More on this book
Community
Kindle Notes & Highlights
Read between
January 20 - February 3, 2024
Perhaps we should develop some rules.
Figuring out if and when to edit our genes will be one of the most consequential questions of the twenty-first century,
kernels of our existence: the atom, the bit, and the gene.
The first half of the twentieth century, beginning with Albert Einstein’s 1905 papers on relativity and quantum theory, featured a revolution driven by physics.
The second half of the twentieth century was an information-technology era, based on the idea that all information could be encoded by binary digits—known as bits—and
Now we have entered a third and even more momentous era, a life-science revolution. Children who study digital coding will be joined by those who study genetic code.
Doudna was a graduate student in the 1990s, other biologists were racing to map the genes that are coded by our DNA. But she became more interested in DNA’s less-celebrated sibling, RNA. It’s the molecule that actually does the work in a cell by copying some of the instructions coded by the DNA and using them to build proteins.
science, which is humanity’s attempt to understand the longest-running mystery we know: the origin and function of the natural world and our place in it.”5
Darwin and Wallace had a key trait that is a catalyst for creativity: they had wide-ranging interests and were able to make connections between different disciplines.
common substance in living cells, nucleic acids, that are the workhorses of heredity. These molecules are composed of a sugar, phosphates, and four substances called bases that are strung together in chains. They come in two varieties: ribonucleic acid (RNA) and a similar molecule that lacks one oxygen atom and thus is called deoxyribonucleic acid (DNA).
The easiest of these viruses to study are the ones that attack bacteria, and they were dubbed (remember the term, for it will reappear when we discuss the discovery of CRISPR) “phages,” which was short for “bacteriophages,” meaning bacteria-eaters.
DNA may be the world’s most famous molecule, so well-known that it appears on magazine covers and is used as a metaphor for traits that are ingrained in a society or organization. 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. RNA, on the other hand, actually goes out and does real work. Instead of just sitting at home curating information, it makes real products, such as proteins. Pay attention to it. From CRISPR
...more
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. Almost every action that takes place in a cell needs to be catalyzed by an enzyme. Pay attention to enzymes. They will be RNA’s costars and dancing partners in this book.
One of the first tweaks to the central dogma came when Thomas Cech and Sidney Altman independently discovered that proteins were not the only molecules in the cell that could be enzymes. In work done in the early 1980s that would win them the Nobel Prize, they made the surprising discovery that some forms of RNA could likewise be enzymes. Specifically, they found that some RNA molecules can split themselves by sparking a chemical reaction. They dubbed these catalytic RNAs “ribozymes,” a word conjured up by combining “ribonucleic acid” with “enzyme.”
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.
he spotted fourteen identical DNA sequences that were repeated at regular intervals.
palindromes, meaning they read the same backward and forward.2
CRISPR, for “clustered regularly interspaced short palindromic repeats.”
the CRISPR-associated (Cas) enzymes.
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 include breaking down starches and proteins in the digestive system, causing muscles to contract, sending signals between cells, regulating metabolism, and (most important for this discussion) cutting and splicing DNA and RNA.
As Marraffini and Sontheimer realized, if the CRISPR system was aimed at the DNA of viruses, then it could possibly be turned into a gene-editing tool.
Researchers had shown that if you deactivated Cas9 in bacteria, the CRISPR system no longer cut up the invading viruses. They had also established the essential role of another part of the complex: CRISPR RNAs, known as crRNAs. These are the small snippets of RNA that contain some genetic coding from a virus that had attacked the bacteria in the past. This crRNA guides the Cas enzymes to attack that virus when it tries to invade again. These two elements are the core of the CRISPR system: a small snippet of RNA that acts as a guide and an enzyme that acts as scissors.
turns out that tracrRNA performs two important tasks. First, it facilitates the making of the crRNA, the sequence that carries the memory of a virus that previously attacked the bacteria. Then it serves as a handle to latch on to the invading virus so that the crRNA can target the right spot for the Cas9 enzyme to chop.
Now Charpentier had a hypothesis: it is this tracrRNA that directs the creation of the short crRNAs.
Charpentier’s little team discovered that the CRISPR-Cas9 system accomplished its viral-defense mission using only three components: tracrRNA, crRNA, and the Cas9 enzyme. The tracrRNA took long strands of RNA and processed them into the small crRNAs that were targeted at specific sequences in an attacking virus.
In her 2011 paper Charpentier showed that tracrRNA was required for producing the crRNA guide. She later said that she suspected it played an even larger, ongoing role, though that possibility had not been part of their initial round of experiments. When those experiments failed, Chylinski decided to throw tracrRNA into his test-tube mix. It worked: the three-component complex reliably chomped up the target DNA. Jinek immediately told Doudna the news: “Without the tracrRNA, the crRNA guide does not bind to the Cas9 enzyme.”
