Kinetics of actin monomer exchange at the slow growing ends of actin filaments and their relation to the elongation of filaments shortened by gelsolin

1986 ◽  
Vol 7 (5) ◽  
pp. 446-454 ◽  
Author(s):  
Paul A. Janmey ◽  
Thomas P. Stossel
Keyword(s):  
Actin Filaments ◽  
Actin Monomer ◽  
Slow Growing ◽  
Kinetics Of

scholarly journals Force-mediated cellular anisotropy and plasticity dictate the elongation dynamics of embryos

2021 ◽  
Vol 7 (27) ◽  
pp. eabg3264
Author(s):  
Chao Fang ◽  
Xi Wei ◽  
Xueying Shao ◽  
Yuan Lin
Keyword(s):  
Plastic Deformation ◽  
Caenorhabditis Elegans ◽  
Muscle Contraction ◽  
Dynamic Model ◽  
Actin Filaments ◽  
Actin Bundles ◽  
Abnormal Embryo ◽  
Intracellular Forces ◽  
Kinetics Of ◽  
Good Agreement

We developed a unified dynamic model to explain how cellular anisotropy and plasticity, induced by alignment and severing/rebundling of actin filaments, dictate the elongation dynamics of Caenorhabditis elegans embryos. It was found that the gradual alignment of F-actins must be synchronized with the development of intracellular forces for the embryo to elongate, which is then further sustained by muscle contraction–triggered plastic deformation of cells. In addition, we showed that preestablished anisotropy is essential for the proper onset of the process while defects in the integrity or bundling kinetics of actin bundles result in abnormal embryo elongation, all in good agreement with experimental observations.

scholarly journals Dynamics of Tpm1.8 domains on actin filaments with single molecule resolution

2020 ◽  
Author(s):  
Ilina Bareja ◽  
Hugo Wioland ◽  
Miro Janco ◽  
Philip R. Nicovich ◽  
Antoine Jégou ◽  
...  
Keyword(s):  
Actin Filament ◽  
Single Molecule ◽  
Actin Filaments ◽  
High Probability ◽  
Actin Binding ◽  
Single Molecule Level ◽  
Filament Dynamics ◽  
Kinetics Of ◽  
Insight Into

ABSTRACTTropomyosins regulate dynamics and functions of the actin cytoskeleton by forming long chains along the two strands of actin filaments that act as gatekeepers for the binding of other actin-binding proteins. The fundamental molecular interactions underlying the binding of tropomyosin to actin are still poorly understood. Using microfluidics and fluorescence microscopy, we observed the binding of fluorescently labelled tropomyosin isoform Tpm1.8 to unlabelled actin filaments in real time. This approach in conjunction with mathematical modeling enabled us to quantify the nucleation, assembly and disassembly kinetics of Tpm1.8 on single filaments and at the single molecule level. Our analysis suggests that Tpm1.8 decorates the two strands of the actin filament independently. Nucleation of a growing tropomyosin domain proceeds with high probability as soon as the first Tpm1.8 molecule is stabilised by the addition of a second molecule, ultimately leading to full decoration of the actin filament. In addition, Tpm1.8 domains are asymmetrical, with enhanced dynamics at the edge oriented towards the barbed end of the actin filament. The complete description of Tpm1.8 kinetics on actin filaments presented here provides molecular insight into actin-tropomyosin filament formation and the role of tropomyosins in regulating actin filament dynamics.

scholarly journals Mechanism of synergistic actin filament pointed end depolymerization by cyclase-associated protein and cofilin

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Tommi Kotila ◽  
Hugo Wioland ◽  
Giray Enkavi ◽  
Konstantin Kogan ◽  
Ilpo Vattulainen ◽  
...  
Keyword(s):  
Molecular Mechanism ◽  
Actin Filament ◽  
Actin Filaments ◽  
Monomeric Form ◽  
Actin Monomer ◽  
Filament Assembly ◽  
Structural Work ◽  
Cytoskeletal Dynamics ◽  
Pointed End ◽  
Dependent Processes

AbstractThe ability of cells to generate forces through actin filament turnover was an early adaptation in evolution. While much is known about how actin filaments grow, mechanisms of their disassembly are incompletely understood. The best-characterized actin disassembly factors are the cofilin family proteins, which increase cytoskeletal dynamics by severing actin filaments. However, the mechanism by which severed actin filaments are recycled back to monomeric form has remained enigmatic. We report that cyclase-associated-protein (CAP) works in synergy with cofilin to accelerate actin filament depolymerization by nearly 100-fold. Structural work uncovers the molecular mechanism by which CAP interacts with actin filament pointed end to destabilize the interface between terminal actin subunits, and subsequently recycles the newly-depolymerized actin monomer for the next round of filament assembly. These findings establish CAP as a molecular machine promoting rapid actin filament depolymerization and monomer recycling, and explain why CAP is critical for actin-dependent processes in all eukaryotes.

