not the white matter nor the gray matter of the cerebellum and spinal cord. Immunoelectron microscopy with colloidal gold has shown that the mAB-B100 decorated the IF in clusters or aggregates around proteinaceous materials associated with the filaments. Results of immunoprecipitation indicate that mAB-B100 reacted with a protein of 50,000 daltons. These findings suggest that the mAB-B100 we have developed recognizes one of the many components of what appears to be an integrated cytoskeletal structure connected with intermediate filaments.

Animals↗
PubMed1983

Cytoplasmic localization of the transforming protein of Fujinami sarcoma virus: salt-sensitive association with subcellular components.

Fujinami sarcoma virus (FSV) encodes a transforming protein of 130,000 daltons (P130) which is associated with a tyrosine-specific protein kinase activity. To elucidate mechanisms involved in cell transformation by FSV, we have studied the intracellular location of P130 in rat cells nonproductively infected with FSV. Immunofluorescent staining of several FSV-transformed rat cell lines with a tumor regressor antiserum specific against the fps sequences of P130 showed that the major staining was localized in the cytoplasm. Staining was also seen in cell ruffles and in some cases at areas of cell contact. The cytoplasmic location of P130 staining in cells infected with temperature-sensitive mutants of FSV was unchanged when they were grown at permissive or nonpermissive temperature. Cell fractionation of FSV-transformed cells under various conditions showed that the ionic strength used during cell fractionation had a striking effect on the distribution of P130. At 10 mM NaCl, 70% of P130 sedimented in the large granule fraction, whereas at 500 mM NaCl 70 to 90% of P130 was recovered in the cytosol fraction. Furthermore, a combination of ionic and nonionic detergents that effectively solubilized subcellular membranes was insufficient to solubilize P130 unless the salt concentration was raised. We conclude that the majority of P130 and its associated protein kinase activity are localized in the cytoplasm and that P130 is not an integral membrane protein.

Animals↗
PubMed1983

Reduced synthesis of pp60src and expression of the transformation-related phenotype in interferon-treated Rous sarcoma virus-transformed rat cells.

Treatment of Rous sarcoma virus-transformed rat cells with rat interferon-alpha (specific activity, 10(6) U/mg of protein) for 24 h caused a 50% reduction in intracellular pp60src-associated protein kinase activity. Staphylococcus aureus V8 protease digestion of pp60src, derived from 32P-labeled monolayer cultures incubated with or without interferon, revealed no differences either in the phosphopeptide pattern or in the phosphoserine-phosphotyrosine ratio. However, [3H]leucine pulse-labeling experiments showed that the synthesis of pp60src was reduced by 42 to 48%, relative to the level of bulk protein synthesis, in the interferon-treated cultures. Rat interferon-alpha also reduced the growth rate of Rous sarcoma virus-transformed rat cells in a dose-dependent manner over a 72-h period. The decrease in growth rate was accompanied by increases in the thickness and number of actin fibers per cell and by a decline in intracellular tyrosine phosphorylation by pp60src. The results suggest that interferon can inhibit the expression of the transformation-related phenotype by selectively reducing the synthesis of the Rous sarcoma virus transforming gene product. However, the interferon effects on the cytoskeletal organization and proliferation of Rous sarcoma virus-transformed cells may be due at least in part to the predominance of interferon-induced phenotypic changes over those caused by pp60src.

Actins↗
PubMed1981

Interferon increases the abundance of submembranous microfilaments in HeLa-S3 cells in suspension culture.

Human beta (fibroblast) interferon inhibits the proliferation of human HeLa-S3 carcinoma cells in suspension culture. Accompanying this effect, the lateral mobility of cell surface receptors for concanavalin A is decreased and the rigidity of the plasma membrane lipid bilayer is increased. The present findings show a marked increase in the number of polymerized actin-containing microfilaments 3 days after treatment of HeLa-S3 cells with beta-interferon (640 units/ml). The cortical region of the treated enlarged cells contains a thick and dense meshwork of 40-70 A microfilaments. The actin nature of the filaments was verified by their ability to bind heavy meromyosin. These results support the concept that beta-interferon induces a coordinated response in the plasma membrane and the underlying microfilaments in both tumor and normal cells.

Cell Membrane↗
PubMed1981

Effect of vanadate on intracellular distribution and function of 10-nm filaments.

