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The shape of most animal cells is controlled by the actin cortex, a thin network of dynamic actin filaments (F-actin) situated just beneath the plasma membrane. The cortex is held far from equilibrium by both active stresses and polymer turnover: Molecular motors drive deformations required for cell morphogenesis, while actin-filament disassembly dynamics relax stress and facilitate cortical remodeling. While many aspects of actin-cortex mechanics are well characterized, a mechanistic understanding of how nonequilibrium actin turnover contributes to stress relaxation is still lacking. To address this, we developed a reconstituted in vitro system of entangled F-actin, wherein the steady-state length and turnover rate of F-actin are controlled by the actin regulatory proteins cofilin, profilin, and formin, which sever, recycle, and assemble filaments, respectively. Cofilin-mediated severing accelerates the turnover and spatial reorganization of F-actin, without significant changes to filament length. We demonstrate that cofilin-mediated severing is a single-timescale mode of stress relaxation that tunes the low-frequency viscosity over two orders of magnitude. These findings serve as the foundation for understanding the mechanics of more physiological F-actin networks with turnover and inform an updated microscopic model of single-filament turnover. They also demonstrate that polymer activity, in the form of ATP hydrolysis on F-actin coupled to nucleotide-dependent cofilin binding, is sufficient to generate a form of active matter wherein asymmetric filament disassembly preserves filament number despite sustained severing.
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This content will become publicly available on March 24, 2026
Gelsolin-mediated actin filament severing, mechanics, and conformational changes at neutral and acidic pH
Gelsolin is a calcium (Ca2+) dependent, pH sensitive actin-binding protein that regulates actin filament dynamics to remodel the actin cytoskeleton. It is known that gelsolin binding induces conformational changes of actin filaments, leading to filament severing. However, the influence of physiological conditions, such as pH variations, on gelsolin-mediated filament severing activities, mechanics and conformations remains unclear despite their role in actin-actin interactions. Using Total Internal Reflection Fluorescence (TIRF) microscopy imaging and pyrene fluorescence assays, we demonstrate that filament severing efficiencies by gelsolin are enhanced in acidic conditions. In addition, analysis of filament thermal fluctuations using TIRF reveals that gelsolin binding stiffens actin filaments. Furthermore, we show that gelsolin binding induces conformational changes in filaments by measuring the filament half-pitch using high resolution Atomic Force Microscopy imaging. Together, our results suggest that pH modulation plays a key role in gelsolin-mediated filament severing activities, bending mechanics, and conformational changes, which have implications in many cellular processes including cell motility and morphogenesis.
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- Award ID(s):
- 1943266
- PAR ID:
- 10618340
- Publisher / Repository:
- Frontiers
- Date Published:
- Journal Name:
- Frontiers in Soft Matter
- Volume:
- 5
- ISSN:
- 2813-0499
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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