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Reversible stress softening of actin networks.

ABSTRACT: The mechanical properties of cells play an essential role in numerous physiological processes. Organized networks of semiflexible actin filaments determine cell stiffness and transmit force during mechanotransduction, cytokinesis, cell motility and other cellular shape changes. Although numerous actin-binding proteins have been identified that organize networks, the mechanical properties of actin networks with physiological architectures and concentrations have been difficult to measure quantitatively. Studies of mechanical properties in vitro have found that crosslinked networks of actin filaments formed in solution exhibit stress stiffening arising from the entropic elasticity of individual filaments or crosslinkers resisting extension. Here we report reversible stress-softening behaviour in actin networks reconstituted in vitro that suggests a critical role for filaments resisting compression. Using a modified atomic force microscope to probe dendritic actin networks (like those formed in the lamellipodia of motile cells), we observe stress stiffening followed by a regime of reversible stress softening at higher loads. This softening behaviour can be explained by elastic buckling of individual filaments under compression that avoids catastrophic fracture of the network. The observation of both stress stiffening and softening suggests a complex interplay between entropic and enthalpic elasticity in determining the mechanical properties of actin networks.

SUBMITTER: Chaudhuri O

PROVIDER: S-EPMC3236682 | biostudies-literature | 2007 Jan

REPOSITORIES: biostudies-literature

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Reversible stress softening of actin networks.

Chaudhuri Ovijit O Parekh Sapun H SH Fletcher Daniel A DA

Nature 20070101 7125

The mechanical properties of cells play an essential role in numerous physiological processes. Organized networks of semiflexible actin filaments determine cell stiffness and transmit force during mechanotransduction, cytokinesis, cell motility and other cellular shape changes. Although numerous actin-binding proteins have been identified that organize networks, the mechanical properties of actin networks with physiological architectures and concentrations have been difficult to measure quantita ...[more]

PMID: 17230186

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Project description:Calcium-calmodulin-dependent kinase II (CaMKII) has an important role in dendritic spine remodeling upon synaptic stimulation. Using fluorescence video microscopy and image analysis, we investigated the architectural dynamics of rhodamine-phalloidin stabilized filamentous actin (F-actin) networks cross-linked by CaMKII. We used automated image analysis to identify F-actin bundles and crossover junctions and developed a dimensionless metric to characterize network architecture. Similar networks were formed by three different CaMKII species with a 10-fold length difference in the linker region between the kinase domain and holoenzyme hub, implying linker length is not a primary determinant of F-actin cross-linking. Electron micrographs showed that at physiological molar ratios, single CaMKII holoenzymes cross-linked multiple F-actin filaments at random, whereas at higher CaMKII/F-actin ratios, filaments bundled. Light microscopy established that the random network architecture resisted macromolecular crowding with polyethylene glycol and blocked ATP-powered compaction by myosin-II miniature filaments. Importantly, the networks disassembled after the addition of calcium-calmodulin and were then spaced within 3 min into compacted foci by myosin motors or more slowly (30 min) aggregated by crowding. Single-molecule total internal reflection fluorescence microscopy showed CaMKII dissociation from surface-immobilized globular actin exhibited a monoexponential dwell-time distribution, whereas CaMKII bound to F-actin networks had a long-lived fraction, trapped at crossover junctions. Release of CaMKII from F-actin, triggered by calcium-calmodulin, was too rapid to measure with flow-cell exchange (<20 s). The residual bound fraction was reduced substantially upon addition of an N-methyl-D-aspartate receptor peptide analog but not ATP. These results provide mechanistic insights to CaMKII-actin interactions at the collective network and single-molecule level. Our findings argue that CaMKII-actin networks in dendritic spines maintain spine size against physical stress. Upon synaptic stimulation, CaMKII is disengaged by calcium-calmodulin, triggering network disassembly, expansion, and subsequent compaction by myosin motors with kinetics compatible with the times recorded for the poststimulus changes in spine volume.

Dataset Information

Reversible stress softening of actin networks.

Publications

Reversible stress softening of actin networks.

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