Too simple, too uniform. Those were the arguments that scientists made for nearly a century to explain why DNA was very likely not the blueprint for life that we consider it to be today.
DNA in water [Courtesy of Bbkkk, CC BY-SA 4.0, via Wikimedia Commons]
The Swiss doctor Friedrich Miescher
, who first isolated DNA in 1869 as a white, clumpy precipitate from the nuclei of white blood cells, suggested this new substance might be a storage center for phosphorus or a precursor for lipid molecules. Over the next several decades, scientists worked out the importance of chromosomes for carrying genetic information and determined that chromosomes are made of DNA and protein. But all bets were on protein as the stuff that genes are made of. Protein, with its 20 amino acids, compared with DNA’s four bases, seemed more apt to encode the complexity of life.
But experiments in the 1940s and 1950s made DNA a prime candidate for hereditary material. The finding by Alfred Hershey and Martha Chase in 1952 that it is the DNA, not the protein, of bacteriophages that gets inserted into bacteria during infection made the most convincing case at the time that genes are actually made of DNA. “That’s when the race began to find the structure of DNA,” says Evelyn Witkin, a 2015 Lasker Laureate. The moment that James Watson and Francis Crick described the now-iconic double helix in their famous 1953 publication, there was no looking back. “It was like a curtain being lifted and you could just see how it worked,” Witkin says of the meeting at Cold Spring Harbor, where she was based at the time, at which she first saw the DNA double helix model. “That was a high point that is hard to duplicate,” she adds.
In the span of just a couple of years the door was thrown open, after decades of being sealed shut, to the paramount importance of DNA. Its structure explained, almost intuitively, how it could be replicated as cells divide and how its information could be transmitted during reproduction. Although it was still formally only a “working hypothesis” that genes were made of DNA, it was getting harder to deny.
And yet, the model of DNA explained little about its functions in the cell. One question rose to the fore, as Lasker Laureate Walter Gilbert put it:
“How does DNA do anything?”
As the following decades revealed, what DNA does — how it is read, regulated, and repaired — turned out to be deceptively complex for such a seemingly simple molecule. From the breakneck progress cracking the genetic code to controversies over editing genomes, Lasker Laureates led some of the innumerable advances over the past 60 years. No one appreciates more than they do the central role that DNA plays in understanding life. “Historically and biologically, our understanding of DNA has been the driving force in all of this,” says Tom Maniatis, recipient of a 2012 Lasker Award.
Learning the Language of DNA
As soon as DNA was cast in the role of likely carrier of genetic information, a frenzy was unleashed in the scientific world to decipher the language of DNA’s four nucleotides. How could the As, Ts, Gs, and Cs, in sets of different sequences and lengths, spell out genes? And how could these genes convey meaning that would get passed on to daughter cells and daughter organisms?
Meeting of the members of the RNA Tie Club, Cambridge, UK Fall 1955 (from upper left): Francis Crick, Leslie Orgel, Alexander Rich and James D. Watson [Courtesy of the James D. Watson Collection, Cold Spring Harbor Laboratory Archives]
In the early 1950s, at the height of the frenzy, 20 esteemed (male) biologists, chemists, and physicists formed a scientific gentleman’s club to speed the answers to these pressing questions. Crick, who was part of the club, later apologized for the “chauvinistic, secret society.”
Nevertheless, it was discussions within the club that inspired Crick to propose at a symposium in London in 1957 that the main function of DNA is to make proteins and that RNA, which had long been suspected to be important for protein synthesis, is some kind of intermediary between DNA and protein.
Crick’s first illustration of the central dogma, from an unpublished note in 1956
The proposal may have been jarring at the time, but it is now the central dogma of biology. In keeping with this proposal, the gentleman’s club, which was dubbed the RNA Tie Club, assigned each of the 20 members one of the 20 amino acids with the mission to determine the nucleotide combinations that encode it.
