Vesicles serve important functions in the construction of artificial cells. They facilitate biochemical reactions by confining reactants and products in space, and delineate the boundaries of the protocell. They allow concentration gradients to form, and control the passage of molecules via embedded proteins. However, to date, manufacturing strategies have focussed on uni-compartmental structures, resulting in vesicles with homogenous internal content. This is in contrast to real cells which have spatial segregation of components and processes. We bridge this divide by fabricating networked multi-compartment vesicles. These were generated by encasing multiple water-in-oil droplets with an external bilayer, using a process of gravity-mediated phase-transfer. We were able to control the content of the compartments, and could define the vesicle architecture by varying the number of encased droplets. We demonstrated the bilayers were biologically functional by inserting protein channels, which facilitated material transfer between the internal compartments themselves, and between the compartments and their external environment. This paves the way for the construction of inter- and intra-vesicle communication networks. Importantly, multi-compartment vesicles allow the spatio-dynamic organisation seen in real cells to be introduced into artificial ones for the first time.
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