BPS2026 – Expressing membrane-less RNA organelles in lipid-based synthetic cells

Bottom-up synthetic biology aims to create biomimetic synthetic cells to perform out-of-equilibrium, “life-like” functions such as growth and division. Giant unilamellar vesicles (GUVs) are good cell mimics due to having sizes similar to living cells and faithfully imitating the properties of their membranes. Recently, Fabrini et al. have developed branched RNA constructs, or “nanostars” transcribed from template DNA, which can fold and assemble co-transcriptionally via complementary kissing loop (KL) domains, creating synthetic biomolecular condensates with controlled number, size, composition, and the ability to selectively recruit proteins. Here, we demonstrate the encapsulation of transcription machinery and the expression of RNA organelles in GUVs. The so-formed synthetic cells can be programmed to produce up to three distinctly addressable organelles, by expressing RNA nanostars with orthogonal KLs. The organelles grow over time to a size dependent on the volume of the GUV and can be made to interact with the membranes thanks to selective adhesive moieties. These moieties can be tuned to vary the interaction strength between the membrane and the RNA organelles. This platform could be used to replicate key behaviors of biological membrane-less organelles in synthetic cells.