摘要:

AbstractMicrotubules assembled from sea urchin eggs with the use of taxol contain a 77,000‐dalton protein as the major nontubulin component [Vallee and Bloom (1983): Proc Natl. Acad. Sci. U.S.A. 80:6259–6263]. We have raised five monoclonal antibodies to this protein to aid in its characterization. Immunoblot analysis of the sea urchin microtubule purification fractions indicated that the protein copurified quantitatively with microtubules. All five antibodies stained the mitotic spindle of dividing sea urchin eggs by immunofluorescence microscopy, indicating that the protein was a component of the mitotic spindle and suggesting that it was actually localized on microtubules in vivo. Immunofluorescent staining of higher resolution was observed in a subpopulation of the coelomic cells found in adult sea urchins, confirming that the 77,000‐dalton protein is indeed present on microtubules in vivo. Because taxol was not used for the immunofluorescence experiments, we conclude that the microtubule‐associated protein (MAP)‐like behavior of the 77,000‐dalton protein in vitro was not induced artifactually by taxol. To determine whether this protein is a component of sea urchin microtubules in general, cilia obtained from blastula stage embryos and sperm tail flagella were analyzed with the antibodies. The protein was undetectable by both immunoblot analysis and immunofluorescence microscopy in both preparations of axonemal microtubules. These results indicated that the 77,000‐dalton MAP is restricted to cytoplasmic and mitotic microtubules in the sea urchin. Furthermore, in view of its particular abundance in embryos, whose microtubules are devoted substantially to mitosis, the 77,000‐dalton MAP is likely to play an important role in regulating the activity of mitotic spindle microtubules in

ISSN:0271-6585    

DOI:10.1002/cm.970050602

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

年代:1985

数据来源: WILEY

摘要:

AbstractEvidence is presented to show that klinokinesis, which was previously demonstrated in bacteria and amoeba only, may also occur in metazoan cells. The chemotactic peptide formyl‐Met‐Leu‐Phe (fMLP) elicited orthokinetic and klinokinetic responses of human blood‐borne polymorphonuclear leucocytes (PMNs) under the test conditions used. Increased speed (orthokinesis) was due to an increase in the proportion of migrating cells as well as in the speed of the locomoting subset. The klinokinetic effect was manifested by a decrease in the klinolocomotion index, the mean angle of changes in direction ⩾ 90°, and the frequency of turns ⩾ 90°. The klinolocomotion index was inversely related to speed. This explains the synergistic effect of klinokinesis and orthokinesis in this system. Colchicine alone had and orthokinetic effect which was exclusively due to alterations in the proportion of migrating cells and it altered the turning behaviour without exerting a klinokinetic effect. However, colchicine had marginal orthokinetic and klinokinetic effects on fMLP‐stimulated cells resulting in reduced translocation. The relationship between klinokinesis and mean angle or frequency of turns has been analysed. Klinokinesis was a substantial though not the major element of the chemokinetic response to fMLP under the conditions used. No other metazoan cells have been shown to possess such a complete pattern of responses, including orthokinesis, klinokinesis, and chemotaxis, which regu

ISSN:0271-6585    

DOI:10.1002/cm.970050603

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

年代:1985

数据来源: WILEY

摘要:

AbstractThe production of both lamellipodia and peculiar thin protuberances (scleropodia) characterizes the preaggregative motility of cells after dissociation of the sponge Clathrina.The locomotory paths taken by cells before aggregation were recorded by time‐lapse microcinematography. Changes of direction in successive 50‐s time intervals and 50‐s mean velocities of each cell were both taken into account as statistical variables. Their distributions give probability density curves that seem to fit bilateral exponential functions. The analysis of the angles of turn indicates a tendency for the cells to persist in their direction of motion and to make counter‐clockwise turns. Implications of such in vitro cellular behaviors in aggregative and in vivo processes are su

ISSN:0271-6585    

DOI:10.1002/cm.970050604

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

年代:1985

数据来源: WILEY

摘要:

AbstractA novel method of digital image analysis of the bends of eukaryotic flagella and cilia was devised. In the analysis system, all image pixels were systematically extracted and processed to measure angular direction and curvature. Simulation experiments on theoretical model pictures of flagella with sine‐generated or arcstraight line bending waves demonstrated that the method can be used with considerable high accuracy. This method then revealed abrupt changes in slope of the curvature in sperm flagella and embryo cilia of the sea urchin, Hemicentrotus pulcherrimus. This indicates that the digital image processing used may be helpful in the study of flagellar and ciliary movement

ISSN:0271-6585    

DOI:10.1002/cm.970050605

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

年代:1985

数据来源: WILEY

摘要:

