Experimental Tests of the Virtual Circular Genome Model for Nonenzymatic RNA Replication

Abstract

The virtual circular genome (VCG) model was proposed as a means of going beyond template copying to indefinite cycles of nonenzymatic RNA replication during the origin of life. In the VCG model, the protocellular genome is a collection of short oligonucleotides that map to both strands of a virtual circular sequence. Replication is driven by templated nonenzymatic primer extensions on a subset of kinetically trapped partially base-paired configurations, followed by the shuffling of these configurations to enable continued oligonucleotide elongation. Here, we describe initial experimental studies of the feasibility of the VCG model for replication. We designed a small 12-nucleotide model VCG and synthesized all 247 oligonucleotides of lengths 2 to 12 corresponding to this genome. We experimentally monitored the fate of individual labeled primers in the pool of VCG oligonucleotides following the addition of activated nucleotides and investigated the effect of factors such as oligonucleotide length, concentration, composition, and temperature on the extent of primer extension. We observe a surprisingly prolonged equilibration process in the VCG system that enables a considerable extent of reaction. We find that environmental fluctuations would be essential for continuous templated extension of the entire VCG system since the shortest oligonucleotides can only bind to templates at low temperatures, while the longest oligonucleotides require hightemperature spikes to escape from inactive configurations. Finally, we demonstrate that primer extension is significantly enhanced when the mix of VCG oligonucleotides is preactivated. We discuss the necessity of ongoing in situ activation chemistry for continuous and accurate VCG replication.

Enhanced nonenzymatic RNA copying within-situactivation of short oligonucleotides

“…Remarkably, activating all of the oligonucleotide components in this system significantly enhanced the observed rate of primer extension (34), consistent with a role for short, activated oligonucleotides in the catalysis of RNA copying chemistry. The combination of the prebiotically plausible activation chemistry presented in our current work with the virtual circular genome replication model has the potential to drive a nonenzymatic replication system through continuous in-situ activation.…”

“…We have recently proposed and provided the first experimental tests of a model for nonenzymatic RNA replication (33,34). This virtual circular genome model invokes a collection of linear oligonucleotides of varying lengths that map onto both strands of a circular genomic sequence.…”

Natural soda lakes provide compatible conditions for RNA and membrane function that could have enabled the origin of life

“…However, protocells may not have needed to synthesize the full complementary strand for a given template. Unlike modern cells that contain large linear or circular genomes, protocells may have contained small, virtual circular genomes where RNA synthesis can be transiently templated by multiple oligomers at different positions for replication ( 42 , 43 ). Additionally, we recognize that 72 hours is not an endpoint for nonenzymatic primer extension in lake water.…”

Sequence Information Transfer in Oligoarylacetylenes

“…[3a,15] We have recently described a method of chemical information transfer for RNA which only requires extension by one or a few nucleotides. [16] This suggests that the copying of long templates may not be a strict requirement for significant information transfer in all chemical systems.…”