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B Geiger

Publications and source records attributed to B Geiger.

At least 127 records · Page 7Linked to original sources
PubMed1981

The association of rhodamine - labelled alpha-actinin with actin bundles in demembranated cells.

Rhodamine-labelled alpha-actinin was specifically bound to actin containing filament bundles of demembranated fibroblasts, and was particularly associated with their termini. Optimal binding of rhodamine-alpha-actinin occurred at pH 6.0 - 6.2 and could be abolished by the addition of unlabeled alpha-actinin or myosin subfragment 1. The spatial relationships between the decorating alpha-actinin and several actin associated proteins was determined by double fluorescence microscopy.

Actinin↗
PubMed1981

Ultrastructure of chicken cardiac muscle as studied by double immunolabeling in electron microscopy.

The ultrastructural localization of alpha-actinin and vinculin in chicken cardiac muscle was studied by double indirect immunoelectron microscopy, using ferritin and iron-dextran (Imposil) as the electron-dense markers conjugated to the secondary antibodies, on ultrathin frozen sections of fixed tissue. Fixation and immunolabeling procedures were developed that permitted maximal retention of the two proteins at their natural sites as well as their adequate labeling. alpha-Actinin was found both on the Z-bands, as expected, and near the fascia adherens of the intercalated discs, whereas vinculin was confined to the latter sites. At the fascia adherens, the double labeling results clearly showed that vinculin was situated closer to the membrane than was alpha-actinin. These results, coupled with earlier observations, suggest that vinculin may participate in the linkage of actin-containing microfilament bundles to membranes in a variety of cell types.

Actinin↗
PubMed1981

Immunoelectron microscope studies of membrane-microfilament interactions: distributions of alpha-actinin, tropomyosin, and vinculin in intestinal epithelial brush border and chicken gizzard smooth muscle cells.

The ultrastructural localization of three cytoskeletal proteins, alpha-actinin, tropomyosin, and vinculin, in the brush border of epithelial cells of chicken small intestine and the smooth muscle cells of chicken gizzard was studied by immunofluorescence and immunonelectron microscope labeling of frozen sections of lightly fixed, intact tissues. In the immunoelectron microscope studies, a recently described new type of electron-dense antibody conjugate, imposil-antibody, has been successfully used, along with ferritin-antibody conjugates, in single and double immunolabeling experiments. In the intestinal brush border shows that vinvulin is sharply confined to the junctional complex close to the membrane region of the zonula adherens, in distinct contrast to the more diffuse distributions of the other two proteins. In the smooth muscle cells, the labeling patterns show that vinculin is sharply confined to the membrane-associated dense plaques, closer to the membrane than the alpha-Actinin is also present in the cytoplastic dense bodies, from which vinculin is absent. Tropomyosin is present diffusely distributed in the cytoplasm, but absent from both dense plaques and dense bodies. These findings with the muscle cells demonstrate, therefore, that the dense plaques and dense bodies are chemically and structurally distinct entities. The results with both tissues, along with those in previous papers (Geiger, 1979, Cell. 18:193-205.; Geiger et al., 1980, Proc. Natl. Acad. Sci. U. S. A. 77:4127-4131), suggest that vinculin may play an important and widespread role in the linkage of actin-containing microfilament bundles to membranes.

Actinin↗
PubMed1980

The localization of acetylcholine receptor clusters in areas of cell-substrate contact in cultures of rat myotubes.

We have used interference reflection and fluorescence microscopy to investigate the relationship between cell-substrate contact and the location of clusters of acetylcholine receptors (AChRs) in cultures of rat myotubes. We have found that AChR clusters on the ventral myotubes surfaces are always located within broad regions of close cell-substrate contact. Detailed analysis of the fine structure of the AChR cluster and its associated contact region showed that AChRs within a cluster are concentrated between the points of closest cell-substrate apposition. Vinculin, a recently discovered intracellular smooth muscle protein, is also concentrated in broad regions of close contact, interdigitating with AChRs within the clusters.

Animals↗
PubMed1980

Vinculin, an intracellular protein localized at specialized sites where microfilament bundles terminate at cell membranes.

