In most modern cells, DNA stores the genetic blueprint and relies on proteins to reproduce, repair, and build from those blueprints. At the same time, proteins require instructions from DNA in the first place.
So which came first? Neither, suggest researchers like Saurja DasGupta, a biochemist at the University of Notre Dame.
DasGupta seeks to better understand the chemical origins of life by studying the structure, function and evolution of RNA, an intermediate molecule that can store genetic information and catalyze biochemical reactions. In a study done today in Nature Communications, DasGupta and colleagues present a key mechanism for sustaining RNA-based life: an engineered enzyme that selectively recognizes and repairs broken RNA.
“Our results suggest that the molecular tools needed to maintain the RNA-based genetic code and pass it on to future generations could have been provided by RNA alone—no proteins required,” said DasGupta, who is an assistant professor in the Department of Chemistry and Biochemistry with a concurrent appointment in the Department of Biological Sciences.
The RNA-based enzyme, or “riboenzyme,” the researchers made, splices together pieces of RNA and targets a hallmark of broken RNA: a phosphate group—a phosphorus atom bonded to four oxygen atoms—at the end of the cleaved RNA chain. Intact RNA strands, by contrast, end up with a hydroxyl group—one oxygen atom and one hydrogen atom.
“The fact that this enzyme seeks out terminal phosphate groups in RNA—and thus broken RNAs—while ignoring strands that end with typical hydroxyl groups suggests that it could have been important for primordial RNA repair,” said DasGupta, who collaborated with Jack W. Szostak of the University of Chicago on the study.
The dual storage and catalysis capabilities of RNA form the basis of the RNA World hypothesis, which holds that the first life forms on Earth that lived for nearly four billion years were powered solely by RNA. The hypothesis suggests that RNA molecules precede DNA and proteins to encode genes and facilitate cellular processes, respectively.
Modern organisms have repair mechanisms to repair broken DNA. If early life forms carried their genes in RNA, then a similar repair process must have existed. Otherwise, when heat, high pH, or other stressors inevitably destroyed the RNA genome, the genetic information would have been lost forever, effectively stopping life in its tracks.”
Saurja DasGupta, biochemist, University of Notre Dame
A major obstacle to studying primitive RNA systems is that they no longer exist. In order to show that these RNA-based organisms and their components could have sustained life, researchers must first engineer new ribozymes through a process called in vitro evolution. The process involves selecting RNA catalysts with particular properties from trillions of RNA molecules in test tubes. Researchers impose certain conditions on these artificial evolution experiments in the hope of engineering ribozymes with specific functions, but often encounter surprises along the way.
“The general consensus is that artificial evolution comes down to a fair amount of luck,” DasGupta said. “Sometimes you get what you aim for, sometimes you don’t. And when you don’t, you start over and do it again.”
First, DasGupta’s research team set out to modify the biochemistry of an existing class of ribozymes using the method. But when they saw unexpected results, the researchers pursued them instead of dismissing them—and discovered something brand new.
“The existence of this riboenzyme has interesting implications for our understanding of the origin of life, and we came across it while looking for something else,” DasGupta said. “What I’m more surprised about, actually, is that it wasn’t found sooner.”
Beyond primordial biology, the importance of the new ribozyme extends into the realm of biotechnology.
Broken RNA is common in viral infections and is a sign of abnormal cell function in some cancers. However, standard RNA-sequencing techniques used to analyze genetic markers of these diseases miss the broken RNA, as the chemical tags that mark the RNA strands for analysis are not designed to attach to broken ends.
“Damaged RNAs are virtually invisible in standard sequencing protocols, which is a barrier to understanding the relationship between RNA decay and disease,” said DasGupta, who is a fellow at the Berthiaume Institute for Precision Health and the Warren Center for Drug Discovery.
Since the RNA repair riboenzyme is selective for broken RNA, it could be used to make the broken strands “visible” by isolating them for special preparation prior to RNA sequencing. As a first step, DasGupta’s research team is in the process of optimizing the reaction efficiency of the riboenzyme while expanding the range of possible molecular targets.
“What began as an RNA-based search for knowledge of the origin of life and resulted in an unexpected finding has also provided a potential solution to a major challenge in biotechnology,” said DasGupta. “We are excited to continue to pursue these new frontiers in ancient RNA biology and modern diagnostics.”
Study collaborators included first author Annyesha Biswas, a postdoctoral researcher in the Department of Chemistry and Biochemistry supported by the University of Industrial and Life Sciences initiative, and Zoe Weiss, MD-Ph.D. student at the Massachusetts Institute of Technology and Harvard University.
