Self-organization and mechanics of actomyosin networks attached to artificial and cellular plasma membranes

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The cortex of animal cells is a thin, highly dynamically crosslinked actin network directly connected to the plasma membrane. Even though cortical actin plays a pivotal role in cellular mechanics and morphogenesis, only very little is known about the organization of the cortex attached to the plasma membrane, its collective dynamics, and how its dynamics affects mechanical properties that eventually generate function. A profound understanding requires to study model systems that permit control over composition and architecture of the network. The overarching goal of this project is to disentangle how network architecture, dynamic attachment to the plasma membrane, and contractility contribute to the viscoelastic properties of the cellular cortex. Convergence of the mechanical behavior of artificial cortices obtained in bottom-up approaches and natural cortices derived in a top-down approach is sought-after. Starting point of the project is a recently established minimal actin cortex (MAC), which dynamically cross-links a lipid membrane doped with the receptor lipid PtdIns(4,5)P2 with F-actin through the protein ezrin. In prior work, we quantified the organization of the F-actin network as a function of ezrin pinning sites on the membrane and related the viscoelastic properties of the F-actin/membrane composite to actin organization. Based on the established MAC, we are now in the position to elucidate the influence of the ezrin cross-linker’s dynamic nature on the F-actin architecture and its viscoelastic properties using ezrin mutants with an altered F-actin binding site. We will further address the question how F-actin cross-linkers alter the organization of the actin network on the membrane and how this influences the dynamics and rheological properties of the system. We will actively drive the actin cortex out of equilibrium by adding non-muscle myosin II motors and ATP to monitor the collective behavior in the networks and quantify the associated athermal fluctuations. In comparison to the MACs, we plan to manipulate the attachment sites of natural cell membrane fragments with preserved cortices to be able to pin down the importance of the different components for mesoscopic organization of the dynamic network. The use of both active and passive microrheology will help us to investigate the effect of athermal fluctuations on the viscoelastic properties of the cortex. Nonlinear behavior of the networks will be explored by using active microrheology with externally applied high-amplitude noise. This will permit us to address the discrepancy between rheological properties of artificial networks and the soft glassy rheology found for living cell.