Ever since the beginning of life, compartmentalization has been playing a crucial role in biological systems. The famous Miller-Urey experiment shows that inorganic molecules can be transformed into organic substances under extreme conditions, catalyzed by, for example, lightnings. However, homogeneously distributed organic matter is not enough for life to emerge. It is almost impossible that all conditions are appropriate for life in the entire primordial soup.That is where the compartments come in.

Only after coacervate droplets form and organic molecules condense inside, can a completely different environment be attained within, thus enabling the emergence of bio-macromolecules, or in other words, making life possible.In higher cells, compartmentalization is mainly achieved by different organelles, i.e. mitochondria, chloroplasts, lysosomes, etc. They play three major roles: A, B, C.

ntuitively, for an organelle to remain a stable compartment, it must acquire a material boundary, or more precisely, a membrane. Membrane-bound organelles are ind eed common and stable, but from the perspective of synthesis, they are too complicated for primordial conditions. However, there are also non-membrane-bound organelles, for instance, stress granules, P granules and nucleoli. More importantly, their formation is guided by simple physical principles. Membrane-less organelles and phase separation. Next came the question how can we synthase membrane-less organelles.

There are a large number of phase separation phenomena in cells, which can be summarized by the principle that interaction and multivalency are two preconditions of phase separation in cells. Based on this principle, we used SUMO-SIM, FKBP-Frb, and similar interacting pairs as interactional modules to provide diverse induction of the condensation, while we fused homo-oligomeric tags (HOTags) to introduce multivalency. We named our system SPOT (Synthetic Phase separation-based Organelle Platform) because it can form granules in yeast (we can see fluorescent spots in yeast under the microscope).

We tested different interactional modules to construct the synthetic organelles and then modeled our system according to the theory of phase separation. As this model predicts, different promoters alter the features and kinetics of our system, which was also validated by the experiments.

We verified the feasibility of several potential functions, both theoretically and experimentally, including reaction compartment, sensor, etc. In the future, by replacing functional modules with other parts, this system can be reprogrammed to conduct functions not included in the current project.