This amazing little system, it quickly became clear, had a truly momentous potential application: the crRNA guide could be modified to target any DNA sequence you might wish to cut. It was programmable. It could become an editing tool.
“Once we figured out the components of the CRISPR-Cas9 assembly, we realized that we could program it on our own,” Doudna says. “In other words, we could add a different crRNA and get it to cut any different DNA sequence we chose.”
As they brainstormed, it became clear to them that they could link the two RNAs together, fusing the tail of one to the head of the other in a way that would keep the combined molecule functional. Their goal was to engineer a single RNA molecule that would have the guide information on one end and the binding handle on the other. That would create what they ended up calling a “single-guide RNA” (sgRNA).
So far Doudna’s collaboration with Charpentier had produced two significant advances. The first was the discovery that the tracrRNA played an essential role not just in creating the crRNA guide but, more important, holding it together with the Cas9 enzyme and binding it all to the target DNA for the cutting process. The second was the invention of a way to fuse these two RNAs into a single-guide RNA.
The enzymes that can cut DNA or RNA are called “nucleases.”
TALENs (transcription activator–like effector nucleases),
But in the CRISPR system, the guide was not a protein but a snippet of RNA. This had a big advantage. With ZFNs and TALENs, you had to construct a new protein guide every time you wanted to target a different genetic sequence to cut; it was difficult and time consuming. But with CRISPR you merely had to fiddle with the genetic sequence of the RNA guide. A good student could do it quickly in a lab.
Much of the attention paid to CRISPR these days involves its potential to make inheritable (germline) edits in humans that will be passed along to all the cells of all of our future descendants and have the potential to alter our species. These edits are done in reproductive cells or early-stage embryos. This is what occurred with the CRISPR baby twins in China in 2018,
CRISPR is used to edit some, but not all, of the body (somatic) cells of a patient and make changes that will not be inherited.
the prospect of CRISPR doing great good is matched by its potential to bankrupt the healthcare system.
Doudna decided that making sickle-cell treatments affordable should become a mission of her Innovative Genomics Institute.
Cure Sickle Cell Initiative funded with $200 million.6 The primary scientific goal of the initiative is to find a method to edit the sickle-cell mutation inside of a patient without needing to extract bone marrow.
CRISPR has been used to fight cancer. China has been the pioneer in this field, and it is two or three years ahead of the United States in devising treatments and getting them into clinical trials.7
CRISPR is also being used as a detection tool to identify precisely what type of cancer a patient has.
diagnostic tools based on CRISPR that can be used on tumors to identify quickly and easily the DNA sequences associated with different types of cancers. Then precision treatments can be tailored for each patient.10
The third use of CRISPR editing that was underway by 2020 was to cure a form of congenital blindness.
Work is also underway on some more ambitious uses of CRISPR gene editing that could make us less vulnerable to pandemics, cancers, Alzheimer’s, and other diseases.
These and other experiences led her to join an effort funded by the U.S. Defense Department to find ways to protect against the misuse of CRISPR.1
2016 when James Clapper, the U.S. Director of National Intelligence, issued the agency’s annual “Worldwide Threat Assessment” and it included for the first time “genome editing” as a potential weapon of mass destruction.
Bondy-Denomy had become a professor at the University of California, San Francisco, and he collaborated with Doudna’s lab to show that the anti-CRISPRs could be delivered into human cells to modulate or stop CRISPR-Cas9 editing.6 It was a basic science discovery about the wonders of nature, showing how the amazing arms race between bacteria and viruses evolved.
Like Church’s lab at Harvard, it was asked to study how to use CRISPR to protect against nuclear radiation.
The geneticist Bentley Glass, in his address on becoming president of the American Association for the Advancement of Science in 1970, argued that the ethical problem was not that people would embrace these new genetic technologies but that they might reject them. “The right that must become paramount is the right of every child to be born with a sound physical and mental constitution,” he said. “No parents will have a right to burden society with a malformed or a mentally incompetent child.”
Princeton professor of Christian ethics Paul Ramsey, a prominent Protestant theologian, published Fabricated Man: The Ethics of Genetic Control. It is a turgid book with one vivid sentence: “Men ought not to play God before they learn to be men.”4 The social theorist Jeremy Rifkin, dubbed by Time America’s “foremost opponent of genetic engineering,” coauthored a book titled Who Should Play God? “Once, all of this could be dismissed as science fiction, the mad ravings of a Dr. Frankenstein,” he wrote. “No more. We are not in the Brave New World yet, but we are well along the road.”
So they issued a letter—which was signed by Berg, James Watson, Herbert Boyer, and others—calling for a “moratorium” on the creation of recombinant DNA until safety guidelines could be formulated.
