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1. Regulation of Actin Bundle Mechanics and Structure by Intracellular Environmental Factors

   https://doi.org/10.3389/fphy.2021.675885  

   Castaneda, Nicholas; Park, Jinho; Kang, Ellen Hyeran (May 2021, Frontiers in Physics)

   null (Ed.)

   The mechanical and structural properties of actin cytoskeleton drive various cellular processes, including structural support of the plasma membrane and cellular motility. Actin monomers assemble into double-stranded helical filaments as well as higher-ordered structures such as bundles and networks. Cells incorporate macromolecular crowding, cation interactions, and actin-crosslinking proteins to regulate the organization of actin bundles. Although the roles of each of these factors in actin bundling have been well-known individually, how combined factors contribute to actin bundle assembly, organization, and mechanics is not fully understood. Here, we describe recent studies that have investigated the mechanisms of how intracellular environmental factors influence actin bundling. This review highlights the effects of macromolecular crowding, cation interactions, and actin-crosslinking proteins on actin bundle organization, structure, and mechanics. Understanding these mechanisms is important in determining in vivo actin biophysics and providing insights into cell physiology. 

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2. Actin Bundle Nanomechanics and Organization Are Modulated by Macromolecular Crowding and Electrostatic Interactions

   https://doi.org/10.3389/fmolb.2021.760950  

   Castaneda, Nicholas; Feuillie, Cecile; Molinari, Michael; Kang, Ellen Hyeran (November 2021, Frontiers in Molecular Biosciences)

   The structural and mechanical properties of actin bundles are essential to eukaryotic cells, aiding in cell motility and mechanical support of the plasma membrane. Bundle formation occurs in crowded intracellular environments composed of various ions and macromolecules. Although the roles of cations and macromolecular crowding in the mechanics and organization of actin bundles have been independently established, how changing both intracellular environmental conditions influence bundle mechanics at the nanoscale has yet to be established. Here we investigate how electrostatics and depletion interactions modulate the relative Young’s modulus and height of actin bundles using atomic force microscopy. Our results demonstrate that cation- and depletion-induced bundles display an overall reduction of relative Young’s modulus depending on either cation or crowding concentrations. Furthermore, we directly measure changes to cation- and depletion-induced bundle height, indicating that bundles experience alterations to filament packing supporting the reduction to relative Young’s modulus. Taken together, our work suggests that electrostatic and depletion interactions may act counteractively, impacting actin bundle nanomechanics and organization. 

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3. Confinement Geometry Tunes Fascin-Actin Bundle Structures and Consequently the Shape of a Lipid Bilayer Vesicle

   https://doi.org/https://doi.org/10.3389/fmolb.2020.610277  

   Y. Bashirzadeh, N.H. Wubshet (January 2020, Frontiers in molecular biosciences)

   null (Ed.)

   Depending on the physical and biochemical properties of actin-binding proteins, actin networks form different types of membrane protrusions at the cell periphery. Actin crosslinkers, which facilitate the interaction of actin filaments with one another, are pivotal in determining the mechanical properties and protrusive behavior of actin networks. Short crosslinkers such as fascin bundle F-actin to form rigid spiky filopodial protrusions. By encapsulation of fascin and actin in giant unilamellar vesicles (GUVs), we show that fascin-actin bundles cause various GUV shape changes by forming bundle networks or straight single bundles depending on GUV size and fascin concentration. We also show that the presence of a long crosslinker, α-actinin, impacts fascin-induced GUV shape changes and significantly impairs the formation of filopodia-like protrusions. Actin bundle-induced GUV shape changes are confirmed by light-induced disassembly of actin bundles leading to the reversal of GUV shape. Our study contributes to advancing the design of shape-changing minimal cells for better characterization of the interaction between lipid bilayer membranes and actin cytoskeleton. 

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4. Liquid-like condensates that bind actin drive filament polymerization and bundling

   https://doi.org/10.1101/2024.05.04.592527  

   Walker, Caleb; Chandrasekaran, Aravind; Mansour, Daniel; Graham, Kristin; Torres, Andrea; Wang, Liping; Lafer, Eileen M; Rangamani, Padmini; Stachowiak, Jeanne C (May 2024, bioRxiv)

   Liquid-like protein condensates perform diverse physiological functions. Previous work showed that VASP, a processive actin polymerase, forms condensates that polymerize and bundle actin. To minimize their curvature, filaments accumulated at the inner condensate surface, ultimately deforming the condensate into a rod-like shape, filled with a bundle of parallel filaments. Here we show that this behavior does not require proteins with specific polymerase activity. Specifically, we found that condensates composed of Lamellipodin, a protein that binds actin but is not an actin polymerase, were also capable of polymerizing and bundling actin filaments. To probe the minimum requirements for condensate-mediated actin bundling, we developed an agent-based computational model. Guided by its predictions, we hypothesized that any condensate-forming protein that binds actin could bundle filaments through multivalent crosslinking. To test this idea, we added an actin-binding motif to Eps15, a condensate-forming protein that does not normally bind actin. The resulting chimera formed condensates that drove efficient actin polymerization and bundling. Collectively, these findings broaden the family of proteins that could organize cytoskeletal filaments to include any actin-binding protein that participates in protein condensation. 

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5. A strong nonequilibrium bound for sorting of cross-linkers on growing biopolymers

   https://doi.org/10.1073/pnas.2102881118  

   Qiu, Yuqing; Nguyen, Michael; Hocky, Glen M.; Dinner, Aaron R.; Vaikuntanathan, Suriyanarayanan (September 2021, Proceedings of the National Academy of Sciences)

   Understanding the role of nonequilibrium driving in self-organization is crucial for developing a predictive description of biological systems, yet it is impeded by their complexity. The actin cytoskeleton serves as a paradigm for how equilibrium and nonequilibrium forces combine to give rise to self-organization. Motivated by recent experiments that show that actin filament growth rates can tune the morphology of a growing actin bundle cross-linked by two competing types of actin-binding proteins [S. L. Freedman et al. , Proc. Natl. Acad. Sci. U.S.A. 116, 16192–16197 (2019)], we construct a minimal model for such a system and show that the dynamics of a growing actin bundle are subject to a set of thermodynamic constraints that relate its nonequilibrium driving, morphology, and molecular fluxes. The thermodynamic constraints reveal the importance of correlations between these molecular fluxes and offer a route to estimating microscopic driving forces from microscopy experiments. 

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