Search PubMed⌕ Search

Biomedical subjects

B M Jockusch

Publications and source records attributed to B M Jockusch.

At least 109 records · Page 6Linked to original sources
PubMed1981

Early changes in the distribution and organization of microfilament proteins during cell transformation.

During the onset of transformation, Rous sarcoma virus-infected cells undergo characteristic morphological changes that reflect the biochemical events induced by the viral src gene. Temperature downshift experiments using chick embryo cells infected with transformation-defective temperature-sensitive viral mutants have shown two major morphological changes occurring at different times in the transformation process: ruffle-like flowers appear on the dorsal cell surface as early as 15 min after temperature shift, while later, between 6 and 12 hr, cytoskeletal stress fibers disappear and the cells round up. We report that flowers contain large accumulations of the cytoskeletal proteins actin, alpha-actinin, myosin and tropomyosin. Furthermore, since flowers stain very intensely with fluorescein-labeled phalloidin, a cyclopeptide that selectively binds to F-actin and not to G-actin, we suggest that these structures result from an early reorganization of microfilaments.

Animals↗
PubMed1981

Interaction of alpha-actinin and vinculin with actin: opposite effects on filament network formation.

The interaction of actin filaments with two actin-associated proteins, alpha-actinin and vinculin (Mr 130,000 protein), was studied in vitro with viscometry and light and electron microscopy. Vinculin, like alpha-actinin, binds to F-actin, and the two proteins were found to have different effects on the formation of filament networks: alpha-actinin crosslinks individual filaments in a manner strongly dependent on temperature and acts as a spacer, whereas vinculin forms actin bundles that display a paracrystalline substructure. In viscometric assays, alpha-actinin mimics the effect of actin gelation factors, whereas vinculin acts as a gelation inhibitor. These findings imply complementary functions of these proteins in the regulation of cellular mobility.

Actinin↗
PubMed1981

The role of clathrin-coated vesicles in acrosome formation.

Ultrastructural and cytochemical observations are presented which suggest that coated vesicles originating from the trans (distal) aspect of the rat spermatid Golgi apparatus transport lysosomal enzymes from the Golgi to the developing acrosome. Using an affinity purified antibody against the clathrin coats of pig brain coated vesicles, immunofluorescence and immuno-electron microscopic studies were carried out on frozen sections of rat seminiferous tubules. The results showed a specific labelling of the Golgi-acrosome region which corresponded precisely with the location of the coated vesicles in Epon sections. We conclude tht a least some of the coated vesicles in this region have clathrin coats.

Acid Phosphatase↗
PubMed1980

Reversible translocation of cytoplasmic actin into the nucleus caused by dimethyl sulfoxide.

The addition of 10% dimethyl sulfoxide (Me2SO) to PtK2 and WI-38 cells caused stress fibers to disappear from the cytoplasm and numerous elongated inclusions to appear in the nucleus. When Me2SO was removed, the stress fibers reformed and the nuclear inclusions disappeared. These nuclear inclusions reacted with fluorescent heavy meromyosin, phalloidin, and actin antibody. In the electron microscope, needle-like structures were seen to be composed of wavy filaments that bound heavy meromyosin. Antibodies against other components of stress fibers--tropomyosin, alpha-actinin, and myosin--did not react with the inclusions. When fluorescently labeled actin was microinjected into living PtK2 and WI-38 cells, the fluorescent actin was incorporated into stress fibers. Subsequent exposure of the same cells to Me2SO led to breakdown of the fluorescent stress fibers and the appearance of fluorescent inclusions in the nucleus. Removal of Me2SO caused reversion to the normal interphase structure. These results indicate that under the influence of Me2SO, dissolution of stress fiber releases actin in a form which allows it to diffuse into the nucleus where it then becomes organized into filamentous bundles.

Actins↗
PubMed1980

Lymphocyte alpha-actinin. Relationship to cell membrane and co-capping with surface receptors.

Mouse spleen lymphocytes synthesize a protein which comigrates with skeletal muscle alpha-actinin on two-dimensional gel electrophoresis and is immunoprecipitated by an antibody directed against skeletal muscle alpha-actinin. Mouse lymphocyte alpha-actinin is present in membrane fractions, and is immunoprecipitated from lymphocyte detergent lysates by an antiserum made against these purified membranes. The anti-alpha-actinin activity of this antiserum is not adsorbed after incubation with fixed intact lymphocytes. Lymphocyte alpha-actinin does not bind concanavalin A and it is inaccessible to lactoperoxidase-catalyzed surface iodination. Double immunofluorescence shows that alpha-actinin moves concurrently along the cell membrane with redistributed surface immunoglobulins and Thy-1 antigen, and remains associated up to 30 min with surface aggregates of these receptors. Our results suggest that lymphocyte alpha-actinin, as defined by molecular weight and cross reactivity with the antibody against the muscle protein, (a) is associated with the cell membrane, (b) is not expressed at the cell surface, and (c) participates in the movement of surface receptors.

Actinin↗
PubMed1980

Antibody labeling of thin sections of skeletal muscle with specific antibodies: a comparison of bovine serum albumin (BSA) embedding and ultracryomicrotomy.

