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Machine Evolution

A 3D model of RNA
by James Kling
for Astrobiology Magazine
Moffett Field CA (SPX) Jul 25, 2008
The question of how life arose on Earth is one of the fundamental questions of biology. Modern life is a complex interplay between DNA that encodes life's instruction manual and the proteins that carry out those instructions.

But DNA has a close chemical relative, called RNA, which acts as an intermediary and may hold the key to life's origins.

Today, RNA is used by the cell to make copies of the gene-encoding elements of DNA. These copies, called messenger RNA (mRNA), are then shuttled to ribosomes, protein factories that "read" the mRNA and use its instructions to direct the assembly of proteins.

But the internal structure of ribosomes also contains a form of RNA.

RNA's critical supporting role in modern life forms - and its central role as part of the structure of the ribosome - has led some scientists to believe that it was the progenitor of all life.

In the so-called 'RNA world,' a hypothetical stage in the origin of life that came before DNA or proteins evolved, RNA molecules would have reproduced themselves and competed against one another, and slowly evolved into new forms.

Eventually, one of those RNA organisms would have evolved to incorporate proteins and DNA and then out-competed the other RNA organisms, driving them to extinction.

It all makes for fascinating speculation, but none of it can be tested because no viable RNA-only life forms are known to exist today. According to the RNA-world model, life's precursor would have been strands of RNA floating loose in a primordial soup. At some point, those must have evolved the ability to coalesce and reproduce themselves from nucleotide components.

Scientists can only study the properties of modern RNA and make inferences about the behavior of its ancient forebears.

Among the most crucial questions: Can individual strands of RNA evolve to reproduce themselves? Researchers have been working for more than a decade to reproduce RNA evolution in the laboratory. One of the leaders of that field is Gerald Joyce, who is a professor of chemistry and molecular biology at the Scripps Research Institute.

Joyce's early studies began with a pool of random RNA sequences and challenged them to attach an oligonucleotide to themselves. The oligonucleotide bore a signal that recruited proteins, also present in the mixture, which in turn replicated the RNA. This enabled successful RNA sequences to be reproduced, just as occurs in natural selection.

This technique was effective: Similar methods have produced novel RNA enzymes that have therapeutic use. But the technique was labor intensive because, in time, the RNAs ran out of 'food' - that is, oligonucleotides.

A researcher had to remove a sample of the mixture and add fresh ingredients to begin the process again. Experimenters may have wanted to conduct hundreds or thousands of iterations, but practical considerations generally limited them to several dozen. A few went into the hundreds. "But it's awkward," Joyce says.

In the April issue of PLoS Biology, Joyce and his postdoc Brian Paegel described a new method that could solve that problem by using computer automation. Using a fluorescent marker to signal when the RNA population had increased 10-fold (which took about 5 minutes), the instrument then removed a small sample and added it to a fresh mixture of reagents.

As that cycle was repeated, some RNAs became dominant in the mixture because of their talent for joining to the oligonucleotide, which caused them to be reproduced more frequently. The replication procedure reproduced these RNAs somewhat faithfully, but introduced various random mutations that might improve, or reduce, their ability to attach to the oligonucleotide.

Beginning with cycle number 101, the instrument reduced the concentration of the oligonucleotide to 10 percent of its original concentration, putting greater pressure on the newly mutated RNAs to scavenge this increasingly rare 'food.'

The instrument then started the process anew, progressively reducing the concentration of the oligonucleotide.

In one experiment, after 500 total cycles it was just 1 percent of the original concentration, and at that point Joyce and Paegel analyzed the resulting RNAs. The conditions had put evolutionary pressure on the RNAs to improve their ability to use an increasingly rare resource, and the results were evident.

"The population got very good at scavenging food," Joyce says. The ability of the RNA molecules to scavenge the food had increased 90-fold, roughly countering the 100-fold decrease in food concentration. All of the RNA sequences in the final solution possessed eleven mutations that had not been present in the starting RNA population.

The mutations had to have occurred in a particular order, Joyce says, because some of the mutations wouldn't have been beneficial unless other mutations had occurred first.

"This is testing evolution - real evolution, not a simulation. The molecules are evolving a new function before your eyes," says Joyce.

The technique could greatly enhance researchers' ability to study evolution. Given the same beginning conditions, would evolution produce the same or similar RNA structures or binding ability? Would the concentration of 'food' alter the path of evolution?

The only way to answer these and other questions was to conduct the experiment repeatedly, but the amount of effort involved in manual studies made that impracticable. "Now we can do the 'what if' experiments. This lowers the barrier," says Joyce.

According to Andrew Ellington, a professor of chemistry and biochemistry at the University of Texas at Austin, the research is a promising step.

He hopes that such experiments will eventually produce what he calls a 'Xeroxase' - an RNA enzyme capable of making more copies of itself with no assistance from proteins like those used in Joyce's experiments. "That would be a sure sign that the RNA world could have arisen from simple elements," he says.

The experimental power of Joyce's instrument could be the key to finding just such an organism. His work was supported by a grant from NASA's Exobiology/Evolutionary Biology program.

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