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January 30 - April 24, 2022
The men who had sequenced DNA taught us how to read the code of life, but the more important step would be learning how to write that code. This would require a different set of tools, ones that would involve the worker-bee molecule that Doudna found more interesting than DNA.
RNA (ribonucleic acid) is a molecule in living cells that is similar to DNA (deoxyribonucleic acid), but it has one more oxygen atom in its sugar-phosphate backbone and a difference in one of its four bases.
But like many famous siblings, DNA doesn’t do much work. It mainly stays at home in the nucleus of our cells, not venturing forth. Its primary activity is protecting the information it encodes and occasionally replicating itself.
A small segment of DNA that encodes a gene is transcribed into a snippet of RNA, which then travels to the manufacturing region of the cell. There this “messenger RNA” facilitates the assembly of the proper sequence of amino acids to make a specified protein.
These proteins come in many types. Fibrous proteins, for example, form structures such as bones, tissues, muscles, hair, fingernails, tendons, and skin cells. Membrane proteins relay signals within cells. Above all is the most fascinating type of proteins: enzymes. They serve as catalysts. They spark and accelerate and modulate the chemical reactions in all living things.
If some RNA molecules could store genetic information and also act as a catalyst to spur chemical reactions, they might be more fundamental to the origins of life than DNA, which cannot naturally replicate themselves without the presence of proteins to serve as a catalyst.
“I was quite surprised to find that there was this young genius, Jack Szostak, at Harvard who wanted to focus a hundred percent on RNA because he thought that it was the key to understanding the origin of life.”
Szostak and Doudna pursued the subject out of pure curiosity about how nature works.
Szostak had a guiding principle: Never do something that a thousand other people are doing.
“I learned from Jack that there was more of a risk but also more of a reward if you ventured into a new area.”
in addition to taking risks by moving into new fields: Ask big questions
The central dogma of biology requires the presence of DNA, RNA, and proteins. Because it’s unlikely that all three of these sprang forth at the exact same time from the primordial stew, a hypothesis arose in the early 1960s—formulated independently by the ubiquitous Francis Crick and others—that there was a simpler precursor system. Crick’s hypothesis was that, early on in the history of earth, RNA was able to replicate itself. That leaves the question of where the first RNA came from. Some
An essential quality of living things is that they have a method for creating more organisms akin to themselves: they can reproduce.
Eventually, she and Szostak were able to engineer a ribozyme that could splice together a copy of itself. “This reaction demonstrates the feasibility of RNA-catalyzed RNA replications,”
Barbara McClintock, who had been a researcher at Cold Spring Harbor for more than forty years and had recently been awarded the Nobel Prize for her discovery of transposons, known as “jumping genes,” that can change their position in a genome.
She learned how to pause, like we all used to do as children, and wonder about how things worked.
structural biologists try to discover the three-dimensional shape of molecules.
Doudna realized that she would need to learn more about structural biology if she wanted to truly understand how some RNA molecules could reproduce themselves.
a slight blunder, like the mold that got on Alexander Fleming’s Petri dishes and led to the discovery of penicillin.
RNA is made up of very few chemicals, so it accomplishes complex tasks based on the different ways it is folded. One of the challenges with RNA is that it’s a molecule made of only four chemical building blocks, unlike proteins, which have twenty. “Because there is a lot less chemical complexity to RNA,” she says, “the challenge is to think about how does it fold into a unique shape.”
the structure discovered by Doudna and her team explained how the RNA could be an enzyme and was able to slice, splice, and replicate itself.
“We hope our discovery will provide clues as to how we might be able to modify the ribozyme so that it can repair defective genes,”
Specifically, she was interested in how the RNA in some viruses, such as coronaviruses, allow them to hijack the protein-making machinery of cells.
Many viruses are composed of DNA, but SARS was a coronavirus that instead contained RNA.
Doudna also became interested in a phenomenon known as RNA interference. Normally, the genes encoded by the DNA in cells dispatch messenger RNAs to direct the building of a protein. RNA interference does just what the name implies: small molecules find a way to mess with these messenger RNAs.
RNA interference operates by deploying an enzyme known as “Dicer.” Dicer snips a long piece of RNA into short fragments. These little fragments can then embark on a search-and-destroy mission: they seek out a messenger RNA molecule that has matching letters, then they use a scissors-like enzyme to chop it up. The genetic information carried by that messenger RNA is thus silenced.