As intracellular protein of 130,000 molecular weight was recently isolated in this laboratory from chicken gizzard smooth muscle. By immunofluorescence observations of cultured chicken fibroblasts, it was shown to be concentrated on the ventral surfaces of the cells where they formed focal adhesions to the substratum [Geiger, B. (1979) Cell 18, 193-205]. Focal adhesions are sites where, inside the fibroblast, microfilament bundles are known to terminate at the cell membrane. The suggestion was made that this new protein (herein named "vinculin") might be involved in the linkage of the termini of microfilament bundles to membranes in various cell types. To explore this possibility, in the present study we examined several chicken tissues, including intestinal epithelium, gizzard smooth muscle, and cardiac striated muscle, by immunoelectron microscopic labeling for vinculin on ultrathin frozen sections of the specimens. In each case, the immunolabeling for vinculin was concentrated close to membrane sites where microfilament bundles terminate: at the zonula adherens in the junctional complex of the brush border of epithelial cells; at the membrane-associated sense plaques of smooth cells; and at the fascia adherens of the intercalated disk membranes of cardiac muscle cells. These results suggest therefore that vinculin may participate in the anchoring of microfilament bundles to specific membrane sites in various cells.

Animals↗
PubMed1980

Association of microtubules and intermediate filaments in chicken gizzard cells as detected by double immunofluorescence.

By double indirect immunofluorescence, using guinea pig and rabbit antibodies to tubulin and to desmin, we have simultaneously labeled microtubules and intermediate filaments in cultured chicken embryo gizzard cells. At the resolution of the light microscope there was extensive but not complete superposition of the labeling patterns for the two filamentous structures within cells in interphase and an essentially complete dissociation of the two labeling patterns in cells in mitosis. These results indicate that there is an extensive association of microtubules and intermediate filaments in these interphase muscle cells and suggest that this association is regulated metabolically.

Animals↗
PubMed1979

Carbohydrate composition of human placental N-acetylhexosaminidase A and B.

The carbohydrate composition of N-acetyl-beta-D-hexosaminidases (EC 3.2.1.52) A, B and heat-converted B was determined by g.l.c. Similar quantities of mannose, N-acetyl-glucosamine and galactose are present in the A and B isoenzymes, whereas N-acetyl-neuraminic acid is found in significant amount in only the A isoenzyme. The heat-converted hexosaminidase B also contains only trace amounts of N-acetylneuraminic acid, but is about 1.5-fold richer in mannose and N-acetylglucosamine and nearly 2-fold richer in galactose than native hexosaminidase B. Since native and converted hexosaminidase B are thought to be composed of four identical protein chains, our results suggest that there may be variable glycosylation of these chains.

Carbohydrates↗
PubMed1979

The participation of alpha-actinin in the capping of cell membrane components.

By means of double fluorescence staining experiments, intracellular alpha-actinin was found to accumulate under caps and patches induced in several cells by a variety of ligands. This phenomenon was demonstrated in lymphocytes and lymphoma cells treated with anti-H-2 sera; spleen lymphocytes treated with concanavalin A or anti-immunoglobulin antibodies, and VSV-infected mouse fibroblast line MC57 treated with antiserum against viral antigens. It occurred during both rapid and slow capping processes, and could be obtained by either direct or indirect ligand-induced redistribution. These observations were carried out on whole cells. For other cytoskeletal proteins such as filamin, tropomyosin and myosin, a similar accumulation under caps was not readily apparent using whole cell mounts, although earlier experiments with frozen-sectioned cells had shown such an enrichment of myosin (as well as actin). The enrichment of alpha-actinin under the clustered surface molecules was already apparent in early stages (patching) of the capping process, with or without 10 mM sodium azide present. Prolonged incubation of the cells with the different ligands resulted in endocytosis of the ligand-receptor complex. alpha-Actinin was not associated with the inernalized complex, however, suggesting that it may dissociate from the patched or capped surface structures at some stage during endocytosis.