Earlier reports from this laboratory suggested that 10-nm filaments and microtubules act together in the movement and positioning of nuclei and centrioles. Sodium vanadate has been found to alter the distribution of 10-nm filaments and separate them from microtubules in virus-induced syncytia and uninfected cells. Accompanying this change in cytoskeletal elements in an alteration in the distribution of nuclei, centrioles, and other organelles. Nuclei in vanadate-treated syncytia were found in a circle or horseshoe arrangement, and 10-nm filaments were aggregated within the circle, whereas microtubules, were found in a network throughout the cytoplasm. Vanadate also caused a perinuclear aggregation of 10-nm filaments in single uninfected cells, whereas microtubules were throughout the cytoplasm, as in syncytia. Centrioles, mitochondria, rough endoplasmic reticulum, and lysosomes were scattered in the perinuclear area, with mitochondria and rough endoplasmic reticulum frequently closely associated, whereas the peripheral region of vanadate-treated cells contained ribosomes, microfilament bundles, and microtubules, but not 10-nm filaments. Vanadate limited virus-induced fusion of cells to polykaryocytes with 5--20 nuclei, in contrast to the massive syncytia found in untreated cells. These results indicate that vanadate separates 10-nm filaments and microtubules topologically and functionally, and support previous evidence that 10-nm filaments and microtubules act together in the movement and positioning of nuclei and other organelles.

Animals↗
PubMed1980

Interferon inhibits the redistribution of cell surface components.

Interferon treatment impairs the ability of cells to redistribute cell surface receptors for concanavalin A (Con A). The effect of interferon becomes evident within 3-6 h and is maximal within 36-48 h. Highly purified human fibroblast interferon (> 2 x 10(8) U/mg of protein sp act; concentration; 640 U/ml) caused approximately 85% inhibition of capping of fluorescein-conjugated Con A in interferon-sensitive HeLa-S3 cells at 36 h from the beginning of treatment.

Cell Membrane↗
PubMed1980

Differences in intracellular location of pp60src in rat and chicken cells transformed by Rous sarcoma virus.

We have investigated the intracellular location of pp60src in Rous sarcoma virus-transformed rat cells (RR1022) by indirect immunofluorescence microscopy and cell fractionation. Immunofluorescence data suggest that pp60src is predominantly associated with the nuclear envelope and the juxtanuclear reticular membrane structures. The bulk of pp60src and of the associated phosphotransferase activity fractionated with nuclei and not with plasma membranes in disrupted cells. This localization contrasts strikingly with the association of pp60src with the plasma membrane of Rous sarcoma virus-transformed chicken fibroblasts. We propose that pp60src is a membrane protein that associates with cellular membranes through hydrophobic regions and that this membrane association is a general feature of the interaction of pp60src with avian and mammalian cells. Although there are major differences in the intracellular localizations of pp60src, it may interact with cellular membranes through one or more NH2-terminal hydrophobic regions.

Animals↗
PubMed1980

Interferon effects on microfilament organization, cellular fibronectin distribution, and cell motility in human fibroblasts.

We have shown previously (Pfeffer et al., 1979, Exp. Cell Res. 121:111-120) that treatment of human fibroblasts, planted at a density of 2x10(3) cells/cm(2), with purified human fibroblasts interferon (640 U/ml) for 3 d at 37 degrees C decreases the overall rate of cell proliferation to 35-40 percent of the control value. In the present experiments we have characterized the phenotype of interferon-inhibited fibroblasts. The mean volume of trypsinized, interferon-treated cells was increased 31 percent abover that of control cells. The interferon-treated population was much more heterogeneous than the control population with respect to volume, and there was a considerable overlap in the volume distributions of the two populations. The cell surface area was, on the average, increased 65 percent after interferon treatment. More than 80 percent of the treated cells had enlarged nuclei, many of which were lobed, and the fraction of binucleated cells was increased fivefold. After interferon treatment, over 40 percent of the cells showed large actin-containing fibers in the form of multiple parallel arrays. Fewer than 5 percent of the control cells contained such large actin fibers. The number of actin fibers of all sizes was tripled in the treated fibroblasts on a per cell basis and, calculated per unit surface area of the cells, the number was increased 82 percent. In contrast, 10-nm filaments and microtubules did not appear to be increased in number per unit surface area of the cells. The increases per cell in the abundance of these structures were directly related to increased cell size. After interferon treatment, fibronection was distributed in arrays of long filaments covering most portions of the cell surface. Interferon treatment markedly decreased the rate of cell locomotion as well as membrane ruffling and saltatory movements of intracellular granules.

