Artificial cells with all-aqueous droplet-in-droplet structures for spatially separated transcription and translation

The design of functional artificial cells involves compartmentalizing biochemical processes to mimic cellular organization. To emulate the complex chemical systems in biological cells, it is necessary to incorporate an increasing number of cellular functions into single compartments. Artificial organelles that spatially segregate reactions inside artificial cells will be beneficial in this context by rectifying biochemical pathways. Here, we develop artificial cells with all-aqueous droplet-in-droplet structures that separate transcription and translation processes like the nucleus and cytosol in eukaryotic cells. This architecture uses protein-based inner droplets and aqueous two-phase outer compartments, stabilized by colloidal emulsifiers. The inner droplet is designed to enrich DNA and RNA polymerase for transcription, coupled to translation at the outer droplet via mRNA-mediated cascade reactions. We show that these processes proceed independently within each compartment, maintaining genotype-phenotype correspondence. This approach provides a practical tool for exploring complex systems of artificial organelles within large ensembles of artificial cells.