| When cells crawl, the protrusion of their leading edge must precede cellular adhesion and traction. However, the biochemical complexity and the dynamic morphology of living cells complicate mechanical characterization of the force-generating engine. Fortuitously, the intracellular bacterial pathogen Listeria monocytogenes have evolved to take advantage of their host cell's actin polymerization machinery to create dynamically polymerizing actin comet tails. As a simplified model of cell crawling, the actin polymerization engine of Listeria has the advantage that it does not depend on motors-driven contraction, and that its motility can be reconstituted in cytoplasmic extracts, which are more amenable to micromechanical and biochemical manipulation. By reconstituting the motility of Listeria in extracts that had been supplemented with methyl cellulose, and quantifying the mechanical environment surrounding motile Listeria using Laser Tracking Microrheology (LTM), we estimated a force-velocity relation for the Listeria engine. With an unusually biphasic relationship, forces >200 pN slowed the bacteria >20-fold. Increased mechanical power appeared to correlate with increased actin content in the tails. To probe the molecular mechanism underlying this “self-strengthening” behavior, we analyzed high-resolution trajectories of Listeria motion. Although the size of Listeria “steps” remained constant (5.2 ± 0.2 nm, n = 63), the pause duration varied inversely with the bacteria speed. Stochastic modeling showed that the biphasic force-velocity relation and constant step size depend on positive feedback loop involving nucleation and released of new actin filaments. Thus, we suggest that the actin polymerization engine is a self-organizing system, and that the saltatory movement, which depends on antagonism between propulsive force due to polymerization and friction due to binding, is a feature of all growth-driven system. |
