Stress-Dependent Elasticity of Composite Actin Networks as a Model for Cell Behavior

Gardel, M. L.; Nakamura, Fumihiko; Hartwig, John H.; Crocker, John C.; Stossel, Thomas P.; Weitz, David A.

doi:10.1103/physrevlett.96.088102

Cited by 147 publications

(148 citation statements)

References 21 publications

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“…As the secondary purse string matures, it comes under increased internal load and the rate of closure slows. This change in internal load may be due to strain stiffening as has been observed in purified solutions of actin, actin plus cross-linkers and actin plus motors (Gardel et al, 2004;Gardel et al, 2006;Mizuno et al, 2007). By this hypothesis, components of the contractile and junctional apparatus would rearrange themselves in response to mechanical forces that drive closure.…”

Section: Discussionmentioning

confidence: 99%

Actomyosin purse strings: Renewable resources that make morphogenesis robust and resilient

et al. 2008

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Section: Discussionmentioning

confidence: 99%

Actomyosin purse strings: Renewable resources that make morphogenesis robust and resilient

et al. 2008

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“…Such a power-law form has no characteristic times at all. In contrast, the low frequency behavior of reconstituted gels is either purely elastic with β = 0 (Janmey, 1990) or nearly so with a very small β exponent (Xu, 1998;Gardel, 2006).…”

Section: Introductionmentioning

confidence: 96%

Multiple‐Particle Tracking and Two‐Point Microrheology in Cells

Crocker

Hoffman

2007

Methods in Cell Biology

195

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Mechanical stress and stiffness are increasingly recognized to play important roles in numerous cell biological processes, notably cell differentiation and tissue morphogenesis. Little definite is known, however, about how stress propagates through different cell structures or how it is converted to biochemical signals via mechanotransduction, due in large part to the difficulty of interpreting many cell mechanics experiments. A newly developed technique, two-point microrheology (TPM), can provide highly interpretable, quantitative measurements of cells' frequency-dependent shear moduli and spectra of their fluctuating intracellular stresses. TPM is a non-invasive method based on measuring the Brownian motion of large numbers of intracellular particles using multiple particle tracking. While requiring only hardware available in many cell biology laboratories-a phase microscope and digital video camera, as a statistical technique, it also requires the automated analysis of many thousands of micrographs. Here we describe in detail the algorithms and software tools used for such large-scale multiple particle tracking, as well as common sources of error and the microscopy methods needed to minimize them. Moreover, we describe the physical principles behind TPM and other passive microrheology methods, their limitations, and typical results for cultured epithelial cells. ABSTRACT Mechanical stress and stiffness are increasingly recognized to play important roles in numerous cell biological processes, notably cell differentiation and tissue morphogenesis. Little definite is known, however, about how stress propagates through different cell structures or how it is converted to biochemical signals via mechanotransduction, due in large part to the difficulty of interpreting many cell mechanics experiments. A newly developed technique, two-point microrheology (TPM), can provide highly interpretable, quantitative measurements of cells' frequency-dependent shear moduli and spectra of their fluctuating intracellular stresses. TPM is a non-invasive method based on measuring the Brownian motion of large numbers of intracellular particles using multiple particle tracking. While requiring only hardware available in many cell biology laboratories-a phase microscope and digital video camera, as a statistical technique, it also requires the automated analysis of many thousands of micrographs. Here we describe in detail the algorithms and software tools used for such large-scale multiple particle tracking, as well as common sources of error and the microscopy methods needed to minimize them. Moreover, we describe the physical principles behind TPM and other passive microrheology methods, their limitations, and typical results for cultured epithelial cells.

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“…Because filamin and ␣-actinin share a similar actin-binding domain, this result suggests that the regulation of the actin cytoskeleton by the ABPs may be governed not only by binding kinetics but also by the actual mechanical structure of the ABP. Moreover, rheological measurements have shown that above some critical load, the network displays substantial softening, suggesting a major disruption of the actin network, possibly because of filament rupture, filament buckling, unfolding of APBs, or unbinding of ABPs (16,30). Although these in vitro network studies provide valuable information about the general behavior of the actin cytoskeleton, a more detailed description at a molecular level is required to better understand how the cell regulates the cytoskeletal machinery.…”

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confidence: 99%

“…To understand the complex mechanical properties exhibited by the cell (21)(22)(23)(24), the primary focus has been directed toward studies of reconstituted actin networks grown in vitro in the presence or absence of ABPs (8,16,17,(25)(26)(27)(28)(29)(30)(31). For example, in one study, actin networks cross-linked by filamin were shown to exhibit dramatically different mechanical properties from networks formed by ␣-actinin (17).…”

mentioning

confidence: 99%

“…Several of these ABPs, including filamin, ␣-actinin, fimbrin, spectrin, and dystrophin, have a conserved actin-binding domain, but their overall structure and function are quite different (7,(9)(10)(11)(12)(13)(14). For instance, cross-linking proteins, such as filamin, promote the formation of a cortical load-bearing isotropic F-actin network at low concentrations near the plasma membrane (10,(15)(16)(17). Conversely, bundling proteins, including ␣-actinin, form thick F-actin cables that help both in maintaining the cell under a prestressed state (1,18) and generating protrusions at the leading edge of a migrating cell (11,19).…”

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confidence: 99%

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Measuring molecular rupture forces between single actin filaments and actin-binding proteins

Ferrer¹,

Lee

Chen

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

194

166

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Actin-binding proteins (ABPs) regulate the assembly of actin filaments (F-actin) into networks and bundles that provide the structural integrity of the cell. Two of these ABPs, filamin and ␣-actinin, have been extensively used to model the mechanical properties of actin networks grown in vitro; however, there is a lack in the understanding of how the molecular interactions between ABPs and F-actin regulate the dynamic properties of the cytoskeleton. Here, we present a native-like assay geometry to test the rupture force of a complex formed by an ABP linking two quasiparallel actin filaments. We readily demonstrate the adaptability of this assay by testing it with two different ABPs: filamin and ␣-actinin. For filamin/actin and ␣-actinin/actin, we measured similar rupture forces of 40 -80 pN for loading rates between 4 and 50 pN/s. Both ABP unfolding and conformational transition events were observed, demonstrating that both are important and may be a significant mechanism for the temporal regulation of the mechanical properties of the actin cytoskeleton. With this modular, singlemolecule assay, a wide range of ABP/actin interactions can be studied to better understand cytoskeletal and cell dynamics.␣-actinin ͉ filamin ͉ optical tweezers ͉ single-molecule force spectroscopy

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Stress-Dependent Elasticity of Composite Actin Networks as a Model for Cell Behavior

Cited by 147 publications

References 21 publications

Actomyosin purse strings: Renewable resources that make morphogenesis robust and resilient

Actomyosin purse strings: Renewable resources that make morphogenesis robust and resilient

Multiple‐Particle Tracking and Two‐Point Microrheology in Cells

Measuring molecular rupture forces between single actin filaments and actin-binding proteins

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