SEARCH · PubMed Health

Results for “actin filaments”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 19 recordsLinked to original sources

PubMed1993

Formation of liquid crystals from actin filaments.

Actin is cross-linked by actin-binding proteins in the cytoplasm to form either isotropic or highly oriented anisotropic structures. The inherent orientation among actin filaments could influence whether an isotropic or highly oriented anisotropic structure is formed. A highly oriented state can arise spontaneously through the formation of liquid crystals as predicted by polymer theory. In this study, the ability of filamentous actin to form liquid crystalline domains was detected using the anisotropic component of scattered light and by observation of birefringence. As liquid crystalline domains formed, the intensity of the anisotropic component of scattered light increased, and birefringent macroscopic oriented domains were directly observed. The formation of liquid crystalline domains was dependent on the concentration of actin filaments and on the average filament length controlled by varying the ratio of gelsolin to actin monomers. The concentration of actin filaments required to form liquid crystalline domains increased moderately as the average length was decreased. At a fixed actin concentration, orientation among the filaments attained a maximum value at a ratio of actin to gelsolin in the range from 1500 to 2000 and decreased as the ratio was increased or decreased from this range. The results are not well explained by theoretical treatments for liquid crystal formation by monodisperse, charged worm-like chains. Differences from the theoretical predictions for formation of liquid crystals are most likely due to the polydisperse filament length of actin. This phenomenon may have important effects on the structural and rheological properties of the cytoplasm in living cells.

Actin Cytoskeleton↗

PubMed2006

Dynamic polymorphism of single actin molecules in the actin filament.

Actin filament dynamics are critical in cell motility. The structure of actin filament changes spontaneously and can also be regulated by actin-binding proteins, allowing actin to readily function in response to external stimuli. The interaction with the motor protein myosin changes the dynamic nature of actin filaments. However, the molecular bases for the dynamic processes of actin filaments are not well understood. Here, we observed the dynamics of rabbit skeletal-muscle actin conformation by monitoring individual molecules in the actin filaments using single-molecule fluorescence resonance energy transfer (FRET) imaging with total internal reflection fluorescence microscopy (TIRFM). The time trajectories of FRET show that actin switches between low- and high-FRET efficiency states on a timescale of seconds. If actin filaments are chemically cross-linked, a state that inhibits myosin motility, the equilibrium shifts to the low-FRET conformation, whereas when the actin filament is interacting with myosin, the high-FRET conformation is favored. This dynamic equilibrium suggests that actin can switch between active and inactive conformations in response to external signals.

Actin Cytoskeleton↗

PubMed1999

Aip1p interacts with cofilin to disassemble actin filaments.

Actin interacting protein 1 (Aip1) is a conserved component of the actin cytoskeleton first identified in a two-hybrid screen against yeast actin. Here, we report that Aip1p also interacts with the ubiquitous actin depolymerizing factor cofilin. A two-hybrid-based approach using cofilin and actin mutants identified residues necessary for the interaction of actin, cofilin, and Aip1p in an apparent ternary complex. Deletion of the AIP1 gene is lethal in combination with cofilin mutants or act1-159, an actin mutation that slows the rate of actin filament disassembly in vivo. Aip1p localizes to cortical actin patches in yeast cells, and this localization is disrupted by specific actin and cofilin mutations. Further, Aip1p is required to restrict cofilin localization to cortical patches. Finally, biochemical analyses show that Aip1p causes net depolymerization of actin filaments only in the presence of cofilin and that cofilin enhances binding of Aip1p to actin filaments. We conclude that Aip1p is a cofilin-associated protein that enhances the filament disassembly activity of cofilin and restricts cofilin localization to cortical actin patches.

Actin Depolymerizing Factors↗

PubMed1996

Nuclear ion channel activity is regulated by actin filaments.