In one of the biggest advances since discovering the structure of DNA, Francis Crick and Sydney Brenner, a fellow RNA Tie Club member and recipient of two Lasker Awards, led work at the University of Cambridge, UK that mapped out how nucleotides are read. Their experiments revealed that the magic number of nucleotides encoding a single amino acid was three, and nucleotide triplets, or codons, did not overlap.
Marshall Nirenberg in the 1960s [Courtesy of National Institutes of Health]
But credit for figuring out, the first codon that corresponds to an amino acid goes to Marshall Nirenberg, in 1961. He clearly didn’t remain an unheralded post-doc researcher at the National Institutes of Health. On Nirenberg’s heels, and building on his experimental system, Gobind Khorana, who shares a 1968 Lasker Award
with Nirenberg, defined the stretch of amino acids that results from a short sequence of 12 nucleotides. In just a few years, largely through the work of those two men (neither of whom belonged to the RNA Tie Club), the codons that give rise to each amino acid were known.
Cracking the genetic code, and learning how the linear molecule of DNA determines the three-dimensional structure of proteins, marked the dawn of molecular biology. “It was a lovely conception and idea behind molecular biology,” recalls Lasker Laureate Gilbert, who was so enthralled by the burgeoning field that he abandoned his physics studies to do molecular biology research.
In the watershed period of the early 1960s, there was also intense focus on describing the mysterious transient RNA molecule — which was dubbed “messenger RNA” or mRNA — that was thought to carry the information in DNA to ribosomes in the cytoplasm to instruct protein synthesis. Although identifying mRNA ultimately involved numerous scientists and multiple experimental approaches, Gilbert recalls many details of the experiments done by his team in Watson’s lab at Harvard University, including pouring radioactive phosphate into a bacterial culture and purifying the molecule from cells. It was becoming clear, in leaps and bounds, how the three main players — DNA, RNA, and protein — worked together in the machine of the cell to read the language of genes.
Revealing Gene Regulation
As the early puzzles were solved about the genetic code and how the information stored in DNA is converted into proteins, scientists were flocking to more nuanced questions about gene regulation. What controls when genes are expressed and their levels of expression?
“The next hallmark in the whole progression of molecular biology… [came from] Francois Jacob and Jacque Monod,” who gave the first hints about how genes are controlled, says Phillip Sharp, 1988 Lasker Laureate. The pair of scientists at the Pasteur Institute, who were also instrumental in describing mRNA, found a gene that repressed gene expression. But the French scientists had no idea how the product of this gene carried out this activity and suspected it was through an RNA molecule. Then in 1966, Gilbert, along with Mark Ptashne also at Harvard, found that repressors are proteins that bind to DNA.
Tom Maniatis at Cold Spring Harbor Laboratory in 1977 [Courtesy of the Cold Spring Harbor Laboratory Archives]
The concept of repressor molecules — and also activators, which were discovered later, in the 1960s — captured the imagination of many scientists. “I found it so interesting that you can regulate gene expression through protein-DNA interactions,” recalls Lasker Laureate Maniatis. He headed to the Ptashne lab in 1973 for his post-doc training, where he uncovered the presence of multiple repressor binding sites in a gene regulatory sequence in bacteriophages.
By the 1970s, research on gene regulation had become so competitive that some scientists avoided it altogether. “Everybody and their brother or sister were studying how RNA polymerase interacts with [regulatory] factors to transcribe a gene. What I wanted to do was work on something that was not crowded,” recalls Michael Grunstein. When he started his own research group in the mid-1970s, Grunstein decided to study “boring” histone proteins and how they package DNA into chromosomes. The plan yielded dramatic and unexpectedly exciting results when he started to notice that yeast mutants he engineered to lack one type of histone protein had higher levels of expression of certain genes, suggesting that seemingly inert histones might actually repress transcription in the living cell. Moreover, sites of posttranslational modification (acetylation) in histones were shown to be required for gene activity in vivo. Thus, his work clarified that histones and their acetylation sites help regulate transcription. Several years later, C. Davis Allis, who shares a 2018 Lasker Award with Grunstein, identified the enzyme that adds chemical groups to certain amino acids in histones and turns on gene expression.
