Reconstituting an active cytoskeleton in a droplet

The actin cytoskeleton is a viscoelastic network of semiflexible actin polymers that dominates many biological processes such as cell locomotion, division, spreading, organelle transport and positioning. It is a dissipative structure that is maintained in and can switch between dynamic steady states through regulated balanced “fluxes”, i.e. spatial movements, polymerization, depolymerization, forming and breaking crosslinks. While equilibrium actin networks have been studied extensively, a dynamic cytoskeleton resembling that of living cells has been impossible to reconstruct from isolated components. We here use Xenopus laevis egg extract to reconstitute dynamic actin networks in water-in-oil emulsion droplets with sizes comparable to cells. We observe convergent steady-state 3D flow of the actin network towards the droplet center, driven by myosin and maintained by constant actin polymerization and depolymerization. Lipid vesicles and other aggregates get concentrated to form an inclusion in the center of the droplets. We constructed a hydrodynamic model of a contracting network with turn-over that can explain the measured actin velocity and density profiles and the resulting stress field, that we probe by local UV laser cutting and magnetic beads.