Bovine serum albumin (BSA) embedding and ultracryomicrotomy were used to prepare thin sections of glutaraldehyde-fixed skeletal muscle; these were then treated with antibodies against alpha-actinin, myosin, and actin. Three criteria were then used to compare these two techniques: 1) The preservation of fine structure; 2) the specificity of labeling with antibodies and 3) the amount of antibody bound to a particular antigen. Fine structure was better preserved using ultracryomicrotomy. Both techniques, under optimal conditions, gave specific labeling of muscle components. The amount of antibody bound was higher for BSA sections than for frozen sections. The conclusion is that, while ultracryomicrotomy gives superior qualitative results, the most reliable quantitative estimates would be obtained by using both methods together. Ultracryomicrotomy has the additional advantage that semithin sections can be visualized by immunofluorescence.

Actinin↗
PubMed1979

Nerve fibers in culture and their interactions with non-neural cells visualized by immunofluorescence.

Cultures of embryonic mouse spinal cord explants, alone or in combination with rat myotubes, were stained by indirect immunofluorescence using antibodies against three structural proteins to: (a) reveal the distribution of these proteins among different cell types, and (b) test the usefulness of antibody staining to reveal the gross morphology of the neurite network in complex cultures. Affinity column purified antibodies were used against chicken gizzard actin, porcine brain tubulin, and skeletal muscle alpha-actinin. Neurites were stained intensely by anti-actin as was the stress fiber pattern of underlying fibroblasts. With anti-tubulin, the staining of neurites was an order of magnitude more intense than the staining of the microtubule pattern of background fibroblasts. Neurite cell bodies and astrocyte-like glia cells were stained with anti-tubulin and their nuclei remained unstained. Anti-tubulin could thus be used to trace even the finest extensions of nerve processes in spinal cord and spinal cord-muscle cultures. Furthermore, it could be combined with the histochemical reaction for acetylcholinesterase (AChE, EC 3.1.1.7) to demonstrate AChE-positive neurons and specialized nerve-muscle contact sites. The staining of neural elements with anti-alpha-actinin was generally much weaker than with anti-actin and anti-tubulin. Neurites were stained only moderately in comparison to myotube Z lines in the same culture. However, a distinct staining of the periphery of dorsal root ganglion cells was observed. Thus, a protein immunologically related to muscle alpha-actinin is present in the nervous system. In myotubes, Z lines were stained intensely with anti-alpha-actinin while I bands were only faintly stained with anti-actin. In isolated myofibrils, both structures were stained intensely with the same antibody preparations.

Actinin↗
PubMed1978

An efficient method to produce specific anti-actin.

The production of affinity column purified (specific) anti-actin is described. With the immunization scheme employed, all rabbits produced precipitating antibodies over several months, so that 30 mg specific anti-actin per rabbit could be isolated in 6 months. The antibodies against native and detergent denatured smooth muscle actin are characterized by immunodiffusion tests, staining of the I-band of isolated myofibrils and stress fibers in tissue culture cells, using indirect immunofluorescence.

Actins↗
PubMed1978

Differential expression of Rous Sarcoma virus-specific transformation parameters in enucleated cells.

Chicken embryo fibroblasts transformed with the Ta and ts68 mutants of Rous Sarcoma virus (RSV) were enucleated and studied for their capacity to express reversibly the transformed phenotype in response to temperature changes. After shift to the permissive temperature (35 degrees C), the cytoplasts acquired a transformed morphology and displayed characteristic ruffles and microvilli at their surface. As detected by immunofluorescence, they also lost their actin filament cables and exhibited characteristic changes in the pattern of cell surface structures containing LETS protein. Expression of all these transformation parameters was reversible after shiftback to the nonpermissive temperature (41 degrees C). These results indicate that a whole set of changes characteristic for the transformed phenotype can be expressed independently of the cell nucleus. In contrast, ts mutant-infected cytoplasts were no longer able to respond to temperature shifts with changes in their hexose transport rate. Cytoplasts prepared from cells grown at 41 degrees C retained their low rate of hexose uptake after shift to 35 degrees C, whereas cytoplasts from cells grown at 35 degrees C exhibited a high rate of hexose transport even after 10 hr of shift to 41 degrees C. These results are in accordance with the hypothesis that the product of the src gene of RSV represents a multifunctional protein which acts independently on nuclear and extranuclear sites.

Actins↗
PubMed1977

Synthesis and transport of myosin in Physarum polycephalum.

Immunological techniques have been used to study the rate of synthesis and intracellular transport of myosin in the slime mould Physarum polycephalum. Quantitative precipitation of myosin in homogenates of Physarum was achieved using an antimyosin antibody produced in rabbit in response to purified Physarum myosin. Dodecylsulphate-gel electrophoresis revealed that about 50% of the precipitated material is myosin. The rates of synthesis of total cellular protein and myosin were measured over the mitotic cycle. Both were found to increase exponentially or linearly between two successive nuclear divisions. Similarly, no difference in the proportion of myosin-synthesising polysomes, assayed by precipitation with antimyosin serum, could be detected between the S phase and G2 phase of the mitotic cycle. Myosin makes up nearly 2% of total plasmodial proteins. Its transport into the nucleus occurs predominantly during the G2 phase.

Animals↗