Eager to push the understanding of gene regulation even further, scientists moved into new technology frontiers: DNA sequencing and gene cloning. As Maniatis says, it was becoming clear that “one could not even imagine studying gene expression in any detail in eukaryotic cells without recombinant DNA,” whose advent was right around the corner.
Taming a Messy Molecule
Just as scientists did not grasp the function of DNA until well after they appreciated the roles of RNA and proteins, their ability to sequence DNA trailed behind their ability to do so for RNA and proteins. In fact, most scientists thought that DNA sequencing was not possible, but the research teams of Gilbert and Frederick Sanger at the University of Cambridge (both recipients of a 1979 Lasker Award and a 1980 Nobel Prize) persevered. “Various members of the [Sanger] lab were all trying different methods for sequencing DNA,” recalls Elizabeth Blackburn, winner of a 2006 Lasker Award, of her PhD with Sanger in the early 1970s. In fact, one of the early success stories involved copying a stretch of DNA into RNA and sequencing the RNA fragments. Using that approach in 1974, it took Gilbert about two years to read 20 nucleotides in the bacterial repressor binding site.
Fred Sanger at Laboratory of Molecular Biology, UK, circa 1969 [Courtesy of MRC Laboratory of Molecular Biology]
By 1976, both Gilbert and Sanger had developed rapid sequencing methods that allowed several hundred nucleotides to be read in half a day. Over the next decade, the two methods were used widely, Gilbert recalls, and scientists around the world collectively sequenced about 8 million bases. But the Gilbert method took a backseat when automated sequencing machines were developed in the late 1980s that relied on the Sanger method. Ever since this technological leap, which was led by Leroy Hood, recipient of a 1987 Lasker Award
, the capacity to sequence DNA has been growing exponentially. According to Gilbert, it has been doubling around every 18 months, comparable to the rate of increase in computing power.
The 1970s saw turning points in the ability not just to sequence DNA but also to study and manipulate it. Donald Brown, recipient of a 2012 Lasker Award, recalls trying to work with DNA in the late 1960s, and it “was just a glop, a mess.” But certain DNA fragments, such as the ribosomal RNA (rRNA) genes that Brown and 2006 Lasker Award recipient Joseph Gall worked with, had properties that made their isolation possible even before the days of recombinant DNA. With the isolated rRNA genes in hand, Blackburn was able to sequence the telomeres at their ends. The rRNA genes also played a pivotal part in the early days of recombinant DNA. Brown sent the purified genes to Herbert Boyer and Stanley Cohen, two of the four scientists to receive a 1980 Lasker Award for developing recombinant DNA tools such as restriction enzymes and bacterial plasmids. In a now-classic experiment, Boyer and Cohen inserted fragments of the genes into a plasmid that they then delivered into Escherichia coli and demonstrated that bacteria could make copies of foreign DNA. Brown proceeded to use these rRNA clones to uncover some of the first details about gene regulation in eukaryotes.
Elizabeth Blackburn in 1975 [Courtesy of Elizabeth Blackburn]
Later that decade, Maniatis and collaborators built on recombinant DNA technology by developing clones of complementary DNA (or cDNA, derived from reverse transcription of mRNA) that could be used as probes to identify genes from genomic DNA libraries, to study gene regulation in detail and churn out mammalian proteins in simple organisms such as bacteria. Before long, new therapies came to market based on this technology, the first of which was insulin (Humulin), approved by the US Food and Drug Administration in 1982 for treating diabetes. Soon after, pharmaceutical companies began turning to technology to update how existing medicines and vaccines were produced.
With the advent of modern molecular biology, concerns were growing that recombinant DNA might present risks, and certain research institutions declared a moratorium on such research. But some of the scientists who initially most vehemently opposed recombinant technology later became staunch supporters of the fledgling Human Genome Project, which by 2003 had relied on the technology to sequence entire human genomes.