Actinin↗
PubMed1979

Immunocytochemical localization of alpha-actinin in intestinal epithelial cells.

alpha-Actinin was localized in chicken intestinal epithelial cells by immunofluorescence and immunoferritin labeling of thin frozen sections. Most of the label of the brush border was confined to the terminal web area. The label there was concentrated mainly along the "roots" of the microvilli core microfilaments and in the vicinity of the zonula adherens. In the latter structure, the narrow electron-dense zones adjacent to the cell membranes, however, were not significantly labeled. This suggests that alpha-actinin does not mediate directly the association of the transverse terminal web microfilaments to the membrane at the zonula adherens. Sparse ferritin labeling was found near the tight junction, whereas the staining associated with the spot desmosome was negligible. The microvilli were not significantly labeled by either immunofluorescence or immunoferritin staining unless the sections were previously treated with detergent. Moreover, alpha-actinin (or a structurally related protein) was not detected in preparations of purified microvillar vesicles, suggesting the possibility that the alpha-actinin staining in the microvilli may be an artificial due to its translocation by the detergent from the terminal web onto the microvilli. The possible roles of alpha-actinin in the organization and function of the brush border are discussed.

Actinin↗
PubMed1979

Selective and unidirectional membrane redistribution of an H-2 antigen with an antibody-clustered viral antigen: relationship to mechanisms of cytotoxic T-cell interactions.

We have studied the co-redistribution of vesicular stomatitis virus (VSV) antigen and of individual H-2 antigens on the surfaces of mouse cells, and in parallel we have also used these VSV-infected cells as targets in cytotoxic T-cell killing experiments. Antibody-induced patching and capping of the VSV antigen caused an extensive co-patching and co-capping of the H-2Kb antigen but not of the H-2Db antigen. In reciprocal experiments, the antibody-induced patching of the H-2Kb or H-2Db antigen did not result in a co-patching of the VSV antigen. Radioimmunoassays showed that the relative numbers of H-2Kb, H-2Db, and VSV antigens on the surfaces of the cells exhibiting such nonreciprocal co-redistributions were closely similar. Furthermore, the H-2 restricted cytotoxic T-cell lysis of these target cells showed a marked preference for H-2Kb compared to H-2Db compatibility. We propose that the VSV and H-2 antigens are molecularly independent entities in the unpreturbed target cell membrane but that the antibody-induced clustering of the VSV antigen causes a selective and unidirectional co-redistribution (which we designate as syn-capping) of H-2Kb with the VSV antigen clusters. It is suggested that such a T-cell-induced syn-capping process involving an antigen and an H-2 molecule on the target cell may play a critical role in the mechanism of cytotoxic T-cell killing.

Antibodies, Viral↗
PubMed1979

Enzyme replacement in Tay-Sachs disease.

Enzyme replacement therapy was attempted with two Tay-Sachs-diseased individuals--a 14-month-old child and a 7-week-old infant. Treatment consisted of repeated weekly intrathecal injections of pure hexosaminidase A. Injection of this enzyme resulted in almost complete disappearance of GM2 from the serum, but did not bring about dissolution of the GM2 membranous cytoplasmic bodies in the brain, as detected by electronmicroscopy. Both patients tolerated the treatment without apparent clinical complications, but no clear-cut improvement was noted as a result of prolonged injections of hexosaminidase A. Since this treatment was initiated in both an advanced stage and a very early stage of the disease, we conclude that enzyme replacement treatment by this route is not beneficial for patients with Tay-Sachs disease.

Biopsy↗
PubMed1979

The use of antibody-coated liposomes as a target cell model for antibody-dependent cell-mediated cytotoxicity.

A simple model system was developed for antibody-dependent cell-mediated cytotoxicity (ADCC) using antibody-coated synthetic membranes (liposomes) as a target cell model. A synthetic hapten, dinitrophenyl-phosphatidylethanolamine (DNP-PE), was incorporated into fluorophore-quencher-loaded liposomes and the latter were coated with pure anti-DNP antibodies. Normal spleen lymphocytes were capable of binding and subsequently lysing these liposomes. This process is dependent upon the presence of an intact (Fc-containing) IgG molecule and independent of exogenous serum complement. The effector lymphocytes are nylon non-adherent, devoid of Thy-1 antigen and present in nude mice, suggesting an identity with K (Fc receptor positive) lymphocytes. These studies indicate that liposomes may be used as a model to study the requirements for the binding and lysis of target cells in this cell-mediated cytotoxic system.

Animals↗
PubMed1979

Normal adult with absent HEX A: immunoreactive HEX A is present.

Fibroblasts from a normal adult with absent hexosaminidase A (HEX A) activity were demonstrated to possess immunoreactive HEX A as measured by GM2 beta-D-N-acetylgalactosaminidase activity which precipitated with specific anti-HEX A antibodies. Possible explanations for the molecular defect are presented.

Antibodies↗