Protein Delivery to Eukaryotic Cells by Engineered Bacteria
Synthetic biologists engineer genetic circuits for applications ranging from biosynthesis to biotherapeutics. Although the application of engineering strategies such as standardization, abstraction, and modularity has long been highlighted as the path to designing complex biological systems, early work generally relied on an ad hoc strategy, limiting applications to relatively simple systems. More recently, several groups have explicitly applied modular design to the development of biosynthetic pathways, biological computation, and increasingly sophisticated logic functions. However, connection of distinct functions to create a useful system-level behavior remains a key challenge. We assessed a possibility and limitation of a modular design when engineering a complex biological system.
By applying a modular design, we engineered E. coli to deliver macromolecules to the cytoplasm of cancer cells in vitro. Fabrication, testing, composition, and troubleshooting of five functional modules produced an efficient system capable of delivering proteins to over 80 % of targeted cancer cells. The modular design strategy enabled facile system modification for both troubleshooting and optimization. These devices were then mixed and matched to build a new type of delivery device that enabled E. coli to escape from the vacuole and secrete payloads. The delivery system was then further modified to deliver payloads to other eukaryotic organisms.
Successful application of modular design to the delivery system demonstrates that abstraction is a simple yet powerful tool for making the design of complex biological systems tractable. We expect that continued refinement of modular design, including incorporation of relevant quantitative information, will enable construction of increasingly complex systems. We envision that the bacterial delivery system developed here may itself become a high level module that can be incorporated into the design of more complex systems, such as a therapeutic bacterium delivering cancer-cell specific microRNA.