Scientists have used sequencing and recombinant DNA technologies to determine the function of genes, how genes make us who we are, and contribute to human disease. In the late 1970s, scientists gained additional tools: Southern blotting and DNA fingerprinting, developed by 2005 Lasker Award recipients Edwin Southern and Alec Jeffreys, respectively. Previously, to scan for specific DNA sequences within genomes, scientists had to grapple with a long, diffuse smear of genomic fragments in an agarose gel, after separating the fragments using gel electrophoresis. Southern devised a technique to transfer the DNA smear to a membrane that could be easily probed using radioactive complementary RNA fragments, and with it, scientists quickly identified sequence differences in genes associated with sickle cell anemia and other diseases. Alec Jeffreys made use of Southern blotting to find regions across the human genome that are highly variable, and he determined how to probe for a combination of such regions to distinguish unique human genomes. This DNA fingerprinting technique was first used in the 1980s to verify a person’s identity and continues to be central to forensic science.
Another major leap forward in understanding human disease was the advent of knockout mice.
Mario Capecchi in the lab in the 1980s [Courtesy of Mario Capecchi]
By the late 1980s, scientists had devised methods to deliver an inactive version of a gene of interest into mouse embryonic cells and obtain mice in which the functional gene was replaced with the inactive version. Thousands of types of knockout mice have since been made that have elucidated the genetic underpinnings of cancer, heart and neurodegenerative diseases, and other conditions. “The mouse will remain a workhorse for understanding human disease from cancer to neuropsychiatric disorders,” says Mario Capecchi, recipient of a 2001 Lasker Award
, along with Martin Evans and Oliver Smithies, for developing methods to engineer knockout mice.
Looking Ahead to the Next Advances
As early as the 1960s, before some of the greatest breakthroughs in molecular biology were even on the horizon, scientists were saying the field was dead. Christiane Nusslein-Volhard, recipient of a 1991 Lasker Award, recalls hearing that prediction when she was working toward her PhD in the early 1970s.
But as it turned out, of course, molecular biology provided the scientific community with both groundbreaking insights and puzzles so confounding that they are still being worked out. Even the most fundamental biological processes, such as gene transcription, are not fully understood, says Stephen Elledge, recipient of a 2015 Lasker Award. Many goals of molecular biology, such as identifying associations between DNA sequence variants and disease, remain the same but have expanded into more complex systems, such as neuropsychiatric diseases, Maniatis says. In some cases, as researchers delve deeper into one aspect of molecular biology, they find connections with other areas. In her ongoing quest to elucidate how telomere length is controlled, Carol Greider, recipient of a 2006 Lasker Award, has ventured into research on DNA replication and how that process is linked to telomere regulation.
DNA molecule [Courtesy Zephyris, CC BY-SA 3.0, via Wikimedia Commons]
There continue to be enormous technological advances. Topping the list is CRISPR, a gene editing tool developed in the early 2000s. “[It] is really the most exciting thing that’s happened in the last few years for sure. I wish the technique had been available when I was still at the bench,” says Lasker Laureate Brown. It has opened up the genome of any organism to mutating and editing, whereas previously such tools were only available for model organisms such bacteria and Drosophila
. “The ability now for people to manipulate the human genome is a different order of magnitude” than in the 1970s, when scientists held meetings to discuss concerns about recombinant DNA technology, says Jack Szostak, recipient of a 2006 Lasker Award
with Blackburn and Greider. (Not surprisingly, concerns about CRISPR have mounted in recent years after a Chinese scientist announced in 2019 that he had used the tool to edit human embryos.) Numerous other technological advances have enabled research that would have been unthinkable back in the days of Nirenberg cracking the genetic code. These include the ability to synthetize larger and larger portions of mammalian genomes from scratch; wield artificial intelligence to predict gene regulatory factor binding sites; and analyze single cells at a scale large enough to reveal how organisms develop.
It can be mind-boggling to think about all the discoveries that have been made about DNA, and thus the biology of living systems, in the last half-century. As Blackburn explains, this is how science and technology advance. “We can look at things in much more sophisticated ways. It is like other fields, take cosmology — they have the technology to look at gravitational waves, and now they can ask enormously interesting questions that were unaskable even a couple decades ago,” she says.
By Carina Storrs