One of the most remarkable features of DNA is its precision. Cells read genetic instructions in sets of three letters called codons, and each codon corresponds to a specific amino acid. Those amino acids are linked together in a defined order to build proteins, the molecules that carry out most of life’s essential tasks.
Researchers at the University of California, Berkeley have now identified a microorganism that challenges this long accepted rule. Their findings show that at least one microbe can tolerate ambiguity in its genetic code, overturning a central assumption in biology.
The organism is a methane producing member of a group of microbes known as Archaea. It treats a particular three letter sequence, typically a stop codon that marks the end of a protein, in two different ways. Sometimes the cell stops building the protein. Other times it inserts an amino acid and keeps going. This produces two distinct proteins from the same genetic instruction. The microbe, Methanosarcina acetivorans, appears to function normally despite this flexible interpretation, demonstrating that life can operate with a slightly imprecise code.
Scientists think this ambiguity may have evolved to allow the organism to insert a rare amino acid called pyrrolysine into an enzyme that breaks down methylamine, a compound commonly found in the environment and in the human gut.
“Objectively, ambiguity in the genetic code should be deleterious; you end up generating a random pool of proteins,” said Dipti Nayak, a UC Berkeley assistant professor of molecular and cell biology and senior author of a paper describing the findings published in the journal Proceedings of the National Academy of Sciences. “But biological systems are more ambiguous than we give them credit to be and that ambiguity is actually a feature — it’s not a bug.”
Why Methylamine Metabolism Matters
Archaea that consume methylamines, along with certain bacteria that may have acquired the same ability, play an important role in human health. When people eat red meat, the liver converts certain byproducts into trimethylamine N-oxide, a compound associated with cardiovascular disease. Microbes that remove methylamines before they reach the liver help limit the production of this potentially harmful molecule.
The discovery also raises the possibility of new medical strategies. Some genetic disorders are caused by premature stop codons in critical genes, which result in incomplete and nonfunctional proteins. These conditions account for roughly 10% of inherited diseases, including cystic fibrosis and Duchenne muscular dystrophy. Researchers have speculated that making stop codons slightly “leaky” could allow cells to produce enough full length protein to ease symptoms.
How the Genetic Code Normally Works
Genetic information stored in DNA is first copied into RNA. Cellular machinery then reads that RNA to assemble proteins. RNA is built from four chemical letters: adenine (A), cytosine (C), guanine (G) and uracil (U). In nearly all organisms studied so far, every three letter codon either specifies one particular amino acid or signals the end of a protein. The translation system follows this one to one relationship with strict consistency.
There is variation across life. Some organisms assign different amino acids to certain codons, some use more than the standard 20 amino acids, and multiple codons can correspond to the same amino acid. Even so, each codon has traditionally been understood to carry only one meaning.
“It’s essentially like a cipher,” Nayak said. “You’re taking something in one language and translating it into another, nucleotides to amino acids.”
For years, scientists have known that many Archaea can produce pyrrolysine, giving them 21 amino acids to work with instead of the usual 20. That extra building block can expand their biochemical capabilities.
“Now that you have a new amino acid, the world’s your oyster,” she said. “You can start playing around with the much larger code. It’s like adding one more letter to the alphabet.”
Researchers had assumed that these organisms simply reassigned the UAG stop codon to represent pyrrolysine.
A Stop Codon With Two Meanings
In the new study, Nayak and former graduate student Katie Shalvarjian surveyed a wide range of Archaea and found that many lineages produce pyrrolysine.
“We found that the machinery required to create pyrrolysine is widespread in the Archaea, especially amongst these methanogenic archaea that consume methylated amines,” said Shalvarjian, now a postdoctoral researcher at Lawrence Livermore National Laboratory.
She wanted to understand how carrying 21 amino acids instead of 20 influences these organisms. While studying how the methanogen controls pyrrolysine production, she noticed something unexpected. The UAG codon was not always translated as pyrrolysine (Pyl).
“The UAG codon is like a fork in the road, where it can be interpreted either as a stop codon or as a pyrrolysine residue,” Shalvarjian said. “We think whether or not a protein exists primarily in its elongated or in its truncated form might form a regulatory cue for the cell.”
The researchers searched for specific sequence or structural signals that might determine how UAG is interpreted, but they did not find any clear triggers.
“The methanogens have not recoded UAG, nor have they added any new factors to make it deterministic,” Nayak said. “They’re flip-flopping back and forth between whether they should call this a stop or whether they should keep going by adding this new amino acid. They cannot decide. They just do both and they seem to be fine by making this random choice.”
Early evidence suggests that the availability of pyrrolysine inside the cell may influence the outcome. When the amino acid is abundant, UAG is more likely to be read as pyrrolysine and the protein continues to grow. When pyrrolysine is scarce, the same codon functions as a stop signal. Between 200 and 300 genes in this organism contain UAG, meaning many proteins could be produced in two forms depending on cellular conditions.
“This really opens the door to finding interesting ways to control how cells interpret stop codons,” Nayak said.
The research was supported by the Searle Scholars Program, a Rose Hills Innovator Grant, a Beckman Young Investigator Award, an Alfred P. Sloan Research Fellowship, a Simons Foundation Early Career Investigator in Marine Microbial Ecology and Evolution Award, and a Packard Fellowship in Science and Engineering. Nayak is also a Chan-Zuckerberg Biohub-San Francisco investigator.
Additional co-authors include Grayson Chadwick and Paloma Pérez of UC Berkeley and Philip Woods and Victoria Orphan of the California Institute of Technology.
