Artificial Symmetry-Breaking for Morphogenetic Engineering Bacterial Colonies

Abstract

Morphogenetic engineering is an emerging field that explores the design and implementation of self-organized patterns, morphologies, and architectures in systems composed of multiple agents such as cells and swarm robots. Synthetic biology, on the other hand, aims to develop tools and formalisms that increase reproducibility, tractability, and efficiency in the engineering of biological systems. We seek to apply synthetic biology approaches to the engineering of morphologies in multicellular systems. Here, we describe the engineering of two mechanisms, symmetry-breaking and domain-specific cell regulation, as elementary functions for the prototyping of morphogenetic instructions in bacterial colonies. The former represents an artificial patterning mechanism based on plasmid segregation while the latter plays the role of artificial cell differentiation by spatial colocalization of ubiquitous and segregated components. This separation of patterning from actuation facilitates the design-build-test-improve engineering cycle. We created computational modules for CellModeller representing these basic functions and used it to guide the design process and explore the design space in silico. We applied these tools to encode spatially structured functions such as metabolic complementation, RNAPT7 gene expression, and CRISPRi/Cas9 regulation. Finally, as a proof of concept, we used CRISPRi/Cas technology to regulate cell growth by controlling methionine synthesis. These mechanisms start from single cells enabling the study of morphogenetic principles and the engineering of novel population scale structures from the bottom up.

Intercellular adhesion promotes clonal mixing in growing bacterial populations

“…As a prototypic single self-recognizing protein, it has proved an attractive method of generating intercellular adhesion [ 45 ]. Here, we combined adhesion with an artificial patterning system based on segregating incompatible plasmids [ 26 , 46 ]. This technique transforms cells with multiple plasmids that can be rendered incompatible, resulting in plasmid segregation at division.…”

Using Synthetic Biology to Engineer Spatial Patterns

“…[40] Finally, the understanding of how physico-chemical laws determine patterning (for example during bacterial colony formation, or in adhesion-driven phase separation) has also benefitted from the building and examination of controllable synthetic systems. [48,51,52,[60][61][62] The ability to build, understand and modify synthetic patterns may enable their use as a tool to study, with a new perspective, not only patterning events but also other varied biological problems. For instance, stripe-forming networks have recently been employed to address questions of GRN evolution.…”

Functional channels in matureE. colicolonies

“…Upon first glance, channels resemble fractal features found in multi-strain colonies which form as a result of the mechanical instability between growth and viscous drag of dividing cells 19 . However, these features have only been reported in multi-strain colonies where the fractal dendrites have been composed of live, fluorescing cells 20–23,47 . We demonstrate that the spatial patterns we observe are different to those outlined previously.…”