Reconstitution of actin polymerization via artificial cells

Abstract

Unilamellar liposomes are sphere-shaped vesicles which are artificially created from natural phospholipids and/or cholesterol to mimic the composition of bilayer plasma membrane. Giant Unilamellar Vesicles (GUVs) are enclosed compartments or “microreactors” which offers a way to study the effect of confinement on cellular systems. Here we demonstrated several preparation approaches of GUVs, namely water-in-oil droplets with microfluidics apparatus, one-pot assembly water-in-oil droplets, GUVs with gentle hydration method and GUVs with reverse emulsion method. Specifically, we highlighted the advantages and challenges of each approach in reconstituting actin cytoskeleton, in an attempt to mimic the interaction between actin and actin-binding proteins – AtFH1, AtVLN4, XopR, Sac6 – in the cell-like vesicles. Moreover, we had also showed that we could vary the lipid compositions to create artificial cell of desired properties, as well as manipulating the proteins-lipids interaction on the membrane interface. Overall, we established capabilities of reconstituting actin polymerization inside an artificial cell-like confinement.