Actin filaments are novel second messengers involved in ion channel regulation. Because cytoskeletal components interact with the nuclear envelope, the actin cytoskeleton may also control nuclear membrane function. In this report, the patch-clamp technique was applied to isolated nuclei from amphibian A6 epithelial cells to assess the role of actin filaments on nuclear ion channel activity under nucleus-attached or -excised conditions. The most prevalent spontaneous nuclear ion channel species, 76% (n = 46), was cation selective and had a maximal single-channel conductance of approximately 420 pS. Nuclear ion channels also displayed multiple subconductance states, including channel activity of 26 pS that was frequently observed. Nuclear ion channel activity on otherwise quiescent patches was induced by either addition of the actin cytoskeleton disrupter cytochalasin D (CD; 5 micrograms/ml, 60%, 3 of 5 patches) or actin (10-1,000 micrograms/ml) to the bathing solution of nucleus-attached patches (59%, 13 of 22 patches). Actin also induced ion channel activity in quiescent excised inside-out patches from the nuclear envelope (80%, 4 of 5 patches). In contrast, addition of bovine serum albumin (10-1,000 micrograms/ml) to the bathing solution of nucleus-attached patches was without effect on nuclear ion channel activity (5 of 5 patches). The monoclonal antibody MAb414, specific for nuclear pore complex proteins, completely prevented either spontaneous or cytosolic actin-induced nuclear ion channels under nucleus-attached conditions (4 of 4 patches) but not intranuclear actin-induced nuclear ion channels under excised inside-out conditions (3 of 3 patches). In nucleus-attached patches, channel activity was readily activated by addition of the G-actin-binding protein deoxyribonuclease I to nucleus-attached patches (56%, 5 of 9 patches) or further addition of the actin-cross-linker filamin in the presence of actin (57%, 4 of 7 patches). The data indicate that dynamic changes in actin filament organization may represent a novel mechanism to control nuclear function.

Actins↗

PubMed1987

Microtubules, intermediate filaments, and actin filaments in the odontoblast of rat incisor.

Actin filaments, intermediate filaments, and microtubules in the odontoblasts of rat incisors were investigated electron microscopically using heavy meromyosin and taxol. Actin filaments were abundant at the periphery of the odontoblast process in the form of a network or in bundles. In a branch of the odontoblast process, longitudinally oriented actin filament bundles were found. Most actin filaments were associated with the plasma membrane via electron-dense material which stained with tannic acid. The intermediate filaments had a diameter of 11 to 13 nm. They were distributed throughout the cytoplasm of odontoblasts. They ran lengthwise in the core of the odontoblast process, which showed a different distribution compared with that of actin filaments. Microtubules, which were disrupted after Triton X-100 but preserved by addition of taxol, tended to be associated with intermediate filaments. Such a relation was also seen in conventional preparations. Coated vesicles, which were abundant at the periphery of the odontoblast process, were often associated with actin filaments. Therefore, it is suggested that actin filaments, in the odontoblast process at least, play a role associated with the coated vesicles at the periphery of the process, and may be involved in coated vesicle transport.

Actins↗

PubMed2004

Eps8 controls actin-based motility by capping the barbed ends of actin filaments.

Actin filament barbed-end capping proteins are essential for cell motility, as they regulate the growth of actin filaments to generate propulsive force. One family of capping proteins, whose prototype is gelsolin, shares modular architecture, mechanism of action, and regulation through signalling-dependent mechanisms, such as Ca(2+) or phosphatidylinositol-4,5-phosphate binding. Here we show that proteins of another family, the Eps8 family, also show barbed-end capping activity, which resides in their conserved carboxy-terminal effector domain. The isolated effector domain of Eps8 caps barbed ends with an affinity in the nanomolar range. Conversely, full-length Eps8 is auto-inhibited in vitro, and interaction with the Abi1 protein relieves this inhibition. In vivo, Eps8 is recruited to actin dynamic sites, and its removal impairs actin-based propulsion. Eps8-family proteins do not show any similarity to gelsolin-like proteins. Thus, our results identify a new family of actin cappers, and unveil novel modalities of regulation of capping through protein-protein interactions. One established function of the Eps8-Abi1 complex is to participate in the activation of the small GTPase Rac, suggesting a multifaceted role for this complex in actin dynamics, possibly through the participation in alternative larger complexes.

Actin Cytoskeleton↗

PubMed1982

Interactions between actin filaments and between actin filaments and membranes in quick-frozen and deeply etched hair cells of the chick ear.

Replicas of the apical surface of hair cells of the inner ear (vestibular organ) were examined after quick freezing and rotary shadowing. With this technique we illustrate two previously undescribed ways in which the actin filaments in the stereocilia and in the cuticular plate are attached to the plasma membrane. First, in each stereocilium there are threadlike connectors running from the actin filament bundle to the limiting membrane. Second, many of the actin filaments in the cuticular plate are connected to the apical cell membrane by tiny branched connecting units like a "crow's foot." Where these "feet" contact the membrane there is a small swelling. These branched "feet" extend mainly from the ends of the actin filaments but some connect the lateral surfaces of the actin filaments as well. Actin filaments in the cuticular plate are also connected to each other by finer filaments, 3 nm in thickness and 74 +/- 14 nm in length. Interestingly, these 3-nm filaments (which measure 4 nm in replicas) connect actin filaments not only of the same polarity but of opposite polarities as documented by examining replicas of the cuticular plate which had been decorated with subfragment 1 (S1) of myosin. At the apicolateral margins of the cell we find two populations of actin filaments, one just beneath the tight junction as a network, the other at the level of the zonula adherens as a ring. The latter which is quite substantial is composed of actin filaments that run parallel to each other; adjacent filaments often show opposite polarities, as evidenced by S1 decoration. The filaments making up this ring are connected together by the 3-nm connectors. Because of the polarity of the filaments this ring may be a "contractile" ring; the implications of this is discussed.

