A synthetic system for asymmetric cell division in Escherichia coli

Abstract

We describe a synthetic genetic circuit for controlling asymmetric cell division in E. coli in which a progenitor cell creates a differentiated daughter cell while retaining its original phenotype. Specifically, we engineered an inducible system that can bind and segregate plasmid DNA to a single position in the cell. Upon cell division, co-localized plasmids are kept by one and only one of the daughter cells. The other daughter cell receives no plasmid DNA and is hence irreversibly differentiated from its sibling. In this way, we achieved asymmetric cell division through asymmetric plasmid partitioning. We then used this system to achieve physical separation of genetically distinct cells by tying motility to differentiation. Finally, we characterized an orthogonal inducible circuit that enables the simultaneous asymmetric partitioning of two plasmid species, resulting in cells that have four distinct differentiated states. These results point the way towards engineering multicellular systems from prokaryotic hosts.

Construction of intracellular asymmetry and asymmetric division in Escherichia coli

“…As high-copy plasmids also tend to localize at nucleoid-free regions 54 , 55 and were used in this study, T7 promoters that are present on plasmids should be accessible to the polarly localized T7 RNA polymerase. If the reporter constructs were to be chromosomally encoded, another layer of polar recruitment of the chromosome via the ParS/ParB system 56 would allow co-localization with the polarized T7 RNA polymerase (for example, by sandwiching the T7 promoters between two ParS arrays).…”

Control of synthetic microbial consortia in time, space, and composition

“…Permanent changes at the genetic level allow for signal responses with much greater dynamic range and longterm stability than transcriptional regulation, and as a result, there have been several efforts to create a differentiation system in microbes [46]. Recently, researchers have utilized the partitioning of plasmids during cell division to create a version of microbial differentiation via asymmetric cell division, creating daughter cells lacking plasmids by binding all copies of the plasmid of interest to each other (Figure 2) [47]. This system allows for the creation of permanent genetic switches, responses that scale with population growth, and simultaneously offers a robust strategy for controlling population ratios by creating new strains within a growing community.…”

Transcriptional factor engineering in microbes for industrial biotechnology

“…68 Constructing TF-containing genetic circuits for protein sciences The discovery and in-depth elucidation of the lac operon opened up an era for scientists to utilize the genetic circuits that naturally exist in the system to sense different compounds 69 or environmental factors, 70,71 to detect gene deletion 72 and bacterial cell division. 73 Engineered TFs are generally indispensable components in designed genetic circuits. 74,75 Besides the influences from its regulators, the expression level of genes is also affected by other genes through the impartment of dynamically available ribosomes in cells.…”

“…The discovery and in‐depth elucidation of the lac operon opened up an era for scientists to utilize the genetic circuits that naturally exist in the system to sense different compounds 69 or environmental factors, 70,71 to detect gene deletion 72 and bacterial cell division 73 . Engineered TFs are generally indispensable components in designed genetic circuits 74,75 …”