摘要:

AbstractComparison of ≏ 160 α‐, β‐, and γ‐tubulins, and excluding the highly divergent C‐terminal peptide, indicates that the three subclasses have similar tertiary structures. Conserved sequences within or between the subclasses have been identified, together with the locations of known epitopes, chemical modifications, and mutations. Evidence is also reviewed concerning the identity of the GTP‐binding sites, about which residues are exposed in the assembled microtubule and at subunit:subunit interfaces. These characteristics constrain the possible tertiary structure of the t

ISSN:0886-1544    

DOI:10.1002/cm.970200302

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1991

数据来源: WILEY

摘要:

AbstractTo investigate the function of vinculin in blood platelets, we studied its localization in relation to other cytoskeletal proteins as well as its state of phosphorylation in platelets allowed to spread on fibrinogen‐coated surfaces. By 5 minutes after loading the platelets onto the surfaces the 47 and 20 kDa polypeptides became phosphorylated, indicating activation. By 30 minutes, platelets formed small, typical bundles of fibers which stained brilliantly with rhodamine phalloidin. Myosin and tropomyosin, detected with specific antibodies, were localized in periodic arrays along these bundles. By indirect immunofluorescence, a discrete patch of vinculin was observed at each end of every actin‐containing bundle. Vinculin phosphorylation was not detected in immunoprecipitates protected against phosphatases. Interference reflection images showed that regions of close binding to the substratum (adhesion plaques) closely matched the vinculin staining sites. Talin appeared diffusely localized. It could be shown to be present in the plaques when platelets were stabilized with ZnCl2by the method of Geiger and then sonicated to remove some of the surface membrane. Localizations of vinculin and myosin were unaltered by this treatment. Talin phosphorylation or proteolysis could not account for vinculin translocation.We conclude that platelets, in response to an appropriate physiological surface, form typical actin bundles with vinculin at the termination of each bundle, in close relation to adhesion plaques. The signal for this translocation does not appear to depend on phosphorylation of vinculin or on phosphorylation or proteolysis of talin. Our findings support the conclusion that in platelets, as in nucleated cells, vinculin serves as at least part of the connection between bundled actin fibers and the extracellular matrix. Such a connection seems required for platelets' known ability to exert tension on surfa

ISSN:0886-1544    

DOI:10.1002/cm.970200303

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1991

数据来源: WILEY

摘要:

AbstractWe have used an in vitro model system to analyze cytomechanical aspects of tissue infiltration by T‐lymphocytes. The interaction of metastatic T‐lymphoma cells with a precultured monolayer of 10T½ fibroblast‐like cells was recorded in time‐lapse video with alternating phase contrast and reflection interference contrast microscopy. Sectioning of embedded specimens as well as cytoskeletal stainings have been performed on matching cocultures.The lymphoma cells did not strongly attach or spread on the dorsal surface of the monolayer cells. Invasion started with the protrusion of a pseudopodium through a narrow gap, and conspicious constriction of the invading cell's body and nucleus was a consistent feature during the later steps. Overt retraction of the target cells was not seen, but the invading lymphoma cells elevated the fibroblasts over relatively large areas, thereby creating dome‐shaped open spaces, allowing for further migration under the monolayer with minimal resistance. Invasion was not unidirectional but was readily reversible at any stage. Due to this wavering character, an invasion event could take more than 1 hour, although the shape alterations involved were fast. Even after the invasion process had been completed, the lymphoma cells could come out from below the monolayer again. Therefore we propose that invasion in this model should be considered as a dynamic equilibrium.Invading T‐lymphoma cells displayed diffuse F‐actin staining and a well‐organized microtubular complex with the centrosomes behind the nucleus in the uropod, which also contained most vesi

ISSN:0886-1544    

DOI:10.1002/cm.970200304

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1991

数据来源: WILEY

摘要:

AbstractWe have established an in vitro migration system for nematocytes of the fresh water cnidarianHydra. Nematocytes display a migratory behavior on isolated sheets of the naturally occurring extracellular matrix, the mesoglea, as well as on surfaces coated with collagen type IV or laminin. Cell behavior was analyzed using video microscopic techniques. Average migration speeds of nematocytes on the mesoglea (140 μm/hr) were lower than values reported from in vivo studies (500 μm/hr). Cells on collagen IV moved at about the same average speed (115 μm/hr) as nematocytes on the natural extracellular matrix; those on laminin were considerably slower (20 μm/hr). Attachment but no movement of cells was found on glass or on surfaces coated with collagen type I and fibronectin. In addition to the differential migration speeds, nematocytes displayed distinct morphologies depending on the substratum. In order to elucidate the causes of the observed cell shape and behavior modulations induced by the offered substratum, the arrangement of major cytoskeletal proteins inHydranematocytes during the in vitro migration or attachment was investigated. The pattern of F‐actin, myosin, and tubulin was determined by immunocytochemical techniques and confocal laser scanning microscopy in nematocytes moving on the mesoglea, on collagen IV, and on laminin, or in cells attaching to fibronectin. We found that the distribution of the cytoskeletal proteins was strikingly different in moving and in stationary cells. The patterns of cytoskeletal proteins in all nematocytes moving on the different substrata, however, was quite si