“Perhaps the most exciting finding of this study is that Dicer can be reengineered,” their 2006 paper noted.4 It was a very useful discovery. It permitted researchers to use RNA interference to turn off a wide variety of genes, both to discover what each gene does and to regulate its activity for medical purposes.
Throughout the history of life on our planet, some organisms (though not humans) have evolved ways to use RNA interference to fight off viruses.
The hope then was that drugs based on RNA interference might someday be a good option for treating severe viral infections, including those from new coronaviruses.
The scientist does not study nature because it is useful. He studies it because he takes pleasure in it, and he takes pleasure in it because it is beautiful. —Henri Poincaré, Science and Method, 1908
“An unusual structure was found,” he wrote. “Five highly homologous sequences of 29 nucleotides were arranged as direct repeats.” In other words, he found five segments of DNA that were identical to each other. These repeated sequences, each twenty-nine base pairs long, were sprinkled between normal-looking sequences of DNA, which he called “spacers.”
The first researcher to figure out the function of the repeated sequences was Francisco Mojica, a graduate student at the University of Alicante on the Mediterranean coast of Spain.
The archaea he was studying thrive in salt ponds that are ten times saltier than the ocean.
The E. coli bacterium that Ishino studied is a very different organism from Mojica’s archaea. So it was surprising that they both had these repeated sequences and spacer segments. This convinced Mojica that the phenomenon must have some important biological purpose.
But he knew that bacteria and archaea have small amounts of genetic material. They cannot afford to waste a lot of it on sequences that have no important function.
Mojica was driving home from his lab one evening when he came up with the name CRISPR, for “clustered regularly interspaced short palindromic repeats.”
discovery of genes that seemed to be associated with CRISPRs. In most organisms that had CRISPRs, the repeated sequences were flanked by one of these genes, which encoded directions for making an enzyme. He named these “CRISPR-associated,” or Cas, enzymes.
What he found was intriguing: the spacer segments matched sequences that were in viruses that attacked E. coli. He found the same thing when he looked at other bacteria with CRISPR sequences; their spacer segments matched those of viruses that attacked that bacteria.
“Bacteria have an immune system. They’re able to remember what viruses have attacked them in the past.”
What Mojica had stumbled upon was a battlefront in the longest-running, most massive and vicious war on this planet: that between bacteria and the viruses, known as “bacteriophages” or “phages,” that attack them. Phages are the largest category of virus in nature. Indeed, phage viruses are by far the most plentiful biological entity on earth.
“When you do curiosity-driven research, you never know what it may someday lead to,” Mojica says. “Something that’s basic can later have wide consequences.”
RNA. “It’s such a versatile molecule—it can do catalysis, it can fold into 3D structures,” he later told Kevin Davies of the CRISPR Journal. “At the same time, it’s a carrier of information. It’s an all-rounder in the world of biomolecules!”
Enzymes are a type of protein. Their main function is to act as a catalyst that sparks chemical reactions in the cells of living organisms, from bacteria to humans. There are more than five thousand biochemical reactions that are catalyzed by enzymes.
These CRISPR-associated (Cas) enzymes enable the system to cut and paste new memories of viruses that attack the bacteria. They also create short segments of RNA, known as CRISPR RNA (crRNA), that can guide a scissors-like enzyme to a dangerous virus and cut up its genetic material. Presto! That’s how the wily bacteria create an adaptive immune system!
They discovered that Cas1 has a distinct fold, indicating that it is the mechanism that bacteria use to cleave a snippet of DNA from invading viruses and incorporate it into their CRISPR array, thus being the key to the memory-forming stage of the immune system.
“It is just as often the case that invention is the parent of science: techniques and processes are developed that work, but the understanding of them comes later,” he writes. “Steam engines led to the understanding of thermodynamics, not the other way round. Powered flight preceded almost all aerodynamics.”
Starter cultures for yogurt and cheese are made from bacteria, and the greatest threats to the $40 billion global market are viruses that can destroy bacteria.
starting with Streptococcus thermophilus, the bacteria that is the great workhorse of the dairy culture industry.
That year Danisco started using CRISPR to vaccinate its bacterial strains.
Without those CRISPR conferences, the field would not have moved at the speed it has or be as collaborative,” Barrangou says. “The camaraderie would never have existed.”
