Actin bundle architecture and mechanics regulate myosin II force generation

The actin cytoskeleton is a soft, structural material that underlies biological processes such as cell division, motility, and cargo transport. The cross-linked actin filaments self-organize into a myriad of architectures, from disordered meshworks to ordered bundles, which are hypothesized to control the actomyosin force generation that regulates cell migration, shape, and adhesion. Here, we use fluorescence microscopy and simulations to investigate how actin bundle architectures with varying polarity, spacing, and rigidity impact myosin II dynamics and force generation. Microscopy reveals that mixed polarity bundles formed by rigid cross-linkers support slow, bidirectional myosin II filament motion, punctuated by periods of stalled motion. Simulations reveal that these locations of stalled myosin motion correspond to sustained, high forces in regions of balanced actin filament polarity. By contrast, mixed polarity bundles formed by compliant, large cross-linkers support fast, bidirectional motion with no traps. Simulations indicate that trap duration is directly related to force magnitude, and that the observed increased velocity corresponds to lower forces resulting from both the increased bundle compliance and filament spacing. Our results indicate that the properties of actin structures regulate the dynamics and magnitude of myosin II forces, highlighting the importance of architecture and mechanics in regulating forces in biological materials.

Synchronized oscillations, metachronal waves, and jammed clusters in sterically interacting active filament arrays

Autonomous active, elastic filaments that interact with each other to achieve cooperation and synchrony underlie many critical functions in biology. A striking example is ciliary arrays in the mammalian respiratory tract; here individual filaments communicate through direct interactions and through the surrounding fluid to generate metachronal traveling waves crucial for mucociliary clearance. The mechanisms underlying this collective response and the essential ingredients for stable synchronization remain a mystery. In this article, we describe Brownian dynamics simulations of multi-filament arrays, demonstrating that short-range steric inter-filament interactions and surface-roughness are sufficient to generate a rich variety of collective spatiotemporal oscillatory, traveling and static patterns. Starting from results for the collective dynamics of two- and three-filament systems, we identify parameter ranges in which inter-filament interactions lead to synchronized oscillations. We then study how these results generalize to large one-dimensional arrays of many interacting filaments. The phase space characterizing the multi-filament array dynamics and deformations exhibits rich behaviors, including oscillations and traveling metachronal waves, depending on the interplay between geometric spacing between filaments, activity, and elasticity of the filaments. Interestingly, the existence of metachronal waves is nonmonotonic with respect to the inter-filament spacing. We also find that the degree of filament surface roughness significantly affects the dynamics — roughness on scales comparable to the filament thickness generates a locking-mechanism that transforms traveling wave patterns into statically stuck and jammed configurations. Our simulations suggest that short-ranged steric inter-filament interactions are sufficient and perhaps even critical for the development, stability and regulation of collective patterns.

Elastohydrodynamical instabilities of active filaments, arrays and carpets analyzed using slender body theory

ABSTRACTThe rhythmic motions and wave-like planar oscillations in filamentous soft structures are ubiquitous in biology. Inspired by these, recent work has focused on the creation of synthetic colloid-based active mimics that can be used to move, transport cargo, and generate fluid flows. Underlying the functionality of these mimics is the coupling between elasticity, geometry, dissipation due to the fluid, and active force or moment generated by the system. Here, we use slender body theory to analyze the linear stability of a subset of these - active elastic filaments, filament arrays and filament carpets - animated by follower forces. Follower forces can be external or internal forces that always act along the filament contour. The application of slender body theory enables the accurate inclusion of hydrodynamic effects, screening due to boundaries, and interactions between filaments. We first study the stability of fixed and freely suspended sphere-filament assemblies, calculate neutral stability curves separating stable oscillatory states from stable straight states, and quantify the frequency of emergent oscillations. When shadowing effects due to the physical presence of the spherical boundary are taken into account, the results from the slender body theory differ from that obtained using local resistivity theory. Next, we examine the onset of instabilities in a small cluster of filaments attached to a wall and examine how the critical force for onset of instability and the frequency of sustained oscillations depend on the number of filaments and the spacing between the filaments. Our results emphasize the role of hydrodynamic interactions in driving the system towards perfectly in-phase or perfectly out of phase responses depending on the nature of the instability. Specifically, the first bifurcation corresponds to filaments oscillating in-phase with each other. We then extend our analysis to filamentous (line) array and (square) carpets of filaments and investigate the variation of the critical parameters for the onset of oscillations and the frequency of oscillations on the inter-filament spacing. The square carpet also produces a uniform flow at infinity and we determine the ratio of the mean-squared flow at infinity to the energy input by active forces. We conclude by analyzing the bending and buckling instabilities of a straight passive filament attached to a wall and placed in a viscous stagnant flow - a problem related to the growth of biofilms, and also to mechanosensing in passive cilia and microvilli. Taken together, our results provide the foundation for more detailed non-linear analyses of spatiotemporal patterns in active filament systems.

Cargo navigation across 3D microtubule intersections

The eukaryotic cell’s microtubule cytoskeleton is a complex 3D filament network. Microtubules cross at a wide variety of separation distances and angles. Prior studies in vivo and in vitro suggest that cargo transport is affected by intersection geometry. However, geometric complexity is not yet widely appreciated as a regulatory factor in its own right, and mechanisms that underlie this mode of regulation are not well understood. We have used our recently reported 3D microtubule manipulation system to build filament crossings de novo in a purified in vitro environment and used them to assay kinesin-1–driven model cargo navigation. We found that 3D microtubule network geometry indeed significantly influences cargo routing, and in particular that it is possible to bias a cargo to pass or switch just by changing either filament spacing or angle. Furthermore, we captured our experimental results in a model which accounts for full 3D geometry, stochastic motion of the cargo and associated motors, as well as motor force production and force-dependent behavior. We used a combination of experimental and theoretical analysis to establish the detailed mechanisms underlying cargo navigation at microtubule crossings.

Effect of Laser-arc Hybrid Welding Energy Parameters on Welding Stability

The high-speed camera was used to collect the droplet transfer pattern and arc pattern of the laser-arc hybrid welding process. Using the methods of image processing and mathematical statistics, the effects of different laser and arc power conditions on the welding stability were studied. The results show that the melting width depends on the welding current, the depth of penetration depends on the laser power, the droplet transition pattern, the actual filament spacing and the arc length determine the welding stability of the laser arc hybrid welding.