The original discovery
of messenger RNA (mRNA) by two Oak Ridge National Laboratory scientists "has
never received the acclaim it deserves," says Alvin M. Weinberg,
former ORNL director and a distinguished fellow of Oak Ridge
The discovery, for which Jacob and Monod received a Nobel Prize, was "next to the original discovery of the molecular structure of DNA, probably the most important event in the history of molecular biology," Weinberg says. Paul Berg, winner of the 1980 Nobel Prize in Chemistry, calls the ORNL research an "unsung but momentous discovery of a fundamental mechanism in genetic chemistry" and a "seminal discovery [that] has never received its proper due."
Messenger RNA is the life-sustaining ribonucleic acid (RNA) that serves as the living cell's template for protein synthesis. Volkin and Astrachan first discovered the acid three years after James Watson and Francis Crick determined the structure of DNA, which makes up genes. For this 1953 discovery Watson, Crick, and Maurice Wilkins received the Nobel Prize for Medicine or Physiology in 1962.
In the mid-1950s scientists knew that genes contained the coding that dictates the molecular structure of proteins, the tens of thousands of fundamental molecules of living cells necessary for the proper functioning of an organism. They also knew that proteins were synthesized on miniature factories called ribosomes, which are found outside the cell's nucleus in the region called the cytoplasm.
Not understood was how the information from inside the nucleus is conveyed to the protein factories in the cytoplasm. One theory at the time was that each ribosome is made in the nucleus, endowed with the DNA code required to direct the assembly of a specific protein, and then exported to the cell's outer region. Some scientists speculated that RNA, a sister molecule of DNA, was involved in protein assembly because RNA is the chief ingredient of ribosomes. However, little experimental information supported the notion that RNA could carry information from the cell's nucleus to its periphery, until Volkin and Astrachan made their discovery.
Elucidation of RNA Structure at ORNL
Using radioisotopes and the ion-exchange chromatography technique developed for the separation of fission products at ORNL's Graphite Reactor, Waldo Cohn was able to isolate uniformly each of the four chemical bases of DNA and RNA molecules. He found that each base bound to the ion-exchange column in a different position, according to its unique ionic strength, depending on the pH of the solvent used to extract one material from another. Because DNA and RNA are organic molecules with pentose-phosphate backbones, Cohn and Volkin incorporated radioactive carbon and phosphorus into these molecules to help determine their structure.
By detecting and measuring the beta radiation of the chemical degradation products, scientists could obtain considerable knowledge about the structure of RNA. However, Volkin convinced Cohn that more insight could be gained using enzymatic hydrolysis.
"Ion-exchange analysis of the digestion products of the enzyme, pancreatic ribonuclease, made it possible to more clearly define the composition of RNA and, in fact, even allowed a partial sequencing of the RNA," Volkin says. Cohn and Volkin then used other enzymes to show that the principal products were mononucleotides with phosphate groups attached to the fifth carbon atom of ribose. These experiments are considered to have been essential to establishing the structure of the ribose-phosphate chain of RNA.
Discovery of Messenger RNA at ORNL
Volkin then became interested in working with bacteriophage, a virus that infects only bacteria. Other researchers had determined that no net synthesis of RNA takes place in these microorganisms. "It occurred to me that no other biological system has both active DNA and protein synthesis but not active RNA synthesis," he says.
Volkin infected bacterial cells of Escherichia coli with the bacteriophage virus, added phosphorus-32, isolated nucleic acid from the preparation, and hydrolyzed it with sodium hydroxide to make alkaline products that were separated using ion-exchange chromatography. The results of experiments with phosphorus-32 were confirmed using a carbon-14 precursor that was specifically incorporated into the nucleic acid bases. Larry Astrachan joined Volkin in performing these experiments, which led to the discovery of messenger RNA, but they called it "DNA-like RNA."
According to Berg, the ORNL researchers "discovered that the virus 'turns off' the [bacterial] cell's machinery for making its own proteins and 'instructs' the cell's machinery to make proteins characteristic of the virus. That instruction entails making a new kind of RNA, a copy of the virus's DNA. This discovery revealed a fundamental mechanism for gene action: the coding sequences of genes are copied into short-lived RNAs that are transported out of the nucleus into the cytoplasm, where they are translated into proteins. Because such RNAs transport information from genes in the nucleus to the cytoplasm, they are designated as messenger RNAs."
Salvatore Luria, who became a Nobel laureate, convinced Volkin and Astrachan to publish their first paper on RNA research in the Journal of Virology in 1956. The paper announcing the discovery of a new kind of RNA is titled "Phosphorus Incorporation in E. Coli Ribonucleic Acid After Infection."
In an interview at his Oak Ridge home in late 2003, Volkin recalled his conversation with Sydney Brenner at Cold Spring Harbor Laboratory in New York, where Volkin conducted research on the hot topic of bacterial viruses during the summers in the late 1950s. "I can well remember sitting on the lawn at Cold Spring Harbor and telling Sydney Brenner about our experiments," Volkin says. "I gave a presentation on our RNA research to the group there." In a 1977 issue of Nature, renowned biophysicist T. H. Jukes wrote that in 1956, "I had squeezed my way into a doorway of a packed room to hear a paper by Volkin and Astrachan on DNA-like RNA."
According to Volkin, the ORNL findings were not widely accepted by the biology community because they challenged prevailing theory. Nevertheless, the ORNL researchers repeated their experiment several times and achieved the same result.
In a book review in a 2001 issue of Nature, Horace Judson, a renowned historian of science who contributed to Time magazine, attributed the discovery of messenger RNA to François Jacob, Sydney Brenner, and Matthew Meselson. Weinberg published a letter in the November 29, 2001, issue of Nature disputing this claim. "In fact," he writes, "Jacob, Brenner, and Francis Crick, at an informal meeting on Good Friday 1960, suddenly 'discovered' the unique RNA found first in 1956 by Elliot Volkin and Lazarus Astrachan. Good accounts of this event can be found in The Statue Within by Jacob and What Mad Pursuit by Crick.
"In several publications from 1956 through 1958, Volkin and Astrachan thoroughly described the unusual properties of this RNA, which they termed DNA-like RNA. These were precisely the properties that Jacob and Jacques Monod sought to assign to the unstable intermediate (which they called X), necessary for the synthesis of galactosidase.
"Out of that Good Friday discussion on the lactose operon came the realization that Volkin and Astrachan's DNA-like RNA was indeed the genetic messenger, hence the messenger RNA (mRNA)."
In his August 2, 2003, obituary for Astrachan in the New York Times, Nicholas Wade cited Judson's history of molecular biology, The Eighth Day of Creation, in his statement that Brenner, in that 1960 meeting in Cambridge, England, with Jacob and Crick, "realized there must be a missing ingredient that carried information from the DNA in the cell's nucleus to the ribosomes in its periphery. This ingredient, he conjectured, must be the same as the transitory form of RNA seen in the Volkin-Astrachan experiment."
In 1965 French scientists Monod, Jacob, and Andre Lwoff received the Nobel Prize in Physiology or Medicine for elucidating the nature of mRNA from their observation of protein synthesis by genes of mutated bacteria in the presence of lactose. Brenner (a 2002 Nobel Prize winner), Crick, and Jacob were internationally acclaimed for the discovery of mRNA. Although these giants of molecular biology are properly credited for their accomplishments, Berg and Weinberg believe that Volkin and Astrachan have never been appropriately recognized for their original discovery.
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