Protocell Progress

Researchers clear hurdle to making primitive, synthetic cells

HMS investigators at Massachusetts General Hospital have accomplished an important step toward their goal of creating primitive synthetic cells. 

The “protocells” they are building consist of a nucleic acid strand encased within a membrane-bound compartment. The scientists faced what could have been a critical problem: incompatibility between a chemical requirement of RNA copying and the stability of the protocell membrane. In the November 28 issue of Science, they described their solution.

“For the first time, we’ve been able to do nonenzymatic RNA copying inside fatty acid vesicles,” said Jack Szostak, HMS professor of genetics and a winner of the 2009 Nobel Prize in Physiology or Medicine for his contribution to the discovery of the enzyme telomerase. “We’ve found a solution to a longstanding problem in the origin of cellular life: RNA copying chemistry requires the presence of the magnesium ion Mg2+, but high Mg2+ levels can break down the simple fatty acid membranes that probably surrounded the first living cells.”

(Upper sequence, from left) As protocells replicate their nucleic acid and grow, their bubble-like vesicles extend into thread-like structures, eventually breaking off to form new vesicles containing their own nucleic acid. (Lower sequence) RNA templates are bound to short RNA primers within vesicles made of fatty acid membranes. Triangular complexes of Mg2+ ions, required for RNA replication, chelated with citrate to protect the membrane from damage, pass into the cell and catalyze extension of the RNA primer (red). Image: Katarzyna Adamala

Szostak’s team has been working for more than a decade to understand how the first cells developed from a primordial soup of chemicals into living organisms capable of copying their genetic material and reproducing. Part of that work is developing a model protocell made from components probably present in the primitive Earth environment. 

The scientists have made significant progress towards developing cell membranes from the kind of fatty acids that would have been abundant and that naturally form themselves into bubble-like vesicles when concentrated in water. But the genetic component—an RNA or DNA molecule capable of replication—has been missing.

Because the primitive environment in which such cells could have developed would not have had the kind of complex enzymes that modern cells use in replicating nucleic acids, Szostak and lead author Katarzyna Adamala, then a graduate student in Szostak’s lab and now a postdoctoral fellow at MIT, investigated whether simple chemical processes could drive nonenzymatic replication of RNA. Many scientists believe RNA was the first nucleic acid to develop. 

To address the incompatibility between the need for the magnesium ion to drive assembly of the RNA molecule and the ion’s ability to degrade fatty acid membranes, they tested several chelators—small molecules that bind tightly to metal ions—for their ability to protect fatty acid vesicles from the destabilizing effects of the magnesium ion. Citrate and several other chelators were found to be effective in protecting the membranes of fatty acid vesicles from disruption.  

To test whether the presence of the tested chelators would allow RNA assembly catalyzed by the magnesium ion, the investigators placed molecules consisting of short primer RNA strands bound to longer RNA templates into fatty acid vesicles. The unbound, single-strand portion of the template consisted of a sequence of cytosine (C) nucleotides. In the presence of the magnesium ion and one of four chelating molecules, one of which was citrate, the researchers then added activated guanine (G), the nucleotide that base-pairs with C in nucleic acids. 

The desired reaction—diffusion of G nucleotides through the vesicle membrane to complete a double-stranded RNA molecule by binding to the C nucleotides of the template—proceeded fastest in the presence of citrate. In fact, two of the other tested chelators completely prevented extension of the RNA primer. 

“While other molecules can protect membranes against the magnesium ion, they don’t let RNA chemistry go on,” Szostak explained. “We think that citrate is able both to protect membranes and to allow RNA copying to proceed by covering only one face to the magnesium ion, protecting the membrane while allowing RNA chemistry to work.”

Szostak and Adamala also found that continually refreshing the activated guanine nucleotide solution by flushing out broken-down molecules and adding fresh nucleotides improved the efficiency of RNA replication.

While citrate may be appropriate for creating artificial cells in a laboratory environment, which Szostak and his team are pursuing, he noted that it probably would not have been present in sufficient quantities on the early Earth. 

“We have shown there is at least one way to make RNA-replication chemistry compatible with primitive, fatty acid-based cell membranes, but this opens up new questions,” he said. “Our current best guess is there must have been some sort of simple peptides that acted in a similar way to citrate. Finding such peptides is something we are working on now.”

The study was supported in part by NASA Exobiology grant NNX07AJ09G. 

Adapted from a Mass General news release.