Engineered Control Systems for Asymmetric Cell Division in Synthetic Bacteria

Synthetic Biology aims to implement new cellular features through the design of original genetic parts and systems. A wide variety of circuit architectures have been built successfully, proving the ability of synthetic genetic parts to predictably interact with each other within a single bacterial cell [4]. The next stage of synthetic biology will involve the de novo engineering of self-organizing multicellular systems. One of the main methods found in nature for creating complex spatially distributed systems is cellular differentiation through asymmetric cell division (ACD), whereby stem cells spawn off irreversibly differentiated daughters. The presence of different cell lineages within an eukaryotic tissue allows for the partitioning of metabolic and structural tasks. Furthermore, if one or both of the daughter cells becomes incapable of dividing, controlling asymmetric division probability can lead to tissue size and shape control [5]. In complex eukaryotic organisms ACD is typically brought about by the differential expression of key genes within each cell type. Here, we engineered a novel method for generating ACD in Escherichia coli and regulating its frequency within the population. This work provides a platform for the construction of complex synthetic tissues and organisms from bacterial cells that differentiate during development of the population.

Current methods for creating differentiated cell types in synthetic bacteria utilize gene regulatory networks that allow for bistable gene expression [2, 1]. While such systems generate bimodal populations, they are very sensitive to both intrinsic and extrinsic noise sources that can randomly force transitions between states [3]. We use a novel method for creating terminally differentiated synthetic bacterial cells. Specifically, we refactored DNA shuttling mechanisms from Caulobacter crescentus

into E. coli to target plasmid DNA into one and only one daughter cell upon division. Thus, the two daughter cells will not just have different gene expression states, but will be irreversibly genetically distinct. Our findings will lead to synthetic "tissues" that can be used in complex settings.

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