Our group developed a system named PhASE, or Photo-Activated Switch in E.coli as an approach to manipulating cellular activities. Our system is based on liquid-liquid phase separation (LLPS) and the light-induced protein-protein interaction (PPI) pairs, through which we can redistribute biomolecules into artificially-induced subcellular liquid-phase micro-compartments in a spatio-temporal and reversible manner, and thus regulate enzymatic catalysis and induce cell heterogeneity.

When we were going through the previous iGEM projects trying to devise our own, we noticed that almost all the previous projects focused on engineering at molecular scale, namely nanoscale, where gene expression circuits and signal transduction pathways were modified and regulated. In this way tissues and individuals at macro scale could be adjusted. However, there were rarely any projects targeting engineering at mesoscale, or subcellular scale, a scale that includes different kinds of organelles such as mitochondria and lysosomes. Few such projects as there were, mesoscale engineering is too important to be ignored because of the importance of these organelles to compartmentalize the intracellular environment, make it well-organized, and maximize the overall efficiency of cellular activities. We immediately agreed that it would be truly awesome if we could engineer at mesoscale to manipulate cellular activities and would probably provide a novel approach to previous iGEM obstacles, and even to solutions of ever unsolved synthetic biological problems.

We decided to try this idea in E. coli first for two reasons. Firstly, E. coli is such a simple model organism that it would be easier to engineer than eukaryotic cell. Besides, the fact that E. coli has no mesoscale organelles could make it more obvious to test the effect of mesoscale engineering. Thinking about a specific plan for such engineering, we were inspired by the polymer physical conception of LLPS that has been extensively studied in biological researches in the past ten years, especially by Clifford P. Brangwynne’s work on the development of technologies for light-activated control of intracellular phase transitions. Phase separation condensate was found to be a great platform of mesoscale engineering and optical induction was a great tool for spatio-temporal and reversible regulation. This is how the idea of PhASE was conceived.

We saw a broad application prospect of PhASE, among which we finally selected two that matter the most to cells to put into practice: the control of cellular reaction efficiency and induction of cell heterogeneity. The enzymatic reaction is the main way cells perform their functions and compartmentalization makes it possible for numerous reactions to be carried out simultaneously. Cell heterogeneity is the key to cell differentiation and biological development, as well as a critical step taken by unicellular organisms towards multicellular organisms in evolution. With these two crucial biological processes successfully engineered at mesoscale in E. Coli, we anticipate our work to be a novel approach to cellular activity modification in E.coli and a starting point for more sophisticated cellular manipulation in a wealth of organisms.

References:

Li, P., Banjade, S., Cheng, H. C., Kim, S., Chen, B., Guo, L., ... & Russo, P. S. (2012). Phase transitions in the assembly of multivalent signalling proteins. Nature, 483(7389), 336.

Shin, Y., Berry, J., Pannucci, N., Haataja, M. P., Toettcher, J. E., & Brangwynne, C. P. (2017). Spatiotemporal control of intracellular phase transitions using light-activated optoDroplets. Cell, 168(1-2), 159-171.

Shin, Y., Chang, Y. C., Lee, D. S., Berry, J., Sanders, D. W., Ronceray, P., ... & Brangwynne, C. P. (2018). Liquid nuclear condensates mechanically sense and restructure the genome. Cell, 175(6), 1481-1491.