Nucleated synthetic cells with genetically driven intercompartment communication.

Eukaryotic cells are characterized by multiple chemically distinct compartments, one of the most notable being the nucleus. Within these compartments, there is a continuous exchange of information, chemicals, and signaling molecules, essential for coordinating and regulating cellular activities. One of the main goals of bottom-up synthetic biology is to enhance the complexity of synthetic cells by establishing functional compartmentalization. There is a need to mimic autonomous signaling between compartments, which in living cells, is often regulated at the genetic level within the nucleus. This advancement is key to unlocking the potential of synthetic cells as cell models and as microdevices in biotechnology. However, a technological bottleneck exists preventing the creation of synthetic cells with a defined nucleus-like compartment capable of genetically programmed intercompartment signaling events. Here, we present an approach for creating synthetic cells with distinct nucleus-like compartments that can encapsulate different biochemical mixtures in discrete compartments. Our system enables in situ protein expression of membrane proteins, enabling autonomous chemical communication between nuclear and cytoplasmic compartments, leading to downstream activation of enzymatic pathways within the cell.

Genetically programmed synthetic cells for thermo-responsive protein synthesis and cargo release.

Synthetic cells containing genetic programs and protein expression machinery are increasingly recognized as powerful counterparts to engineered living cells in the context of biotechnology, therapeutics and cellular modelling. So far, genetic regulation of synthetic cell activity has been largely confined to chemical stimuli; to unlock their potential in applied settings, engineering stimuli-responsive synthetic cells under genetic regulation is imperative. Here we report the development of temperature-sensitive synthetic cells that control protein production by exploiting heat-responsive mRNA elements. This is achieved by combining RNA thermometer technology, cell-free protein expression and vesicle-based synthetic cell design to create cell-sized capsules able to initiate synthesis of both soluble proteins and membrane proteins at defined temperatures. We show that the latter allows for temperature-controlled cargo release phenomena with potential implications for biomedicine. Platforms like the one presented here can pave the way for customizable, genetically programmed synthetic cells under thermal control to be used in biotechnology.

Engineered Bacteria as Living Biosensors in Dermal Tattoos.

Dermal tattoo biosensors are promising platforms for real-time monitoring of biomarkers, with skin used as a diagnostic interface. Traditional tattoo sensors have utilized small molecules as biosensing elements. However, the rise of synthetic biology has enabled the potential employment of engineered bacteria as living analytical tools. Exploiting engineered bacterial sensors will allow for potentially more sensitive detection across a broad biomarker range, with advanced processing and sense/response functionalities using genetic circuits. Here, the interfacing of bacterial biosensors as living analytics in tattoos is shown. Engineered bacteria are encapsulated into micron-scale hydrogel beads prepared through scalable microfluidics. These biosensors can sense both biochemical cues (model biomarkers) and biophysical cues (temperature changes, using RNA thermometers), with fluorescent readouts. By tattooing beads into skin models and confirming sensor activity post-tattooing, our study establishes a foundation for integrating bacteria as living biosensing entities in tattoos.