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1. Cofilin drives rapid turnover and fluidization of entangled F-actin

   https://doi.org/10.1073/pnas.1818808116  

   McCall, Patrick M.; MacKintosh, Frederick C.; Kovar, David R.; Gardel, Margaret L. (June 2019, Proceedings of the National Academy of Sciences)

   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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2. Agent‐Based Computational Modeling of the Stochastic Dynamic Behavior of Actin Filaments Recapitulates the Homeostatic Cortical Array in Plant Epidermal Cells

   https://doi.org/10.1002/cm.70014  

   Kim, June Hyung; Zhang, Weiwei; Iyer‐Pascuzzi, Anjali; Staiger, Christopher J; Kim, Taeyoon (June 2026, Cytoskeleton)

   ABSTRACT The homeostatic cortical actin array in plant cells plays important roles in fundamental processes, including intracellular transport, secretion, cell expansion, and cytoplasmic streaming. In response to diverse chemical and mechanical signals, the cortical array can remodel within minutes to assume new configurations or altered filament abundance. The homeostatic cortical actin array of some plant epidermal cells comprises sparsely distributed individual actin filaments and actin bundles, which enable tracking and quantitative analysis of dynamic properties over many minutes at high spatiotemporal resolution. Previous studies using quantitative live‐cell imaging, small molecule inhibitors, and genetic mutations reveal the robust dynamic steady state of the cortical actin array, with individual filaments showing a behavior termed stochastic dynamics. Compared to experimental findings, computational efforts focused on the plant actin cytoskeleton are lacking, although computational models have the potential to define underlying mechanisms of actin array homeostasis and remodeling. Here, we used an agent‐based computational model to reproduce the stochastic dynamic behavior of individual actin filaments in epidermal cells with consideration of key governing factors, including the nucleation, polymerization, depolymerization, severing, capping, and branching of filaments. Our model was able to reproduce experimental observations with respect to the abundance and length of filaments as well as the rates or frequencies of dynamic events. This model can be used to study the role of myosin motors and other actin‐binding proteins, as well as the effects of signaling events and fluxes in cellular second messengers, on actin dynamics in plant cells. 

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3. A generalized Flory-Stockmayer kinetic theory of connectivity percolation and rigidity percolation of cytoskeletal networks

   https://doi.org/10.1371/journal.pcbi.1010105  

   Bueno, Carlos; Liman, James; Schafer, Nicholas P.; Cheung, Margaret S.; Wolynes, Peter G. (May 2022, PLOS Computational Biology)

   Gov, Nir (Ed.)

   Actin networks are essential for living cells to move, reproduce, and sense their environments. The dynamic and rheological behavior of actin networks is modulated by actin-binding proteins such as α-actinin, Arp2/3, and myosin. There is experimental evidence that actin-binding proteins modulate the cooperation of myosin motors by connecting the actin network. In this work, we present an analytical mean field model, using the Flory-Stockmayer theory of gelation, to understand how different actin-binding proteins change the connectivity of the actin filaments as the networks are formed. We follow the kinetics of the networks and estimate the concentrations of actin-binding proteins that are needed to reach connectivity percolation as well as to reach rigidity percolation. We find that Arp2/3 increases the actomyosin connectivity in the network in a non-monotonic way. We also describe how changing the connectivity of actomyosin networks modulates the ability of motors to exert forces, leading to three possible phases of the networks with distinctive dynamical characteristics: a sol phase, a gel phase, and an active phase. Thus, changes in the concentration and activity of actin-binding proteins in cells lead to a phase transition of the actin network, allowing the cells to perform active contraction and change their rheological properties. 

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4. A mechanism with severing near barbed ends and annealing explains structure and dynamics of dendritic actin networks

   https://doi.org/10.7554/eLife.69031  

   Holz, Danielle; Hall, Aaron R; Usukura, Eiji; Yamashiro, Sawako; Watanabe, Naoki; Vavylonis, Dimitrios (June 2022, eLife)

   Single molecule imaging has shown that part of actin disassembles within a few seconds after incorporation into the dendritic filament network in lamellipodia, suggestive of frequent destabilization near barbed ends. To investigate the mechanisms behind network remodeling, we created a stochastic model with polymerization, depolymerization, branching, capping, uncapping, severing, oligomer diffusion, annealing, and debranching. We find that filament severing, enhanced near barbed ends, can explain the single molecule actin lifetime distribution, if oligomer fragments reanneal to free ends with rate constants comparable to in vitro measurements. The same mechanism leads to actin networks consistent with measured filament, end, and branch concentrations. These networks undergo structural remodeling, leading to longer filaments away from the leading edge, at the +/-35° orientation pattern. Imaging of actin speckle lifetimes at sub-second resolution verifies frequent disassembly of newly-assembled actin. We thus propose a unified mechanism that fits a diverse set of basic lamellipodia phenomenology. 

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5. Single-molecule analysis of actin filament debranching by cofilin and GMF

   https://doi.org/10.1073/pnas.2115129119  

   Chung, Johnson; Goode, Bruce L.; Gelles, Jeff (July 2022, Proceedings of the National Academy of Sciences)

   Eukaryotic cells contain branched actin networks that are essential for endocytosis, motility, and other key cellular processes. These networks, which are formed by filamentous actin and the Arp2/3 complex, must subsequently be debranched to allow network remodeling and to recycle the Arp2/3 complex. Debranching appears to be catalyzed by two different members of the actin depolymerizing factor homology protein family: cofilin and glial maturation factor (GMF). However, their mechanisms of debranching are only partially understood. Here, we used single-molecule fluorescence imaging of Arp2/3 complex and actin filaments under physiological ionic conditions to observe debranching by GMF and cofilin. We demonstrate that cofilin, like GMF, is an authentic debrancher independent of its filament-severing activity and that the debranching activities of the two proteins are additive. While GMF binds directly to the Arp2/3 complex, cofilin selectively accumulates on branch–junction daughter filaments in tropomyosin-decorated networks just prior to debranching events. Quantitative comparison of debranching rates with the known kinetics of cofilin–actin binding suggests that cofilin occupancy of a particular single actin site at the branch junction is sufficient to trigger debranching. In rare cases in which the order of departure could be resolved during GMF- or cofilin-induced debranching, the Arp2/3 complex left the branch junction bound to the pointed end of the daughter filament, suggesting that both GMF and cofilin can work by destabilizing the mother filament–Arp2/3 complex interface. Taken together, these observations suggest that GMF and cofilin promote debranching by distinct yet complementary mechanisms. 

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