Periodic Reporting for period 4 - EvoTrap (Mechanisms to emerge and replicate the first sequence information of life in geothermal microfluidics of early Earth)
Reporting period: 2023-04-01 to 2023-09-30
How life emerged is a long standing problem of science. That it could not be resolved yet, sparked concern that science and especially biology is not yet complete. We performed highly diverse experiments that explore physical non-equilibria to bring about prebiotic reactions to now much better understand how Darwinian evolution could be triggered on an early Earth and how these reactions gave rise to an ever increasing complexification of biological information.
For this, we either focused on creating a most simple RNA world where a low number of molecules, in our case only two nucleotides of RNA, would trigger the evolution of sequence information. We found that at alkaline conditions, these molecules trigger ring opening polymerization of short random sequences, if they were activated by 2’,3’-cyclic phosphates. The usage of elevated pH at dry conditions without buffering molecules enabled a most simple and start of biological molecules by wet-dry cycling at room temperature. Very interestingly, the same conditions in wet state triggered an albeit slow, but highly precise replication reaction by templated ligation. In nature, this reaction cascade would be triggered by the low salt wet state in morning dew that separated the RNA strands, which would increase salt concentrations as the humidity drops over the course of the day, triggering replication by ligation and lead to a dry state at noon, which would polymerize again the raw material for the next cycle. Recycling is very natural since the hydrolysis of RNA leads again to the 2’,3’-cyclic phosphate ends of RNA.
On the other hand, we explored geological settings such as hydrothermal vents, but also looked into long-term evolution of replicating processes, by either templated ligation or templated polymerization with the use of proteins which speed up the reactions. Sequencing was established for these reactions and we found autocatalytic loops in templated ligation and the formation of long strands in templated polymerization. We could also show that Ribozymes flourish under the air-water interface conditions which were speeding up replication and selection reactions. What was most exciting was that the same settings which were very good for RNA evolution also triggered the accumulation of modern biochemistry in the form of cell-free extracts. These materials are basically the inner molecule mix of cells which are very sensitive to conditions. However, we found that both thermophoretic accumulation and accumulation at heated air-water interfaces triggered their coordinated accumulation and constituted a cellular state without cells and without membranes, offering a completely new way to think about synthetic cells in the future.
As a result, we now see new ways of life to emerge and we think the hard hurdles are overcome. The fog is lifting and straight forward experiments become clear now to demonstrate in ever more detail how some initially lonely molecules could create an autocatalytic process of sequence replication that brought about the wonders of life onto an early Earth – and possibly also to other planets.
For this, we either focused on creating a most simple RNA world where a low number of molecules, in our case only two nucleotides of RNA, would trigger the evolution of sequence information. We found that at alkaline conditions, these molecules trigger ring opening polymerization of short random sequences, if they were activated by 2’,3’-cyclic phosphates. The usage of elevated pH at dry conditions without buffering molecules enabled a most simple and start of biological molecules by wet-dry cycling at room temperature. Very interestingly, the same conditions in wet state triggered an albeit slow, but highly precise replication reaction by templated ligation. In nature, this reaction cascade would be triggered by the low salt wet state in morning dew that separated the RNA strands, which would increase salt concentrations as the humidity drops over the course of the day, triggering replication by ligation and lead to a dry state at noon, which would polymerize again the raw material for the next cycle. Recycling is very natural since the hydrolysis of RNA leads again to the 2’,3’-cyclic phosphate ends of RNA.
On the other hand, we explored geological settings such as hydrothermal vents, but also looked into long-term evolution of replicating processes, by either templated ligation or templated polymerization with the use of proteins which speed up the reactions. Sequencing was established for these reactions and we found autocatalytic loops in templated ligation and the formation of long strands in templated polymerization. We could also show that Ribozymes flourish under the air-water interface conditions which were speeding up replication and selection reactions. What was most exciting was that the same settings which were very good for RNA evolution also triggered the accumulation of modern biochemistry in the form of cell-free extracts. These materials are basically the inner molecule mix of cells which are very sensitive to conditions. However, we found that both thermophoretic accumulation and accumulation at heated air-water interfaces triggered their coordinated accumulation and constituted a cellular state without cells and without membranes, offering a completely new way to think about synthetic cells in the future.
As a result, we now see new ways of life to emerge and we think the hard hurdles are overcome. The fog is lifting and straight forward experiments become clear now to demonstrate in ever more detail how some initially lonely molecules could create an autocatalytic process of sequence replication that brought about the wonders of life onto an early Earth – and possibly also to other planets.
With the title page of Nature Chemistry 2019 how water-air interfaces enrich, crystallize, dry, phosphorylate and encapsulate prebiotic molecules we started our science journey to better understand the origins of life. The story continued with many results, shown below, and culminated in a Nature paper showing how heated cracks in rock accumulate and deplete molecules highly selectively, creating pockets of enhanced molecular reactions.
While these are diverse results, shown in many high impact publications, the 2’,3’-cyclic activation at high pH has the potential to establish Darwinian evolution from only two nucleotides without other helping chemical reactions, a breakthrough for understanding the Origin of Life. Also, the positive evaluation of many non-equilibrium systems showed us that our strategy to combine physics and chemistry was correct and much progress could be achieved by these microscale experiments. Since we are seeing that cell-free systems are equally enhanced by the systems we actually developed for an early RNA world, we can see a red line through early biology in porous rock samples, allowing us to recreate the first evolution mechanisms to create from simple nucleotides, stepwise, the rich molecular systems of biochemistry.
While these are diverse results, shown in many high impact publications, the 2’,3’-cyclic activation at high pH has the potential to establish Darwinian evolution from only two nucleotides without other helping chemical reactions, a breakthrough for understanding the Origin of Life. Also, the positive evaluation of many non-equilibrium systems showed us that our strategy to combine physics and chemistry was correct and much progress could be achieved by these microscale experiments. Since we are seeing that cell-free systems are equally enhanced by the systems we actually developed for an early RNA world, we can see a red line through early biology in porous rock samples, allowing us to recreate the first evolution mechanisms to create from simple nucleotides, stepwise, the rich molecular systems of biochemistry.
We could show a number of firsts in the field. We first showed that ligation creates an autocatalytic replication network from random sequences, two demonstrations that physical binding between oligonucleotides can create a replicator, one for tRNA-like molecules and one for oligonucleotide phase transitions that sediment in a sequence-dependent manner. We showed how CO2 is helping replication for the first time, that coacervates are not only getting together, but can also be divided by the help of air-water interfaces. We understood how the often dismissed 2’,3’-cyclic phosphate activation of RNA, the natural activation since it is recyclized by hydrolysis, can at elevated pH trigger both polymerization and templated ligation spontaneously in a wet-dry cycle of morning dew and afternoon dryness at room temperature. This opens a new and very promising way for the emergence of Life, enabled by this ERC. We could show that templated ligation, while slow, has a unique high prevision in its replication, reaching 98% fidelity per base pair replicated. For the first time, we could show that Ribozymes can replicate RNA in an autonomous setting, either with a fast thermal convection or – and that is not yet fully possible – by partially reassembling the Ribozyme itself by templated ligation at an air-water interface. We have explored geological settings such as pores in rock and self-generated alkaline vents, which were found to trap RNA. This work culminated in a Nature paper on how a random network of thermophoretic traps enhance prebiotic reactions by their selective accumulation of small biomolecules such as nucleotides or amino acids. The last author is our still young scientist and assistant Christof Mast in the group.