Mechanical properties of reconstituted actin networks at an oil–water interface determined by microrheology

Abstract

There have been various attempts to investigate the mechanical properties of the actin cortex in cells, but the factors that control them remain poorly understood. To make progress, we develop a reconstituted model of the actin cortex that mimics its structure. We attach actin filaments to lipids lining the surface of an oil droplet using biotin–streptavidin bonds. In this way we can form a thin actin network that can be visualized and studied by confocal microscopy. Our approach allows incorporation of different actin-binding and motor proteins into this 2D network and characterization of their effect on its mechanical properties in a quantitative way. To study the viscoelasticity of the network, we use passive particle tracking microrheology, which allows storage and loss moduli to be extracted from the mean square displacement of tracer particles. We show that adding cross-linkers to the cortex increases its elasticity by several orders of magnitude and addition of myosin in the presence of ATP results in a strong and rapid stiffening of the network. This approach opens up a variety of possibilities to study viscoelastic properties of the actin cortex in vitro, allowing incorporation of any protein of interest into the system.