TY - JOUR
T1 - Darwinian Evolution of Self-Replicating DNA in a Synthetic Protocell
AU - Abil, Zhanar
AU - Restrepo Sierra, Ana María
AU - Stan, Andreea R.
AU - Châne, Amélie
AU - Del Prado, Alicia
AU - de Vega, Miguel
AU - Rondelez, Yannick
AU - Danelon, Christophe
PY - 2024
Y1 - 2024
N2 - Replication, heredity, and evolution are characteristic of Life. We and others have postulated that the reconstruction of a synthetic living system in the laboratory will be contingent on the development of a genetic self-replicator capable of undergoing Darwinian evolution. Although DNA-based life dominates, the in vitro reconstitution of an evolving DNA self-replicator has remained challenging. We hereby emulate in liposome compartments the principles according to which life propagates information and evolves. Using two different experimental configurations supporting intermittent or semi-continuous evolution (i.e., with or without DNA extraction, PCR, and re-encapsulation), we demonstrate sustainable replication of a linear DNA template – encoding the DNA polymerase and terminal protein from the Phi29 bacteriophage – expressed in the ‘protein synthesis using recombinant elements’ (PURE) system. The self-replicator can survive across multiple rounds of replication-coupled transcription-translation reactions in liposomes and, within only ten evolution rounds, accumulates mutations conferring a selection advantage. Combined data from next-generation sequencing with reverse engineering of some of the enriched mutations reveal nontrivial and context-dependent effects of the introduced mutations. The present results are foundational to build up genetic complexity in an evolving synthetic cell, as well as to study evolutionary processes in a minimal cell-free system.
AB - Replication, heredity, and evolution are characteristic of Life. We and others have postulated that the reconstruction of a synthetic living system in the laboratory will be contingent on the development of a genetic self-replicator capable of undergoing Darwinian evolution. Although DNA-based life dominates, the in vitro reconstitution of an evolving DNA self-replicator has remained challenging. We hereby emulate in liposome compartments the principles according to which life propagates information and evolves. Using two different experimental configurations supporting intermittent or semi-continuous evolution (i.e., with or without DNA extraction, PCR, and re-encapsulation), we demonstrate sustainable replication of a linear DNA template – encoding the DNA polymerase and terminal protein from the Phi29 bacteriophage – expressed in the ‘protein synthesis using recombinant elements’ (PURE) system. The self-replicator can survive across multiple rounds of replication-coupled transcription-translation reactions in liposomes and, within only ten evolution rounds, accumulates mutations conferring a selection advantage. Combined data from next-generation sequencing with reverse engineering of some of the enriched mutations reveal nontrivial and context-dependent effects of the introduced mutations. The present results are foundational to build up genetic complexity in an evolving synthetic cell, as well as to study evolutionary processes in a minimal cell-free system.
UR - http://www.scopus.com/inward/record.url?scp=85206967453&partnerID=8YFLogxK
U2 - 10.1038/s41467-024-53226-0
DO - 10.1038/s41467-024-53226-0
M3 - Article
C2 - 39433731
AN - SCOPUS:85206967453
SN - 2041-1723
VL - 15
JO - Nature Communications
JF - Nature Communications
IS - 1
M1 - 9091
ER -