Actins↗

PubMed1991

Osmotically induced electrical signals from actin filaments.

Actin filaments, F-actin, a major component of the cortical cytoskeleton, play an important role in a variety of cell functions. In this report we have assessed the role of osmotic stress on the electrochemical properties of F-actin. The spontaneous Donnan potential of a polymerized actin solution (5 mg/ml) was -3.93 +/- 1.84 mV, which was linearly reduced by osmotic stress on the order of 1-20 mOsm (0.28 +/- 0.06 mV/mM). Calculated surface charge density was reduced and eventually reversed by increasing the osmotic stress as expected for a phase transition behavior. The electro-osmotic behavior of F-actin disappeared at pH 5.5 and was dependent on its filamentous nature. Furthermore, osmotically stressed F-actin displayed a nonlinear electric response upon application of electric fields on the order of 500-2,000 V/cm. These data indicate that F-actin in solution may display nonideal electro-osmotic properties consistent with ionic "cable" behavior which may be of biological significance in the processing and conduction of electrical signals within the cellular compartment.

Actins↗

PubMed1983

Direct measurement of critical concentrations and assembly rate constants at the two ends of an actin filament.

Actin filament bundles isolated from Limulus sperm were used for quantitative electron microscope studies of F-actin assembly. The assembly rate constants were calculated. In addition, the critical concentrations (Cos) for both filament ends were directly determined. In 75 mM KCl and 1-5 mM Mg++, the Cos were 0.1 microM and 0.5 microM for the barbed and pointed ends, respectively. Substitution of Ca++ (20-200 microM) for Mg++ resulted in Cos of 0.4 microM for both filament ends. Consistent with these findings, filament growth occurred only from the barbed ends of Limulus bundles "seeded" into F-actin solutions in KCl and Mg++. Finally, filaments originally grown from the pointed filament ends of Limulus bundles were gradually lost as the actin solution reached steady state. These results demonstrate that actin filaments can "treadmill" under physiological conditions, albeit at very slow rates.

Actins↗

PubMed1994

The actin side-binding domain of gelsolin also caps actin filaments. Implications for actin filament severing.

Gelsolin is an actin filament-severing and -capping protein which is inhibited by polyphosphoinositides (PPI). Severing requires gelsolin binding to the side of the filaments through a site in segments 2 and 3 (S2-3) to position another site in segment 1 (S1) to sever filaments. In this paper, we report that S2-3, like S1, caps actin filaments. Since neither S1 and S2-3 caps as well as gelsolin, and neither severs actin filament, S2-3 may actively contribute to severing by capping filaments cooperatively with S1. We used deletional mutagenesis to locate the S2-3 sequence required for actin filament side binding, capping, and PPI binding and found that these sites are located close to the NH2 terminus of S2 (residues 161-172). S3, a segment which has no known function up to now and does not by itself bind actin, contributes to stable capping and may contain an additional PPI-binding site.

Actins↗

PubMed1986

Cryo-electron microscopy and three-dimensional reconstruction of actin filaments.

Actin filaments have been examined by electron microscopy whilst in a frozen-hydrated state. Filaments embedded in a vitreous water layer are basically similar to those prepared by negative staining and show characteristic helical substructure, where the pitches of the helices are about 70 nm and 6 nm. Variability in spacing between long pitch helix cross-over points has been observed, which is consistent with intrinsic angular disorder between successive filament subunits. Fourier transforms of the most ordered filaments show four strong layer lines that index as the first, fifth, sixth and seventh orders of a 35 nm repeat. A three-dimensional helical reconstruction, calculated to a resolution of about 4 nm, shows the individual subunits to be orientated with their long axes roughly perpendicular to the filament axis. Each subunit is somewhat curved and is resolved into two domains. Most connections between successive subunits appear to be made close to the filament axis. We also report on the performance of the specimen holder (Philips PW 5699) used in this work.

Actins↗

PubMed2000

Muscle force arises by actin filament rotation and torque in the Z-filaments.

