2- and 3-dimensional synthetic large-scale de novo patterning by mammalian cells through phase separation

Abstract

Synthetic biology provides an opportunity for the construction and exploration of alternative solutions to biological problems - solutions different from those chosen by natural life. To this end, synthetic biologists have built new sensory systems, cellular memories, and alternative genetic codes. There is a growing interest in applying synthetic approaches to multicellular systems, especially in relation to multicellular self-organization. Here we describe a synthetic biological system that confers large-scale de novo patterning activity on 2-D and 3-D populations of mammalian cells. Instead of using the reaction-diffusion mechanisms common in real embryos, our system uses cadherin-mediated phase separation, inspired by the known phenomenon of cadherin-based sorting. An engineered self-organizing, large-scale patterning system requiring no prior spatial cue may be a significant step towards the construction of self-assembling synthetic tissues.

Engineering multicellular systems: Using synthetic biology to control tissue self-organization

“…Activating genes involved in differentiation, proliferation, and secretion [23] could contribute to the functional aspect of the engineered tissue. For example, differential expression of cadherin family adhesion molecules has been shown to enable formation of spatial patterns based, at least in part, on differential adhesion [24,25]. Implementation of this approach could lead to the creation of more complex morphogenesis and dynamically evolving structures.…”

“…More complex intracellular signal processing like memory and recombinase-based networks are available [60], and will be interesting to see if they can be coupled to extracellular sensing in multicellular contexts as they have been implicated in driving tissue development. Finally, responses could also result in changes to cell shape or to mechanical properties such as cell adhesion; in a recent example, drug-inducible expression of cadherin molecules in epithelial monolayer has been used to generate 2D and 3D synthetic patterning based on phase separation [24]. The implementation of these responses under control of synthetic pathways will be important for the implementation and the study of more complex morphogenetic trajectories.…”

Self-organizing multicellular structures designed using synthetic biology

“…It has been more challenging to find evidence that meltwater from the break-up of the Cordilleran Ice Sheet freshened the North Pacific Ocean. Near-coastal sediments in the northeast Pacific reveal that large abundances of freshwater biota were transported to the region by glacial-era meltwater 6 , and glacial debris has been uncovered in the region that can be associated with some Heinrich events 7 .…”

“…Cadherin proteins mediate cell-cell adhesion, and so are essential for holding cells together and creating tissue boundaries during development 4 . Much like oil and water, cell populations that have different patterns or levels of cadherins can sort themselves into separate groups after being mixed together, and can self-assemble into a range of structures in vitro 5,6 . To create a synthetic program to guide shape formation, Toda et al built several genetic circuits composed of different synNotch sensors that, when activated by a neighbouring cell, drive the expression not only of different levels or types of cadherin, but also of different ligands to bind to other sensors.…”

Using synthetic biology to explore principles of development

“…Living cells of different adhesiveness separate rather like oil and water in an unconstrained system (Steinberg, 1970;Foty and Steinberg, 2005), so it is possible that these, too, might generate complex patterns under constraint. To test this idea, Cachat et al (2016) constructed two populations of the poorly adhesive HEK239 human cell line, which could be induced by tetracycline to express E-cadherin and P-cadherin, respectively, and which were also labelled with fluorescent proteins. Cultured in the absence of tetracycline, the cells mixed statistically randomly but, on tetracycline induction, they underwent phase separation to form spots or patches, depending on the ratio of cells (Fig.…”