ISSN:0886-1544    

DOI:10.1002/cm.970200305

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1991

数据来源: WILEY

摘要:

AbstractThe cytoskeleton of the amoeboid spermatozoa ofAscaris suumconsists of major sperm protein (MSP) filaments arranged into long, branched fiber complexes that span the length of the pseudopod and treadmill rearward continuously due to assembly and disassembly at opposite ends of the complexes (Sepsenwol et al.,Journal of Cell Biology108:55–66, (1989)). Examination by video‐enhanced microscopy showed that this cytoskeletal flow is tightly coupled to sperm locomotion. The fiber complexes treadmilled reaward at the same rate (10–50 μm/ min) as the cell crawled forward. Only fiber complexes with their plasmalemmal ends within a limited sector along the leading edge of the pseudopod underwent continuous assembly. Thus, the location of this sector, which occupies about 50% of the pseudopod perimeter, determined the direction of sperm locomotion. Treatment of sperm with agents that lower intracellular pH, such as, weak acids and protonophores, caused the fiber complexes to disassemble completely in 4–5 sec. Removal of these compounds resulted in reassembly of the cytoskeleton in a pattern that mimicked treadmilling in intact sperm. The fiber complexes were reconstructed by assembly at their plasmalemmal ends so that within 30–60 sec the entire filament system reformed and the cell resumed locomotion. Both cytoskeletal reassembly and treadmilling required exogenous HCO3−. These results suggest that variation in intracellular pH may help regulate cytoskeletal treadmilling and thereby play a significant role in sper

ISSN:0886-1544    

DOI:10.1002/cm.970200306

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1991

数据来源: WILEY

摘要:

AbstractThe fragment of smooth muscle α‐actinin, comprising the four spectrin‐like structural repeating units, has a high α‐helix content, similar to that of spectrin, and a hydrodynamic frictional coefficient, indicative of an elongated, probably bent or kinked rod‐like structure, as found for spectrin dimer and tetramer. The fragment exists in solution as an extremely stable dimer, which is dissociated only under denaturing conditions and is much more resistant to dissociation by urea than is the spectrin heterodimer. High‐resolution proton magnetic resonance spectra reveal that a part of the polypeptide chain gives rise to sharp resonances; this is also true of spectrin and it implies that the individual structural repeating units contain segmentally mobile elements, which may be required to generate the elastic properties of the spectrin family of proteins. Again like spectrin, the α‐actinin fragment contains multiple binding sites for long‐chain fatty acids, as revealed by quenching of tryptophan fluorescence by 2‐bromostearate (though not by 9(10)‐bromostearate). The results point to extensive structural and functional similarities between the repeating units of all the proteins of

ISSN:0886-1544    

DOI:10.1002/cm.970200307

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1991

数据来源: WILEY

摘要:

AbstractAn antiserum against tubulin, NS20, has been previously shown to inhibit anterograde and retrograde axonal transport by 50% in vivo and in vitro. We report here that Protein A purified NS20 antibodies also attenuate sperm motility by 50% in demembranated sea urchin sperm. This inhibition is absorbed out by preincubating the NS20 antibodies with a biochemically purified porcine microtubule preparation, with recombinant Trypanosoma β‐ (but not α‐) tubulin and most specifically, with a 37 amino acid (a.a.) synthetic peptide corresponding to a domain near (but not including) the porcine β‐tubulin C terminus. Furthermore, addition of this β‐tubulin peptide alone is sufficient to attenuate motility by 50% in demembranated sperm, indicating that this critical 37a.a. NS20 antigen is a motor binding domain. Together, the results suggest that at least two phenotypically distinct forms of microtubule‐based motility, axonal transport and flagellar beating, are homologous at the fundamental level of the microtubule domains (the β‐tubulin peptide and we suggest a distinct but similarly located α‐tubulin domain) mediating the attachment of tubuli

ISSN:0886-1544    

DOI:10.1002/cm.970200308

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1991

数据来源: WILEY

ISSN:0886-1544    

DOI:10.1002/cm.970200309

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1991

数据来源: WILEY

ISSN:0886-1544    

DOI:10.1002/cm.970200301

出版商:Wiley Subscription Services, Inc., A Wiley Company    

年代:1991

数据来源: WILEY