摘要:

AbstractWe have utilized a video‐enhanced contrast system coupled to a DIC‐equipped microscope to examine the motility of both whole coelomocytes and individual filopodia. When the cells are left in diluted coelomic fluid, they exhibit a fibroblast‐like mode of translocation across the substrate. These cells extend lamellipodia at their advancing margin and develop retraction fibers at the trailing edge. Filopodia are actively extended from the lamellipodia of the advancing margin. Cells that are washed free of the coelomic fluid and placed in an isotonic buffer lose their ability to translocate. Filopodia on these stationary cells are seen to undergo a series of waving and bending motions. These motions are rapid and result in a filopodium folding back upon itself only to reextend later. Both forms of motility are discussed in light of the existing structural and biochemical knowledge of this and other cell

ISSN:0271-6585    

DOI:10.1002/cm.970030202

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

年代:1983

数据来源: WILEY

摘要:

AbstractTaxol induces the in vitro assembly of calcium stable microtubules from flagellar tubulin solubilized from sea urchin (Strongylocentrotus purpuratus) sperm tail outer doublets by sonication. Assembly occurs in the presence or absence of exogenous GTP. The drug (10 μM) reduces the critical concentration of protein required for assembly to ⩽0.04 mg/ml.3H‐Taxol binds specifically to both isolated flagellar outer doublets and to reassembled microtubules with calculated maximal binding ratios of 0.25 and 1.32 moles taxol/mole polymerized flagellar tubulin dimer, respectively. We suggest that the discrepancy in maximal binding ratios may result from the presence of an endogenous molecule(s) along the surface of outer doublet microtubules that restricts taxol binding to that struc

ISSN:0271-6585    

DOI:10.1002/cm.970030203

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

年代:1983

数据来源: WILEY

摘要:

AbstractUsing a uniflagellate mutant of Chlamydomonas and flash photomicrography at 300 Hz, we have obtained detailed information on the forward and reverse beating modes of Chlamydomonas flagella and on the relationship between rotation of the uniflagellate cell and the bending cycle of the forward mode. Flagella ranging in length from 5 to 15.5 μm were photographed. There is a decrease in wavelength and an increase in curvature in the principal bends when the length of the flagellum is less than the normal length of 12–13 μm, but these changes are not sufficient to maintain similarity of the bending pattern. In the reverse mode, the flagellum propagates symmetrical, planar, undulatory waves with a shear amplitude which is the same as in the forward mode: there is a 19% increase in beat frequency and a similar decrease in wave length. The reorientation of the flagellar beat direction towards the axis of the cell in the reverse mode is caused both by the decrease in asymmetry of beat and by activation of sliding in the principal bends at an earlier time in the beat cycle, relative to the time of activation of sliding in reverse bends. There are additional rare modes of beating which may be related to intermediate stages in the transition between forward and reverse beating mo

ISSN:0271-6585    

DOI:10.1002/cm.970030204

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

年代:1983

数据来源: WILEY

摘要:

AbstractWe have investigated the Ca2+‐dependent interactions of villin, a protein of the intestinal microvillar core, with actin by monitoring resonance energy tranfer between fluorescently labeled actin subunits. In the presence of elevated free Ca2+(∼20 μM), villin affects both the nucleation and the elongation phases of actin polymerization. Consistent with previous reports, villin stimulates the nucleation process and will form stable nuclei under depolymerization conditions. Compared to the control, the net rate of polymerization is slightly inhibited at low con‐centrations of villin (villin/actin ∼ 1:400) but is stimulated at higher concentrations (villin/actin>1:100). Villin also significantly increases the critical concentration of actin polymerization. Addition of either villin or villin‐actin complexes induces depolymerization of preassembled actin filaments. This villin‐induced depolymerization is reversible upon removal of free Ca2+or upon the addition of phalloidin. The exchange of actin subunits at steady state is inhibited at low concentrations of villin (villin/actin ∼ 1:200) but is stimulated at higher concentrations (villin/actin ∼ 1:50). None of the above effects is observed at<1

ISSN:0271-6585    

DOI:10.1002/cm.970030205

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

年代:1983

数据来源: WILEY

摘要:

AbstractTo test the idea that saltatory organelle movements of nonmuscle cells might be driven by microtubule‐dynein interactions, we microinjected vanadate into several different types of cultured cell. Solutions of sodium metavanadate made up in a simple buffered salt solution were pressure microinjected into fully spread cells in an open‐topped culture chamber placed on the stage of an inverted microscope. The cells were observed by oil‐immersion phase‐contrast optics and results were recorded on movie film. Vanadate, at 10−5‐10−2M, microinjected into cultured chick embryo fibroblasts, failed to inhibit organelle movements. To test the effectiveness of vanadate's inhibitory action under living cell conditions, ciliated epithelial cells were micro‐injected. In these cells even the smallest microinjection of 5 × 10−5M vanadate caused an immediate cessation of ciliary beating. Moreover, in cells that were well spread it was found that whereas vanadate, at 5 × 10−5× 10−3M, inhibited ciliary motion, it failed to inhibit organelle saltations in the same cell. To determine whether vanadate would inhibit a living actin‐myosin system, myocardial cells were also microinjected. Following microinjection of 5 × 10−5and 5 × 10−4M vanadate a temporary tonic contraction (which also occurred following microinjection of buffer alone) was followed by regular beating. Taken together these results demonstrate that in living cell systems microtubule‐dynein interactions are as sensitive to vanadate inhibition as they are in demembranated model systems, and that a working actin‐myosin system in a living muscle cell does not share this great sensitivity. In light of the pronounced differential inhibitory effects of vanadate on the movements of cilia and organelles, our results suggest that saltatory organelle movements in chick embryo fibroblasts and rabbit oviduct epithelial cells are unlikely to be brought abou

ISSN:0271-6585    

DOI:10.1002/cm.970030206

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

年代:1983

数据来源: WILEY

摘要:

AbstractPatients with Kartagener syndrome (KS) show defects in ciliary and flagellar movement that are usually associated with the partial or total absence of dynein side arms from axonemal microtubules. Dynein is essential for such movements, but its involvement in other cellular (particularly microtubule‐related) processes is unknown. It has recently been reported that neutrophils from KS patients show impaired motility including responses to chemotactic stimuli, suggesting that dynein‐like proteins may be generally involved in motile processes. In support of this, we have now found that spontaneous motility of cultured skin fibroblasts from KS patients is also markedly impaired. Three cell lines derived from skin explants of KS patients with deficient dynein side arms in nasal cilia and eight cell lines derived from normal volunteers were studied. Fibroblasts were seeded into dishes containing colloidal gold‐coated cover glasses [Albrecht‐Buehler, 1977], incubated for 24 h at 37°C, and the area of cell “phagokinetic” tracks determined.Each cell line studied in this manner reproducibly displayed an amount of spontaneous motility characteristic for that cell line. The mean track area (± SE) for all control cells studied was 14.6 ± 0.5 × 103μm2whereas for KS fibroblasts was 8.7 ± 0.4 × 103μm2(P<0.001). Immunofluorescence microscopy using antitubulin and antihuman 210 K MAP antibodies revealed no differences in the staining patterns between control and KS fibroblasts. Pinocytic rates were identical, and the complement of tubulin and major microtubule associated proteins as seen on one‐dimensional SDS polyacrylamide gel autoradio‐graphs appeared similar for control and KS cells. Thus, the observed motility defect is probably not the result of alterations in the occurrence or distribution of microtubules or in the occurrence or binding of the major microtubule‐associated proteins. This defect in cellular motility may be related to the absence of dynein or may reflect another in

ISSN:0271-6585    

DOI:10.1002/cm.970030207

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

年代:1983

数据来源: WILEY

摘要:

AbstractDemembranated and membrane disrupted bull sperm models exhibit an increase in motility when exposed to cAMP. Tritium‐labeled cAMP was used to locate the initial site of action of cAMP in the modeled sperm preparations. cAMP did not bind selectively to the modeled cells, and the presence or absence of plasma membrane fragments on the models did not significantly alter this result. When suspension medium taken from modeled sperm preparations was subjected to gel filtration on Sephadex G25‐150 columns, cAMP bound to a high molecular weight component that eluted with the void volume. The responsible binding factor is a soluble component that is released when the plasma membranes of the sperm are disrupted during the modeling procedure. To test the importance of the cAMP binding factor, modeled sperm were centrifuged, the super‐natant solution was decanted, and the cells were resuspended in fresh medium. After this treat‐ment the cells could be restored to motility with Mg‐ATP but no longer exhibited a response to cAMP. Furthermore, addition of cAMP binding factor isolated by gel filtration partially restored the response of these sperm to cAMP. Investigation of the properties of the cAMP‐binding factor have confirmed that it is specific for cAMP, with a much lower affinity for AMP and cGMP. In the pre‐sence of a large excess of unlabeled cAMP the labeled complex has a half‐life of approximately 1 hour. Our results indicate that the action of cAMP on the motility of modeled sperm is mediated by its attachment to a high molecular weight, soluble component of the

ISSN:0271-6585    

DOI:10.1002/cm.970030208

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

年代:1983

数据来源: WILEY

ISSN:0271-6585    

DOI:10.1002/cm.970030209

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

年代:1983

数据来源: WILEY

ISSN:0271-6585    

DOI:10.1002/cm.970030201

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

年代:1983

数据来源: WILEY