摘要:

AbstractWe used video‐fluorescence microscopy to directly observe the sliding movement of single fluorescently labeled actin filaments along myosin fixed on a glass surface. Single actin filaments labeled with phalloidin‐tetramethyl‐rhodamine, which stabilizes the filament structure of actin, could be seen very clearly and continuously for at least 60 min in O2‐free solution, and the sensitivity was high enough to see very short actin filaments less than 40 nm long that contained less than eight dye molecules. The actin filaments were observed to move along double‐headed and, similarly, single‐headed myosin filaments on which the density of the heads varied widely in the presence of ATP, showing that the cooperative interaction between the two heads of the myosin molecule is not essential to produce the sliding movement. The velocity of actin filament independent of filament length (>1 μm) was almost unchanged until the density of myosin heads along the thick filament was decreased from six heads/14.3 nm to 1 head/34 nm. This result suggests that five to ten heads are sufficient to support the maximum sliding velocity of actin filaments (5 μm/s) under unloaded conditions. In order for five to ten myosin heads to achieve the observed maximum velocity, the sliding distance of actin filaments during one ATP cycle must be mo

ISSN:0886-1544    

DOI:10.1002/cm.970100112

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

年代:1988

数据来源: WILEY

摘要:

AbstractAmoebae of the cellular slime moldDictyostelium discoideumare an excellent model system for the study of amoeboid chemotaxis. These cells can be studied as a homogeneous population whose response to chemotactic stimulation is sufficiently synchronous to permit the correlation of the changes in cell shape and biochemical events during chemotaxis. Having demonstrated this synchrony of response, we show that actin polymerization occurs in two stages during stimulation with chemoattractants. The assembly of F‐actin that peaks between 40 and 60 sec after the onset of stimulation is temporally correlated with the growth of new pseudopods. F‐actin, which is assembled by 60 sec after stimulation begins, is localized in the new pseudopods that are extended at this time. Both stages of actin polymerization during chemotactic stimulation involve polymerization at the barbed ends of actin filaments based on the cytochalasin sensitivity of this response. We present a hypothesis in which actin polymerization is one of the major driving forces for pseudopod extension during chemotaxis. The predictions of this model, that localized regulation of actin nucleation activity and actin filament cross‐linking must occur, are discussed in the context of current models for signal transduction and of recent information regarding the types of actin‐binding proteins that are present in the cell

ISSN:0886-1544    

DOI:10.1002/cm.970100113

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

年代:1988

数据来源: WILEY

摘要:

AbstractA computer‐assisted Dynamic Morphology System (DMS) is described that allows the rapid quantitation of more than 30 parameters of motility and dynamic morphology for up to 40 amebae in parallel. This system also generates “difference pictures” for characterizing the dynamics of pseudopod formation. A 3‐D DMS is described, and application of DMS to problems of motility and chemotaxis in normal and mutant cells ofDictyostelium discoideumis r

ISSN:0886-1544    

DOI:10.1002/cm.970100114

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

年代:1988

数据来源: WILEY

摘要:

AbstractBirefringent fibrils (BRFs) with a positive sign composed of bundles of F‐actin were found throughout thePhysarumplasmodium with the mode of existence differing regionally. In the zone behind the leading edge of an advancing plasmodium, where cytoplasmic sol and gel were still not well differentiated, more BRFs came to the foreground when the endoplasm flowed backward (emptying phase), and a substantial portion disappeared when the endoplasm flowed forward (filling phase), except for nodes, from which BRFs were reorganized in the early emptying phase of each cycle. BRFs found in the wall of the streaming channel in the posterior network and the branched vein section ran in parallel to or helically around the channel. They were much more stable and maintained strong birefringence irrespective of the direction of the cytoplasmic flow. When the fan‐like expanse ceased moving forward, the BRFs no longer appeared and disappeared cyclically but persisted in the area which had previously been the front. We concluded that the site of the active contraction‐relaxation rhythm in an advancing plasmodium with antero‐posterior polarity is restricted to its frontal zone and that the rest of the plasmodium is in a state of “tonus” which continuously imparts a certain level of hydrostatic pressure to the interior. The meaning of the tonus and the mechanics of tensile force production in the plasmodium are discussed in terms of a working hypothesis arrived at from the phase relationship between isometric and isotonic contra

ISSN:0886-1544    

DOI:10.1002/cm.970100115

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

年代:1988

数据来源: WILEY

摘要:

AbstractTo move, leukocytes extend portions of their cortical cytoplasm as pseudopods. These pseudopods are filled with a three‐dimensional actin filament skeleton, the reversible assembly of which in response to receptor stimulation is thought to play a major role in providing the mechanical force for these protrusive movements. The organization of this actin skeleton occurs at different levels within the cell, and a number of macrophage proteins have been isolated and shown to affect the architecture, assembly, stability, and length of actin filaments in vitro. The architecture of cytoplasmic actin is regulated by proteins that cross‐link filaments in higher‐order structures. Actin‐binding protein plays a major role in defining network structure by cross‐linking actin filaments into orthogonal networks. Gelsolin may have a central role in regulating network structure. It binds to the sides of actin filaments and severs them, and binds the “barbed” filament end, thereby blocking monomer addition at this end. Gelsolin is activated to bind actin filaments by μM calcium. Dissociation of gelsolin bound on filament ends occurs in the presence of the polyphosphoinositides, PIP and PIP2. Calcium and PIP2have been shown to be intracellular messengers of ce

ISSN:0886-1544    

DOI:10.1002/cm.970100116

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

年代:1988

数据来源: WILEY

摘要:

AbstractMicrotubules are the major cytoskeletal component of foraminiferan reticulopodia. Video‐enhanced differential interference contrast light microscopy has demonstrated that the microtubules serve as the intracellular tracks along which rapid bidirectional organelle transport and cell surface motility occurs. Microtubules appear to move, both axially and laterally within the pseudopodial cytoplasm, and these microtubule translocations appear to drive the various reticulopodial movements. F‐actin is localized to discrete filament plaques form at sites of pseudopod‐substrate adhesion. Correlative immunofluorescence and electron microscopy reveals a structural interaction between microtubules and the actin‐containing filament plaques. Our recent data on reticulopodial motility are discussed in an historical context, and a model for foram motility, based on motile microtubules, is pr

ISSN:0886-1544    

DOI:10.1002/cm.970100117

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

年代:1988

数据来源: WILEY

摘要:

AbstractEvidence for a structural precursor of the focal contact in cultured fibroblasts and continuing studies on the development of the precursor and contact are discussed. The structural precursor consists of an F‐actin‐rich, rib‐like fiber within the motile lamellipodium. The focal contact forms beneath the fiber, part of which is retained at the contact as the initial adhesion plaque. Therefore, F‐actin is present at the contact from the beginning. Vinculin accumulates at the plaque during a 90‐second period after the contact forms. A novel feature of the distribution of talin has been found. The protein is present along the distal margin of the lamellipodium, where it is further concentrated as a series of nodes at the tips of each precursor and between precursors. This distribution of talin is independent of that which develops at the plaque after the contact forms. The structural development of the precursor has been followed with AVEC‐DIC optics. The process begins with the development of fine oblique fibers from small structural nodes at the margin of the lamellipodium, and continues with the fusion of the nodes at the margin and inward coalescence of the fibers. It is suggested that talin may function as a cross‐linking protein in the convergence of actin filaments at the membrane, while other actin‐bundling proteins participate in the inward coalescence of the filaments to form fibers. The F‐actin core of the precursor could provide a structural framework against which differences at the external surface of the membrane develop prior to

ISSN:0886-1544    

DOI:10.1002/cm.970100118

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

年代:1988

数据来源: WILEY

摘要:

AbstractIn goldfish xanthophores, the formation of pigment aggregate requires: (1) that a pigment organelle (carotenoid droplet) protein p57 be in the unphosphorylated state; (2) that self‐association of pigment organelles occur in a microtubule‐independent manner; and (3) that pigment organelles via p57 associate with microtubules. In the fully aggregated state, the pigment organelles are completely stationary. Pigment dispersion is initiated by activation of a cAMP‐dependent protein kinase, which phosphorylates p57 and allows pigment dispersion via an active process dependent on F‐actin and a cytosolic factor. This factor is not an ATPase, and its function is unknown. However, its abundance in different tissues parallels secretory activity of the tissues, suggesting a similarity between secretion and pigment dispersion in xanthophores. The identity of the motor for pigment dispersion is unclear. Experimental results show that pigment organelles isolated from cells with dispersed pigment have associated actin and ATPase activity comparable to myosin ATPase. This ATPase is probably an organelle protein of relative molecular mass ∼72,000, and unlikely to be an ion pump. Isolated pigment organelles without associated actin have 5× lower ATPase activity. Whether this organelle ATPase is the motor for pigment dispersion is under investigation. The process of pigment aggregation is poorly understood, with conflicting results for and against the involvement of intermediate

ISSN:0886-1544    

DOI:10.1002/cm.970100119

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

年代:1988

数据来源: WILEY

摘要:

AbstractThe endoplasmic reticulum (ER) and associated organelle and particle movements in onion (Allium cepa) bulb scale epidermal cells were observed, recorded, and analyzed using computer‐assisted video (AVEC‐DIC, AVEC‐POL and fluorescence) microscopy. The ER is composed of two interconnected sets of filamentous membrane tubules with diameters ranging from 0.1 to 0.5 μm. The first form a more stable, stationary network of intersecting polygonal membrane tubules lying closely appressed to the plasma membrane and continuous with a second very dynamic set of longer membrane tubules that often are located parallel to each other, shifting rapidly around the cytoplasm and forming dynamic knots or organization centers. The ER, mitochondria, and spherosomes fluoresced upon chlortetracycline treatment and are therefore presumed to sequester calcium. ER and mitochrondria also stain with the fluorescent dye, rhodamine 123. Mitochrondria and spherosomes are seen to move in the cytoplasm only along paths parallel to the axis of the ER tubules. Smaller particles (0.5 μm) tend to follow these same paths but may occasionally move independently. Particles and organelles move in close, but not in direct, association with the ER tubules. In optically favored cells, actin filaments were occasionally recorded located in parallel with the ER tubules and directly associated with moving particles. Streaming ceased promptly and reversibly upon treatment with cytochalasin B, which did not visibly disrupt the ER. Short‐term treatment with colchicine did not inhibit streaming or disrupt the ER network, whereas long‐term (hours) colchicine treatments caused the disappearance of the stationary, cortical polygonal networks and an aggregation of still slowly moving organelles and particles onto now visible actin filaments. This suggests that microtubule breakdown disrupts the three‐dimensional distribution of the ER and rearranges actin filaments in the cell's cytoplasm. Actin filaments must be directly involved in generation of movement of the particles and organelles. A three‐dimensional model, based on optical sectioning of the epidermal cells, is proposed to illustrate the distribution of the endoplasmic reticulum in onion epidermal

ISSN:0886-1544    

DOI:10.1002/cm.970100120

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

年代:1988

数据来源: WILEY

ISSN:0886-1544    

DOI:10.1002/cm.970100121

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

年代:1988

数据来源: WILEY