Exploiting heterogeneity in coupled, two plasmid systems for dynamic population adaptation

In synthetic multi-plasmid systems, it is standard to use only plasmids with orthogonal replication mechanisms to avoid phenotypic heterogeneity and ensure plasmid stability. In nature, however, microbial populations actively exploit heterogeneity to survive in fluctuating environments. Here we show that the intentional use of distinct plasmids with identical origins of replication (oris) can help an engineered bacterial population adapt to its environment. We find that copy number coupling between distinct plasmids in such systems allows for copy number buffering of an essential, but high-burden construct through the action of a stably maintained, nonessential plasmid. Plasmid coupling also generates population state memory without additional layers of regulatory control. This work reimagines how we design synthetic populations to survive and adapt by strategically giving control back to the cells.