AbstractA discrete stage in two different morphogenetic processes has been examined employing fluorescently labelled alpha‐actinin as a probe to localize native alpha‐actinin and antibodies to localize fibronectin and collagen type I. The stage of somitogenesis examined is the transition from the compact mesenchymal somitic mass to the epithelial somitic vesicle (ie, epithelialization of the somite). The stage of neurulation examined is the transition from the relatively flat neuroepithelium to the approximation of the neural folds. Before these morphogenetic movements begin, the neuroepithelium is sitting upon a basal lamina and interstitial collagen, and the somite is surrounded by a meshwork of interstitial collagen. During both of these processes, the cells become narrowed at their apices in the region of the tissue that is becoming concave, and alpha‐actinin is localized in the apices. The localization of intracellular alpha‐actinin and extracellular fibronectin, and the distribution of collagen, suggest that there is a coordinated appearance and distribution of these molecules that is temporally associated with these discrete morphogenetic

ISSN:0271-6585    

DOI:10.1002/cm.970050606

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

年代:1985

数据来源: WILEY

摘要:

AbstractA model for fast axonal transport is developed in which the essential features are that organelles may interact with mechanochemical cross‐bridges that in turn interact with microtubules, forming an organelle‐engine‐microtubule complex which is transported along the microtubules. Computer analysis of the equations derived to describe such a system show that most of the experimental observations on fast axonal transport can be simulated by the model, indicating that the model is useful for the interpretation and design of experiments aimed at clarifying the mechanism of fast axonal tran

ISSN:0271-6585    

DOI:10.1002/cm.970050607

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

年代:1985

数据来源: WILEY

摘要:

AbstractWe have isolated a 30,000‐dalton protein from Dictyostelium which cosedimented with and affected the low shear viscosity of actin. At low concentrations, this protein increased the low shear viscosity to greater than that of the actin control, whereas higher concentrations decreased viscosity. The viscosity decrease correlated with the formation of actin filament bundles, as seen electron microscopically. This protein resembled a previously reported actin‐binding protein from Dictyostelium [Fechheimer and Taylor, 84, J Biol Chem 259:4514] in electrophoretic mobility, Stokes radius, and ability to crosslink filaments, but was shown to be different by peptide mapping, lack of immunologic crossreactivity, and lack of sensitivity to calc

ISSN:0271-6585    

DOI:10.1002/cm.970050608

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

年代:1985

数据来源: WILEY

摘要:

AbstractThe studies presented here characterize a simple, quantitative NBDphallacidin extraction assay for determining the F‐actin content of fMLP‐activated neutrophils. The NBDphallacidin extraction assay is based upon the specificity of NBDphallacidin binding to F‐actin and the solubility of NBDphallacidin in methanol. Cells are fixed, permeabilized, and stained with NBDphallacidin; the cells are then pelleted, the bound NBDphallacidin is extracted into methanol, and the RFI (excite 465; emit 535) of the solution is determined. Binding of NBDphallacidin to neutrophils is saturable and 90% of bound NBDphallacidin is displaced by nonfluorescent phalloidin. The extraction of bound NBDphallacidin into methanol is complete and the excitation/emission characteristics of NBDphallacidin are not altered by extraction. The assay is relatively inexpensive, applicable to the study of cells in suspension or on substratum, allows kinetic studies with 5–10s time resolution, and is not affected by the shape of the cell or the distribution of the probe. We used the NBDphallacidin extraction assay to study the kinetics of fMLP‐induced change in the F‐actin content of neutrophils and the effect of tBOC peptide, an inhibitor of fMLP binding, on these changes. The extraction assay reveals a rapid, sequential fMLP‐induced increase followed by a decrease in F‐actin content. The tBOC peptide inhibits fMLP‐induced actin polymerization. Addition of tBOC during fMLP‐induced polymerization or at times when F‐actin content is maximal enhances F‐actin depolymerization. The rate of F‐actin depolymerization is ⩾ fourfold faster in the presence than in the absence of tBOC. The results show that (1) The NBDphallacidin extraction assay is useful for studying the kinetics of change in F‐actin content of nonmuscle cells; (2) fMLP receptor occupancy is required for fMLP‐dependent polymerization but not depolymerization; and (3) both the actin polymerizing and depolymerizing processes are active in the cell within 5 s after fMLP stimulation. Implications of these observations for understanding the observed increase and, then, decrease in F‐actin content of f

ISSN:0271-6585    

DOI:10.1002/cm.970050609

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

年代:1985

数据来源: WILEY

ISSN:0271-6585    

DOI:10.1002/cm.970050601

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

年代:1985

数据来源: WILEY