Actin filament rotation in skeletal muscle is studied by a mechanical model that simulates structure and tension. The four anchoring Z-filaments are twisted around and change the structure of the Z-lattice. The "small square" without twist represents the resting stage of muscle. Torque causes contraction by clockwise rotation (as seen from the Z-band), drilling into the A-band and transition of the "small square" to "basket weave" by increasing the twist and decreasing the torque. Release decreases the torque ("force-depression") by passive clockwise rotation. Stretch causes increased torque ("stretch activation") by passive counterclockwise rotation. Torque arises during Ca(2+)-activation by a conformational change in the highly charged coiled-coils: The four alpha-actinin Z-filaments generate strong torque for the isometric tension. Quick release experiments show that less than one rotation reduces this torque to zero. The 5-12 rotations necessary for isotonic shortening result from torque-generation in the two long tropomyosin coiled-coils. Myosin controls the velocity of active and passive rotations.

Actinin↗

PubMed2000

Evidence of a new type of protein-protein interaction: desensitized actomyosin blocks Ca(2+)-sensitivity of the natural one. A possible model for an intracellular signalling system related to actin filaments.

Actin filaments are certainly believed to function as an intracellular signalling system; however, this is not confirmed by direct evidence. We used a two-layer actomyosin gel with a concentration gradient of the troponin-tropomyosin complex (TT-complex, Ca(2+)-sensitive system) between the two layers. To prepare one layer of the system, natural actomyosin (nAM) rich in TT-complex was used. To prepare the second layer, we used desensitized actomyosin (dAM) without the complex. All experimental studies were made in medium with a low ionic strength. Two phenomena were observed: (1) dAM blocks Ca(2+)-sensitivity of nAM when the dAM weight portion in the system (as well as in mixed nAM + dAM suspension) reaches 40% and more; further increase of the dAM portion does not affect the Ca(2+)-sensitivity; (2) it was electrophoretically shown that a rapid diffusion of the TT-complex from nAM gel into the dAM gel took place. The apparent diffusion coefficient for the TT-complex in dAM gel is about (1-4).10(-4) cm2/sec, i.e. three orders higher than the same values for protein diffusion in water.

Actins↗

PubMed1986

Nonlinear increase of elongation rate of actin filaments with actin monomer concentration.

The nonlinear increase of the elongation rate of actin filaments above the critical monomer concentration was investigated by nucleated polymerization of actin. Significant deviations from linearity were observed when actin was polymerized in the presence of magnesium ions. When magnesium ions were replaced by potassium or calcium ions, no deviations from linearity could be detected. The nonlinearity was analyzed by two simple assembly mechanisms. In the first model, if the ATP hydrolysis by polymeric actin is approximately as fast as the incorporation of monomers into filaments, terminal subunits of lengthening filaments are expected to carry to some extent ADP. As ADP-containing subunits dissociate from the ends of actin filaments faster than ATP-containing subunits, the rate of elongation of actin filaments would be nonlinearly correlated with the monomer concentration. In the second model (conformational change model), actin monomers and filament subunits were assumed to occur in two conformations. The association and dissociation rates of actin molecules in the two conformations were thought to be different. The equilibrium distribution between the two conformations was assumed to be different for monomers and filament subunits. The ATP hydrolysis was thought to lag behind polymerization and conformational change. As under the experimental conditions the rate of ATP hydrolysis by polymeric actin was independent of the concentration of filament ends, the observed nonlinear increase of the rate of elongation with the monomer concentration above the critical monomer concentration was unlikely to be caused by ATP hydrolysis at the terminal subunits. The conformational change model turned out to be the simplest assembly mechanism by which all available experimental data could be explained.

Actins↗

PubMed1985

Calcium-dependent interaction of actin filaments with actin binding protein in the presence of calmodulin and caldesmon.

Caldesmon, calmodulin-, and actin-binding protein of chicken gizzard did not affect the process of polymerization of actin induced by 0.1 M KCl. Caldesmon binds to F-actin, thus inhibiting the gelation action of actin binding protein (ABP; filamin). Low shear viscosity and flow birefringence measurements revealed that in a system of calmodulin, caldesmon, ABP, and F-actin, gelation occurs in the presence of micromolar Ca2+ concentrations, but not in the absence of Ca2+. Electron microscopic observations showed the Ca2+-dependent formation of actin bundles in this system. These results were interpreted by the flip-flop mechanism: in the presence of Ca2+, a calmodulin-caldesmon complex is released from actin filaments on which ABP exerts its gelating action. On the other hand, in the absence of Ca2+, caldesmon remains bound to actin filaments, thus preventing the action of ABP.

Actins↗

PubMed1993

The actin/actin interactions involving the N-terminus of the DNase-I-binding loop are crucial for stabilization of the actin filament.

