Reconstitution of an Actin Cortex Inside a Liposome

The composite and versatile structure of the cytoskeleton confers complex mechanical properties on cells. Actin filaments sustain the cell membrane and their dynamics insure cell shape changes. For example, the lamellipodium moves by actin polymerization, a mechanism that has been studied using simplified experimental systems. Much less is known about the actin cortex, a shell-like structure underneath the membrane that contracts for cell movement. We have designed an experimental system that mimicks the cell cortex by allowing actin polymerization to nucleate and assemble at the inner membrane of a liposome. Actin shell growth can be triggered inside the liposome, which offers a useful system for a controlled study. The observed actin shell thickness and estimated mesh size of the actin structure are in good agreement with cellular data. Such a system paves the way for a thorough characterization of cortical dynamics and mechanics.     

Mechanics of Biomimetic Liposomes Encapsulating an Actin Shell

Cell-shape changes are insured by a thin, dynamic, cortical layer of cytoskeleton underneath the plasma membrane. How this thin cortical structure impacts the mechanical properties of the whole cell is not fully understood. Here, we study the mechanics of liposomes or giant unilamellar vesicles, when a biomimetic actin cortex is grown at the inner layer of the lipid membrane via actin-nucleation-promoting factors. Using a hydrodynamic tube-pulling technique, we show that tube dynamics is clearly affected by the presence of an actin shell anchored to the lipid bilayer. The same force pulls much shorter tubes in the presence of the actin shell compared to bare membranes. However, in both cases, we observe that the dynamics of tube extrusion has two distinct features characteristic of viscoelastic materials: rapid elastic elongation, followed by a slower elongation phase at a constant rate. We interpret the initial elastic regime by an increase of membrane tension due to the loss of lipids into the tube. Tube length is considerably shorter for cortex liposomes at comparable pulling forces, resulting in a higher spring constant. The presence of the actin shell seems to restrict lipid mobility, as is observed in the corral effect in cells. The viscous regime for bare liposomes corresponds to a leakout of the internal liquid at constant membrane tension. The presence of the actin shell leads to a larger friction coefficient. As the tube is pulled from a patchy surface, membrane tension increases locally, leading to a Marangoni flow of lipids. As a conclusion, the presence of an actin shell is revealed by its action that alters membrane mechanics.     

Spreading dynamics of biomimetic actin cortices

Reconstituted systems mimicking cells are interesting tools for understanding the details of cell behavior. Here, we use an experimental system that mimics cellular actin cortices, namely liposomes developing an actin shell close to their inner membrane, and we study their dynamics of spreading. We show that depending on the morphology of the actin shell inside the liposome, spreading dynamics is either reminiscent of a bare liposome (in the case of a sparse actin shell) or of a cell (in the case of a continuous actin shell). We use a mechanical model that qualitatively accounts for the shape of the experimental curves. From the data on spreading dynamics, we extract characteristic times that are consistent with mechanical estimates. The mechanical characterization of such stripped-down experimental systems paves the way for a more complex design closer to a cell. We report here the first step in building an artificial cell and studying its mechanics.     

Unexpected Membrane Dynamics Unveiled by Membrane Nanotube Extrusion

In cell mechanics, distinguishing the respective roles of the plasma membrane and of the cytoskeleton is a challenge. The difference in the behavior of cellular and pure lipid membranes is usually attributed to the presence of the cytoskeleton as explored by membrane nanotube extrusion. Here we revisit this prevalent picture by unveiling unexpected force responses of plasma membrane spheres devoid of cytoskeleton and synthetic liposomes. We show that a tiny variation in the content of synthetic membranes does not affect their static mechanical properties, but is enough to reproduce the dynamic behavior of their cellular counterparts. This effect is attributed to an amplified intramembrane friction. Reconstituted actin cortices inside liposomes induce an additional, but not dominant, contribution to the effective membrane friction. Our work underlines the necessity of a careful consideration of the role of membrane proteins on cell membrane rheology in addition to the role of the cytoskeleton.     

Cis and Trans Cooperativity of E-Cadherin Mediates Adhesion in Biomimetic Lipid Droplets

The regulation of cell-cell adhesion is important in cell motility, tissue growth, and for the mechanical integrity of tissues. Although the role of active cytoskeleton dynamics in regulating cadherin interactions is crucial in vivo, here we present a biomimetic emulsion system to characterize the passive E-cadherin-mediated adhesion between droplets. The visualization of a three-dimensional assembly of lipid droplets, functionalized with extracellular E-cadherin domains, reveals a hierarchy of homophilic interactions. First, the high interfacial tension of droplets facilitates trans cadherin-cadherin adhesion, which is strong enough to stabilize looser than random close packing configurations. Second, fluorescence enhancement shows that adding clustering agents, such as calcium or chelating ligands, favor the lateral cis adhesion of the already bound cadherin pairs over the clustering of monomer cadherin on the surface. Finally, above a threshold cadherin and calcium concentration, the cis and trans protein interactions become strong enough to trigger and promote droplet fusion. While E-cadherin is not known to participate in cellular fusion, this mechanism is general because replacing calcium with cholesterol to cluster the cadherin-carrying lipids also promotes fusion. These results suggest that passive clustering, via calcium-induced dimerization or membrane ordering, may contribute to the reinforcement of cell-cell contacts. Alternatively, a molecular switch for fusion offers a route to mixing droplet contents and controlling their size in situ.