Cracked actin filaments as mechanosensitive receptors.

Actin filament networks are exposed to mechanical stimuli, but the effect of strain on actin filament structure has not been well established in molecular detail. This is a critical gap in understanding because the activity of a variety of actin-binding proteins has recently been determined to be altered by actin filament strain. We therefore used all-atom molecular dynamics simulations to apply tensile strains to actin filaments and find that changes in actin subunit organization are minimal in mechanically strained, but intact, actin filaments. However, a conformational change disrupts the critical D-loop to W-loop connection between longitudinal neighboring subunits, which leads to a metastable cracked conformation of the actin filament whereby one protofilament is broken prior to filament severing. We propose that the metastable crack presents a force-activated binding site for actin regulatory factors that specifically associate with strained actin filaments. Through protein-protein docking simulations, we find that 43 evolutionarily diverse members of the dual zinc-finger-containing LIM-domain family, which localize to mechanically strained actin filaments, recognize two binding sites exposed at the cracked interface. Furthermore, through its interactions with the crack, LIM domains increase the length of time damaged filaments remain stable. Our findings propose a new molecular model for mechanosensitive binding to actin filaments.

Motor crosslinking augments elasticity in active nematics.

In active materials, uncoordinated internal stresses lead to emergent long-range flows. An understanding of how the behavior of active materials depends on mesoscopic (hydrodynamic) parameters is developing, but there remains a gap in knowledge concerning how hydrodynamic parameters depend on the properties of microscopic elements. In this work, we combine experiments and multiscale modeling to relate the structure and dynamics of active nematics composed of biopolymer filaments and molecular motors to their microscopic properties, in particular motor processivity, speed, and valency. We show that crosslinking of filaments by both motors and passive crosslinkers not only augments the contributions to nematic elasticity from excluded volume effects but dominates them. By altering motor kinetics we show that a competition between motor speed and crosslinking results in a nonmonotonic dependence of nematic flow on motor speed. By modulating passive filament crosslinking we show that energy transfer into nematic flow is in large part dictated by crosslinking. Thus motor proteins both generate activity and contribute to nematic elasticity. Our results provide new insights for rationally engineering active materials.

Machine learning interpretable models of cell mechanics from protein images.

Cellular form and function emerge from complex mechanochemical systems within the cytoplasm. Currently, no systematic strategy exists to infer large-scale physical properties of a cell from its molecular components. This is an obstacle to understanding processes such as cell adhesion and migration. Here, we develop a data-driven modeling pipeline to learn the mechanical behavior of adherent cells. We first train neural networks to predict cellular forces from images of cytoskeletal proteins. Strikingly, experimental images of a single focal adhesion (FA) protein, such as zyxin, are sufficient to predict forces and can generalize to unseen biological regimes. Using this observation, we develop two approaches-one constrained by physics and the other agnostic-to construct data-driven continuum models of cellular forces. Both reveal how cellular forces are encoded by two distinct length scales. Beyond adherent cell mechanics, our work serves as a case study for integrating neural networks into predictive models for cell biology.

Highly flexible PEG-LifeAct constructs act as tunable biomimetic actin crosslinkers.

In vitro studies of actin filament networks crosslinked with dynamic actin binding proteins provide critical insights into cytoskeletal mechanics as well as inspiration for new adaptive materials design. However, discontinuous variance in the physiochemical properties of actin binding proteins impedes holistic relationships between crosslinker molecular parameters, network structure, and mechanics. Bio-synthetic constructs composed of synthetic polymer backbones and actin binding motifs would enable crosslinkers with engineered physiochemical properties to directly target the desired structure-property relationships. As a proof of concept, bio-synthetic crosslinkers composed of highly flexible polyethylene glycol (PEG) polymers functionalized with the actin binding peptide LifeAct, are explored as actin crosslinkers. Using bulk rheology and fluorescence microscopy, these constructs are shown to modulate actin filament network structure and mechanics in a contour length dependent manner, while maintaining the stress-stiffening behavior inherent to actin filament networks. These results encourage the design of more diverse and complex peptide-polymer crosslinkers to interrogate and control semi-flexible polymer networks.