Cell Line↗
PubMed1980

Phorbol myristate acetate stimulates pinocytosis and membrane spreading in mouse peritoneal macrophages.

Phorbol myristate acetate (PMA) at a concentration of 0.01 microgram/ml causes an approximately threefold increase in surface area of resident, proteose-peptone-elicited, and thioglycolate-broth-elicited mouse peritoneal macrophages. Resident and proteose-peptone-elicited macrophages, cultured for 24 h in the presence of PMA, increase their pinocytic rate twofold in response to addition of PMA (0.01 microgram/ml) to the medium. Thioglycolate-broth-elicited macrophages, cultured for 24 h in the absence of PMA, immediately increase their pinocytic rate 2- to 3.5-fold in response to a single challenge with PMA (0.01 microgram/ml). Cytochalasin B, colchicine, and podophyllotoxin have only modest inhibitory effects on the basal rate of pinocytosis and on PMA-induced cellular spreading, but completely block the stimulatory effects of PMA on pinocytosis in thioglycolate-broth-elicited macrophages. Cytochalasin D markedly inhibits both basal and PMA-stimulated pinocytosis in these cells. Thus, PMA is a useful tool for studying mechanisms of macrophage spreading and for enhancing the overall rate of pinosome formation.

Animals↗
PubMed1980

Phorbol myristate acetate stimulates microtubule and 10-nm filament extension and lysosome redistribution in mouse macrophages.

Phorbol myristate acetate (PMA) stimulates cell spreading and fluid-phase pinocytosis in mouse peritoneal macrophages. Colchicine (10(-5) M) and cytochalasin B (10(-5) M) abolish PMA stimulated pinocytosis but have little effect on cellular spreading (Phaire-Washington et al., 1980, J. Cell Biol., 86:634-640). We report here that PMA also alters the organization of the cytoskeleton and the distrubution of organelles in these cells. Neither control nor PMA-treated macrophages contain actin cables. PMA-treated resident thioglycolate-elicited macrophages exhibit beneath their substrate-adherent membranes many randomly distributed punctate foci that stain brightly for actin. The appearance and distribution of these actin-containing foci are not altered by colchicine (10(-5) M) or cytochalasin B (10(-5) M). In thioglycolate-elicited macrophages PMA causes the extension and radial organization of microtubules and 10-nm filaments and promotes the movement of secondary lysosomes from their perinuclear location to the peripheral cytoplasm. Depending upon the concentration of PMA used, 45-71% of thioglycolate-elicited macrophages and 32-44% of proteose-peptone-elicited macrophages and numerous lysosomes, radiating from the centrosphere region, arranged linearly along microtubule and 10-nm filament bundles. Colchicine (10(-5) M) and podophyllotoxin (10(-5) M) prevent the radial redistribution of microtubules, 10-nm filaments, and lysosomes in these cells. Cytochalasins B and D (10(-5) M) have no inhibitory effects on these processes. These findings indicate that microtubules and 10-nm filaments respond in a coordinated fashion to PMA and to agents that inhibit microtubule function; they suggest that these cytoskeletal elements regulate the movement and distribution of lysosomes in the macrophage cytoplasm.

Acid Phosphatase↗
PubMed1979

Relationship between movement and aggregation of centrioles in syncytia and formation of microtubule bundles.

Previous reports from this laboratory have provided evidence suggesting that microtubules and 10-nm filaments serve both cytoskeletal and force-generating functions in the intracellular movement and positioning of nuclei in syncytia. It has been found that, during the process of cell fusion and nuclear migration in syncytia induced by the paramyxovirus simian virus 5, centrioles are transported in the cytoplasm and form large aggregates. These aggregates are usually found in regions adjacent to rows of aligned nuclei and large bundles of microtubules and 10-nm filaments. Colchicine prevents the translocation and aggregation of centrioles, but cytochalasin B has little effect on this process. These results suggest that the same cytoskeletal elements that are involved in nuclear migration and positioning--i.e., microtubules and 10-nm filaments--are also involved in the transport of centrioles. The possibility that aggregates of centrioles may serve as centers for the organization of microtubules and 10-nm filaments into the large bundles observed in the syncytia is discussed.

Animals↗