Actin can be specifically cleaved between residues 42 and 43 with a novel protease from Escherichia coli A2 strain (ECP) [Khaitlina, S. Y., Collins, J. H., Kuznetsova, I.M., Pershina, V.P., Synakevich, I.G., Turoverov, K.K. & Usmanova, A.M. (1991) FEBS Lett. 279, 49-51]. The resulting C-terminal and N-terminal fragments remained associated to one another in the presence of either Ca2+ or Mg2+. The protease-treated actin was, however, neither able to spontaneously assemble into filaments nor to copolymerize with intact actin unless its tightly bound Ca2+ was replaced with Mg2+. Substitution of Mg2+ for the bound Ca2+ was also necessary to partially restore the ability of the protease-treated actin to inhibit the DNase I activity. The critical concentration for KCl-induced polymerization of ECP-treated ATP-Mg-G-actin, determined by measuring the fluorescence of pyrenyl label, was approximately threefold higher than that for actin cleaved between residues 47 and 48 using subtilisin, and 36-fold higher than the critical concentration for polymerization of intact actin under the same conditions. Morphologically, the filaments of ECP-treated actin were indistinguishable from those of intact actin. Comparison of the fluorescence spectra of pyrenyl-labelled actins and chemical cross-linking with N,N'-1,2-phenylenebismaleimide have, however, revealed structural differences between the filaments assembled from ECP-treated actin and those of intact as well as subtilisin-treated actin. Moreover, the filaments of ECP-treated actin were easily disrupted by centrifugal forces or shearing stress unless they were stabilized by phalloidin. The results are consistent with the direct participation of the region around residues 42 and 43 in the monomer/monomer interactions as predicted from the atomic model of F-actin [Holmes, K.C., Popp, D., Gebhard, W. & Kabsch, W. (1990) Nature 347, 44-49] and suggest that the interactions involving this region are of primary importance for stabilization of the actin filament. The mechanism of the regulation of actin polymerization by the tightly bound divalent cation is also discussed.

Actins↗

PubMed2003

Regulation of actin filament dynamics by actin depolymerizing factor/cofilin and actin-interacting protein 1: new blades for twisted filaments.

Actin depolymerizing factor (ADF)/cofilin enhances turnover of actin filaments by severing and depolymerizing filaments. A number of proteins functionally interact with ADF/cofilin to modulate the dynamics of actin filaments. Actin-interacting protein 1 (AIP1) has emerged as a conserved WD-repeat protein that specifically enhances ADF/cofilin-induced actin dynamics. Interaction of AIP1 with actin was originally characterized by a yeast two-hybrid system. However, biochemical studies revealed its unique activity on ADF/cofilin-bound actin filaments. AIP1 alone has negligible effects on actin filament dynamics, whereas in the presence of ADF/cofilin, AIP1 enhances filament fragmentation by capping ends of severed filaments. Studies in model organisms demonstrated that AIP1 genetically interacts with ADF/cofilin and participates in several actin-dependent cellular events. The crystal structure of AIP1 revealed its unique structure with two seven-bladed beta-propeller domains. Thus, AIP1 is a new class of actin regulatory proteins that selectively enhances ADF/cofilin-dependent actin filament dynamics.

Actin Cytoskeleton↗

PubMed2000

Multiple-particle tracking measurements of heterogeneities in solutions of actin filaments and actin bundles.

One of the central functions of actin cytoskeleton is to provide the mechanical support required for the establishment and maintenance of cell morphology. The mechanical properties of actin filament assemblies are a consequence of both the available polymer concentration and the actin regulatory proteins that direct the formation of higher order structures. By monitoring the displacement of well-dispersed microspheres via fluorescence microscopy, we probe the degree of spatial heterogeneity of F-actin gels and networks in vitro. We compare the distribution of the time-dependent mean-square displacement (MSD) of polystyrene microspheres imbedded in low- and high-concentration F-actin solutions, in the presence and absence of the F-actin-bundling protein fascin. The MSD distribution of a 2. 6-microM F-actin solution is symmetric and its standard deviation is similar to that of a homogeneous solution of glycerol of similar zero-shear viscosity. However, increasing actin concentration renders the MSD distribution wide and asymmetric, an effect enhanced by fascin. Quantitative changes in the shape of the MSD distribution correlate qualitatively with the presence of large heterogeneities in F-actin solutions produced by increased filament concentration and the presence of actin bundles, as detected by confocal microscopy. Multiple-particle tracking offers a new, quantitative method to characterize the organization of biopolymers in solution.

Actins↗