scholarly journals Basic Characterization of Plant Actin Depolymerizing Factors: A Simplified, Streamlined Guide

2019 ◽  
Author(s):  
Sonali Sengupta ◽  
Kanniah Rajasekaran ◽  
Niranjan Baisakh
Keyword(s):  
Ionic Strength ◽  
Actin Filaments ◽  
Actin Dynamics ◽  
Actin Binding ◽  
Actin Monomer ◽  
Oligomeric State ◽  
Binding Partners ◽  
Helical Twist

Abstract Actin depolymerizing factors (ADFs) are small monomeric actin-binding proteins that alter the oligomeric state of cellular actin. Members of the ADF family can bind both the G-actin and F-actin in plants, and their functions are regulated by cellular pH, ionic strength and availability of other binding partners. Actin depolymerization activity is reportedly essential for plant viability. By binding to the ADP-bound form of actin, ADFs severe actin filaments and thereby provide more barbed filament ends for polymerization. They also increase the rate of dissociation of F-actin monomer by changing the helical twist of the actin filament. These two activities together make ADF the major regulator of actin dynamics in plant cell. Therefore, it is essential to measure the binding and depolymerization activity of the plant ADFs. Here, we present a simplified, streamlined step-by-step protocol to quickly measure these important functions of the ADF proteins in vitro.

scholarly journals Insight into the kinetics and the mode of the interaction between smooth muscle calponin and F-actin.

2002 ◽  
Vol 49 (2) ◽  
pp. 471-479
Author(s):  
Janusz Kołakowski ◽  
Renata Dabrowska
Keyword(s):  
Smooth Muscle ◽  
Light Scattering ◽  
Conformational Changes ◽  
Actin Filaments ◽  
Molar Ratio ◽  
Flow Measurements ◽  
Pyrene Fluorescence ◽  
Kinetics Of ◽  
Insight Into ◽  
Parallel Measurements

Kinetics of the smooth muscle calponin-F-actin interaction was studied by stopped-flow measurements of light scattering and fluorescence intensity of pyrene-labelled F-actin. The intensity and character of the changes in light scattering, and thus the mode of calponin binding to actin filaments leading to changes in their shape and bundling, depend on the molar ratio of the two proteins. Parallel measurements of pyrene-fluorescence quenching upon calponin binding revealed that intrinsic conformational changes in actin filaments are delayed relative to the binding process and are not markedly influenced by the mode of calponin binding. Bundling of actin filaments by calponin was not correlated with fluorescence changes and thus with alterations in the structure of actin filaments.

Reaktionen und relative Affinitäten zwischen verschiedenen Nucleotiden und F-Actin unter Ultraschall

1965 ◽  
Vol 20 (10) ◽  
pp. 977-989 ◽  
Author(s):  
Irene Kakol ◽  
Hans H. Weber
Keyword(s):  
Actin Filaments ◽  
Mechanical Treatment ◽  
Quaternary Structure ◽  
Optimal Conditions ◽  
Single Nucleotide ◽  
Teflon Piston ◽  
Affinity Constants ◽  
Relative Affinity ◽  
Kinetics Of ◽  
Zero Time

1. Like the polymerization of G-actin, the repolymerization of F-actin treated with USV occurs in the presence of every nucleosidetriphosphate (NTP) as well as nucleosiddiphosphate (NDP) that have been investigated.2. At 4°C solutions of F-actin exchange about 9 per cent of their bound nucleotide instantaneously and about 7 percent during the next 24 hours even then when they are not mechanically treated.3. On the other hand, every nucleotide is exchanged with considerable velocity if the F-actin solution at pH ~ 7 and 4°C is mechanically treated with USV or in the homogenisator. The rate of exchange decreases considerably if the KCl of the solution is replaced by MgCl2. Moreover, the rate of exchange decreases with the increase of the concentration of F-actin from 1,5·10-5 M to 5·10-5 M if the solution is treated with USV (but not when treated in the homogenisator).4. The exchanged amounts of all investigated nucleotides are identical if during the mechanical treatment the nucleotides are present in a concentration of ≧ 10-3 M. Under optimal conditions (KCl, 1,6 ·10-5 M F-actin) either in a USV-period of 40′ or after a thousand movements of the teflon piston in the homogenisator, the exchange of the nucleotides reaches an average of 90 per cent. Even after mechanical treatment one, F-actin still binds one mole nucleotide.5. The kinetics of exchange is monomolecular in any case.6. If the nucleotide entering by exchange is a triphosphate one phosphate is split off. The kinetics of the splitting is linear.7. At zero time the rate of exchange and that of splitting are virtually the same, but later the measured rate of exchange lags more and more behind the rate of splitting. At a given moment the ratio of split nucleotide, to exchanged nucleotide is the ratio one would expect under the following two conditions: (a) The single nucleotide ion is split only during the regeneration of a break of the F-actin filament if it has been bound to the point of breaking immediately before the regeneration, (b) The breaks between the single monomeres are statistically distributed over the lengths of the filament (i. e. the interaction of all monomers is of equal strength).8. If the exchange occurs in the presence of two competing nucleotides the ratio of the affinities of the competing nucleotides to F-actin can be calculated. Thus it has been found that the relative affinities decrease in the following sequence: ATP>ITP>ADP>IDP>GTP>CTP. This sequence coincides with that of the relative affinity constants to G-actin; in addition the numerical values of the affinity constants to F-actin and to G-actin are similar.9. The same rules obey the binding of the different nucleotides to G-actin and to F-actin as well as the splitting of the nucleotides during the polymerization of G-actin and during the repolymerization of broken F-actin filaments. Thus the seemingly different behaviour of G-actin nucleotide compounds and of F-actin nucleotide compounds becomes the same or nearly so if the steric hindrance of the nucleotide reactions (because of the special quaternary structure of F-actin) is removed by mechanical treatment such as USV or treatment in the homogenisator.

