Topological and thermodynamic factors that influence the evolution of small networks of catalytic RNA species.

An RNA-directed recombination reaction can result in a network of interacting RNA species. It is now becoming increasingly apparent that such networks could have been an important feature of the RNA world during the nascent evolution of life on the Earth. However, the means by which such small RNA networks assimilate other available genotypes in the environment to grow and evolve into the more complex networks that are thought to have existed in the prebiotic milieu are not known. Here, we used the ability of fragments of the Azoarcus group I intron ribozyme to covalently self-assemble via genotype-selfish and genotype-cooperative interactions into full-length ribozymes to investigate the dynamics of small (three- and four-membered) networks. We focused on the influence of a three-membered core network on the incorporation of additional nodes, and on the degree and direction of connectivity as single new nodes are added to this core. We confirmed experimentally the predictions that additional links to a core should enhance overall network growth rates, but that the directionality of the link (a "giver" or a "receiver") impacts the growth of the core itself. Additionally, we used a simple mathematical model based on the first-order effects of lower-level interactions to predict the growth of more complex networks, and find that such a model can, to a first approximation, predict the ordinal rankings of nodes once a steady-state distribution has been reached.

Dynamics of prebiotic RNA reproduction illuminated by chemical game theory.

Many origins-of-life scenarios depict a situation in which there are common and potentially scarce resources needed by molecules that compete for survival and reproduction. The dynamics of RNA assembly in a complex mixture of sequences is a frequency-dependent process and mimics such scenarios. By synthesizing Azoarcus ribozyme genotypes that differ in their single-nucleotide interactions with other genotypes, we can create molecules that interact among each other to reproduce. Pairwise interplays between RNAs involve both cooperation and selfishness, quantifiable in a 2 × 2 payoff matrix. We show that a simple model of differential equations based on chemical kinetics accurately predicts the outcomes of these molecular competitions using simple rate inputs into these matrices. In some cases, we find that mixtures of different RNAs reproduce much better than each RNA type alone, reflecting a molecular form of reciprocal cooperation. We also demonstrate that three RNA genotypes can stably coexist in a rock-paper-scissors analog. Our experiments suggest a new type of evolutionary game dynamics, called prelife game dynamics or chemical game dynamics. These operate without template-directed replication, illustrating how small networks of RNAs could have developed and evolved in an RNA world.

Group I Intron Internal Guide Sequence Binding Strength as a Component of Ribozyme Network Formation.

Origins-of-life research requires searching for a plausible transition from simple chemicals to larger macromolecules that can both hold information and catalyze their own production. We have previously shown that some group I intron ribozymes possess the ability to help synthesize other ribozyme genotypes by recombination reactions in small networks in an autocatalytic fashion. By simplifying these recombination reactions, using fluorescent anisotropy, we quantified the thermodynamic binding strength between two nucleotides of two group I intron RNA fragments for all 16 possible genotype combinations. We provide evidence that the binding strength (KD) between the 3-nucleotide internal guide sequence (IGS) of one ribozyme and its complement in another is correlated to the catalytic ability of the ribozyme. This work demonstrates that one can begin to deconstruct the thermodynamic basis of information in prebiotic RNA systems.

RNA-directed recombination of RNA in vitro.

Construction of long RNAs can be achieved in vitro by using ribozymes to recombine shorter RNAs. This can be a useful technique to prepare RNAs when the final product is either very long or contains chemical modifications that are difficult to incorporate using standard in vitro transcription techniques. Here, we describe the use of the Azoarcus group I intron ribozyme to recombine shorter RNAs into longer ones. This ribozyme is a generalized RNA recombinase ribozyme that operates rapidly and with high efficiency.

Prebiotic network evolution: six key parameters.

The origins of life likely required the cooperation among a set of molecular species interacting in a network. If so, then the earliest modes of evolutionary change would have been governed by the manners and mechanisms by which networks change their compositions over time. For molecular events, especially those in a pre-biological setting, these mechanisms have rarely been considered. We are only recently learning to apply the results of mathematical analyses of network dynamics to prebiotic events. Here, we attempt to forge connections between such analyses and the current state of knowledge in prebiotic chemistry. Of the many possible influences that could direct primordial network, six parameters emerge as the most influential when one considers the molecular characteristics of the best candidates for the emergence of biological information: polypeptides, RNA-like polymers, and lipids. These parameters are viable cores, connectivity kinetics, information control, scalability, resource availability, and compartmentalization. These parameters, both individually and jointly, guide the aggregate evolution of collectively autocatalytic sets. We are now in a position to translate these conclusions into a laboratory setting and test empirically the dynamics of prebiotic network evolution.