α-Actinin and Filamin Cooperatively Enhance the Stiffness of Actin Filament Networks

BACKGROUND: The close subcellular proximity of different actin filament crosslinking proteins suggests that these proteins may cooperate to organize F-actin structures to drive complex cellular functions during cell adhesion, motility and division. Here we hypothesize that alpha-actinin and filamin, two major F-actin crosslinking proteins that are both present in the lamella of adherent cells, display synergistic mechanical functions. METHODOLOGY/PRINCIPAL FINDINGS: Using quantitative rheology, we find that combining alpha-actinin and filamin is much more effective at producing elastic, solid-like actin filament networks than alpha-actinin and filamin separately. Moreover, F-actin networks assembled in the presence of alpha-actinin and filamin strain-harden more readily than networks in the presence of either alpha-actinin or filamin. SIGNIFICANCE: These results suggest that cells combine auxiliary proteins with similar ability to crosslink filaments to generate stiff cytoskeletal structures, which are required for the production of internal propulsive forces for cell migration, and that these proteins do not have redundant mechanical functions

GTPase Activity, Structure, and Mechanical Properties of Filaments Assembled from Bacterial Cytoskeleton Protein MreB

MreB, a major component of the recently discovered bacterial cytoskeleton, displays a structure homologous to its eukaryotic counterpart actin. Here, we study the assembly and mechanical properties of Thermotoga maritima MreB in the presence of different nucleotides in vitro. We found that GTP, not ADP or GDP, can mediate MreB assembly into filamentous structures as effectively as ATP. Upon MreB assembly, both GTP and ATP release the gamma phosphate at similar rates. Therefore, MreB is an equally effective ATPase and GTPase. Electron microscopy and quantitative rheology suggest that the morphologies and micromechanical properties of filamentous ATP-MreB and GTP-MreB are similar. In contrast, mammalian actin assembly is favored in the presence of ATP over GTP. These results indicate that, despite high structural homology of their monomers, T. maritima MreB and actin filaments display different assembly, morphology, micromechanics, and nucleotide-binding specificity. Furthermore, the biophysical properties of T. maritima MreB filaments, including high rigidity and propensity to form bundles, suggest a mechanism by which MreB helical structure may be involved in imposing a cylindrical architecture on rod-shaped bacterial cells

Gelation kinetics of actin filament networks in the presence of equimolar concentrations of F-actin crosslinking proteins α-actinin, and filamin.

<p>Time-dependent elastic modulus is measured using a strain-controlled rheometer. The imposed deformation amplitude to measure the elastic modulus was 1% and the shear frequency was 1 rad/s. The concentration of actin was 24 μM.</p

Aggregate Productivity Growth in Indian Manufacturing: An Application of Domar Aggregation

An attempt is made to compute the aggregate productivity growth using the Domar aggregation technique. Building up from the Total Factor Productivity Growth (TFPG) estimates for 3-digit industries, we have used Domar weights to computed total factor productivity (TFP) growth for selected 10, 2-digit industries for the period 1980-2000. [ICRIER WP no. 239].total factor productivity (TFP), TFP, Productivity growth, Domar aggregation, aggregate value added, productivity, industries, Indian manufacturing, industrial sector, Indian economy, manufacturing, Indian, manufacturing,

Non-linear rheology of F-actin networks in the presence of α-actinin, filamin or both.

<p>A and B. Time-dependent shear modulus <i>G</i>(t, γ₀) of an F-actin network in the presence of (A) both α-actinin and filamin or (B) filamin alone for low shear deformation amplitude γ₀. The modulus increases for increasing deformation amplitudes, indicating shear-induced network hardening (or stiffening). <i>Inset</i>, Time-dependent shear modulus of the same network for high deformation amplitudes. The modulus decreases for increasing deformation amplitude, indicating shear-induced network softening. C. Shear modulus of F-actin networks in the presence of α-actinin, filamin, or both as a function of deformation amplitude. The modulus is estimated at a time of 1 second.</p

Elastic modulus of F-actin networks in the presence of α-actinin, filamin or both.

<p>Steady state elastic modulus of F-actin networks in the presence of α-actinin only (black columns), both filamin and α-actinin (50:50 molar ratio; grey columns), or filamin only (blue columns). was measured using a strain-controlled rheometer. The total concentration of F-actin crosslinking proteins is indicated. The imposed deformation amplitude to measure the elastic modulus was 1% and the shear frequency was 1 rad/s. The concentration of actin was 24 μM.</p

Viscoelastic properties of F-actin networks in the presence of α-actinin, filamin or both.

<p>A. Frequency-dependent elastic modulus of F-actin networks in the presence of either 0.12 μM α-actinin, 0.12 μM filamin, or 0.06 μM α-actinin+0.06 μM filamin. The amplitude of the deformation was 1%. Elastic moduli are normalized by their value at 1 rad/s. B. Slope <i>a</i> of the elastic modulus obtained from a power-law fit of <i>G'</i>(ω)∼ω<i>^a</i> shown in panel A. C. Phase angle of F-actin networks, which compares the viscous modulus <i>G”</i> to the elastic modulus, <i>G'</i> as δ = tan⁻¹(<i>G”</i>/<i>G'</i>). The phase angle of water and F-actin without crosslinking proteins is 90° and 30°, respectively.</p

Mechanical properties of crescentin and its eukaryotic counterparts <i>in vitro</i>.

<p>Elasticity, G′, and phase angle, δ, were measured using a cone-and-plate strain-controlled rheometer, which applied oscillatory shear deformations of small 1%-amplitude and a frequency of 1 rad/s. Rheological parameters G′ and δ were measured at steady state, i.e. after these parameters had reached a steady state after onset of assembly. The phase angle measures the delay in the response of the stress induced in the filament networks by the rheometer. An elastic solid shows no delay (phase angle of 0°); a viscous liquid without elasticity like glycerol shows a maximum delay (phase angle of 90°). The mechanical resilience of cytoskeleton proteins is defined as the shear amplitude at which the elastic modulus started to fall (e.g. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0008855#pone-0008855-g003" target="_blank">Fig. 3</a>). Protein concentration for measurements of G′, δ, and resilience was 1 mg/ml. For the range of concentrations for the concentration-dependent elasticity, G′(c), see text and references.</p

Response of crescentin to mechanical deformation.

<p><b><i>A</i></b>, Frequency-dependent elastic modulus, G′(ω), of crescentin as different concentrations. <b><i>B</i></b>, Elasticity, G′, of crescentin as a function of strain amplitude, γ (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0008855#s2" target="_blank">Methods</a> section). Crescentin does not undergo strain-hardening, whereby G′ would increase with γ. Crescentin concentrations in panels A and B are 0.25 mg/ml (filled squares), 0.5 mg/ml (open squares), 0.75 mg/ml (filled circles), and 1 mg/ml (open circles). <b><i>C</i></b>, Recovery of the elasticity of crescentin following the application of a large short-lived shear deformation as a function of crescentin concentration. Percent recovery is defined as the ratio of the recovered elasticity after application of a couple of oscillatory shear deformations of 1000% and the initial elasticity. Elasticity was evaluated at a frequency of 1 rad/s and a strain amplitude of 1%.</p