scholarly journals Structure of the actin-depolymerizing factor homology domain in complex with actin

2008 ◽  
Vol 182 (1) ◽  
pp. 51-59 ◽  
Author(s):  
Ville O. Paavilainen ◽  
Esko Oksanen ◽  
Adrian Goldman ◽  
Pekka Lappalainen
Keyword(s):  
Crystal Structure ◽  
Atomic Structure ◽  
Actin Filaments ◽  
Driving Force ◽  
Actin Dynamics ◽  
Actin Binding ◽  
Actin Monomer ◽  
Nucleotide Exchange ◽  
Actin Depolymerizing Factor ◽  
Cellular Processes

Actin dynamics provide the driving force for many cellular processes including motility and endocytosis. Among the central cytoskeletal regulators are actin-depolymerizing factor (ADF)/cofilin, which depolymerizes actin filaments, and twinfilin, which sequesters actin monomers and caps filament barbed ends. Both interact with actin through an ADF homology (ADF-H) domain, which is also found in several other actin-binding proteins. However, in the absence of an atomic structure for the ADF-H domain in complex with actin, the mechanism by which these proteins interact with actin has remained unknown. Here, we present the crystal structure of twinfilin's C-terminal ADF-H domain in complex with an actin monomer. This domain binds between actin subdomains 1 and 3 through an interface that is conserved among ADF-H domain proteins. Based on this structure, we suggest a mechanism by which ADF/cofilin and twinfilin inhibit nucleotide exchange of actin monomers and present a model for how ADF/cofilin induces filament depolymerization by weakening intrafilament interactions.

scholarly journals Actin turnover–dependent fast dissociation of capping protein in the dendritic nucleation actin network: evidence of frequent filament severing

2006 ◽  
Vol 175 (6) ◽  
pp. 947-955 ◽  
Author(s):  
Takushi Miyoshi ◽  
Takahiro Tsuji ◽  
Chiharu Higashida ◽  
Maud Hertzog ◽  
Akiko Fujita ◽  
...  
Keyword(s):  
Single Molecule ◽  
Actin Filaments ◽  
Dissociation Rate ◽  
Actin Network ◽  
Lim Kinase ◽  
Capping Protein ◽  
Filament Dynamics ◽  
Actin Turnover ◽  
Kinetics Of

Actin forms the dendritic nucleation network and undergoes rapid polymerization-depolymerization cycles in lamellipodia. To elucidate the mechanism of actin disassembly, we characterized molecular kinetics of the major filament end-binding proteins Arp2/3 complex and capping protein (CP) using single-molecule speckle microscopy. We have determined the dissociation rates of Arp2/3 and CP as 0.048 and 0.58 s−1, respectively, in lamellipodia of live XTC fibroblasts. This CP dissociation rate is three orders of magnitude faster than in vitro. CP dissociates slower from actin stress fibers than from the lamellipodial actin network, suggesting that CP dissociation correlates with actin filament dynamics. We found that jasplakinolide, an actin depolymerization inhibitor, rapidly blocked the fast CP dissociation in cells. Consistently, the coexpression of LIM kinase prolonged CP speckle lifetime in lamellipodia. These results suggest that cofilin-mediated actin disassembly triggers CP dissociation from actin filaments. We predict that filament severing and end-to-end annealing might take place fairly frequently in the dendritic nucleation actin arrays.

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