The ability to evolve is a key characteristic that distinguishes living things from non-living chemical compounds. The construction of an evolvable cell-like system entirely from non-living molecules has been a major challenge. Here we construct an evolvable artificial cell model from an assembly of biochemical molecules. The artificial cell model contains artificial genomic RNA that replicates through the translation of its encoded RNA replicase. We perform a long-term (600-generation) replication experiment using this system, in which mutations are spontaneously introduced into the RNA by replication error, and highly replicable mutants dominate the population according to Darwinian principles. During evolution, the genomic RNA gradually reinforces its interaction with the translated replicase, thereby acquiring competitiveness against selfish (parasitic) RNAs. This study provides the first experimental evidence that replicating systems can be developed through Darwinian evolution in a cell-like compartment, even in the presence of parasitic replicators.

In all living systems, the genome is replicated by proteins that are encoded within the genome itself. This universal reaction is essential to allow the system to evolve. Here, we have constructed a simplified system involving encapsulated macromolecules termed a "self-encoding system", in which the genetic information is replicated by self-encoded replicase in liposomes. That is, the universal reaction was reconstituted within a microcompartment bound by a lipid bilayer. The system was assembled by using one template RNA sequence as the information molecule and an in vitro translation system reconstituted from purified translation factors as the machinery for decoding the information. In this system, the catalytic subunit of Qbeta replicase is synthesized from the template RNA that encodes the protein. The replicase then replicates the template RNA that was used for its production. This in-liposome self-encoding system is one of the simplest such systems available; it consists of only 144 gene products, while the information and the function for its replication are encoded on different molecules and are compartmentalized into the microenvironment for evolvability.

To date, various cellular functions have been reconstituted in vitro such as self-replication systems using DNA, RNA, and proteins. The next important challenges include the reconstitution of the interactive networks of self-replicating species and investigating how such interactions generate complex ecological behaviors observed in nature. Here, we synthesized a simple replication system composed of two self-replicating host and parasitic RNA species. We found that the parasitic RNA eradicates the host RNA under bulk conditions; however, when the system is compartmentalized, a continuous oscillation pattern in the population dynamics of the two RNAs emerges. The oscillation pattern changed as replication proceeded mainly owing to the evolution of the host RNA. These results demonstrate that a cell-like compartment plays an important role in host-parasite ecological dynamics and suggest that the origin of the host-parasite coevolution might date back to the very early stages of the evolution of life.arious functions of living organisms have been reconstituted in vitro from biological molecules to understand the basic principles underlying complex biological phenomena observed in the cell or in nature (1, 2). Several types of continuous systems of self-replication of genetic information have been constituted in vitro from RNA alone (3-6), and from the combinations of RNA and proteins (7) or of DNA, RNA, and proteins (8). One of the next important challenges is the reconstitution of interacting networks of self-replicating species, including the investigation of how such interactions generate complex ecological behaviors such as those observed in nature.Organisms in the wild self-reproduce and interact with each other to form ecosystems. Such interaction generates complex ecological behaviors such as the oscillation dynamics observed in prey-predator or host-parasite populations (9-12). The causes and results of such periodic patterning have been a focus of intensive debate for decades (13)(14)(15)(16)(17). Such interactions and the resultant coevolution of the interacting species are thought to be an important factor explaining the high degree of extant biodiversity and complexity (16,18). However, to date, the in vitro reconstitution of such ecological behavior has been limited; one example is of prey-predator oscillation dynamics constructed using a combination of small hybridizing DNA fragments and several enzymes, although the DNA fragment did not evolve in this system (19).Previously, we had constructed a simple translation-coupled RNA replication system in which an artificial genomic RNA replicated through translation of the self-encoded RNA replicase and evolved (7). This system consists of a reconstituted translation system of Escherichia coli and an RNA encoding the RNA replicase (Qβ replicase), which is composed of the catalytic β-subunit and the translational factors, EF-Tu and EF-Ts. We found that parasitic RNAs, which had lost their replicase encoding region, spontaneously appeared in this system at ...

Lysylphosphatidylglycerol (LPG) is a basic phospholipid in which L-lysine from lysyl-tRNA is transferred to phosphatidylglycerol (PG). This study examined whether the Staphylococcus aureus mprF gene encodes LPG synthetase. A crude membrane fraction prepared from wild-type S. aureus cells had LPG synthetase activity that depended on PG and lysyl-tRNA, whereas the membrane fraction from an mprF deletion mutant did not. When S. aureus MprF protein was trans-expressed in wild-type Escherichia coli cells, LPG synthesis was induced, whereas it was not observed in E. coli pgsA3 mutant cells in which the amount of PG is significantly reduced. In addition, LPG synthetase activity and a 93 kDa protein whose molecular size corresponded to that of MprF protein were co-induced in the crude membrane fraction prepared from E. coli cells expressing MprF protein. The K m values of the LPG synthetase activity for PG and for lysyl-tRNA were 56 mM and 6?9 mM, respectively, consistent with those of S. aureus membranes. These results suggest that the MprF protein is LPG synthetase.