Five big mysteries about CRISPR’s origins
Where did it come from? How do organisms use it without self-destructing? And what else can it do?
Heidi Ledford
12 January 2017 Corrected: 13 January 2017
Francisco Mojica was not the first to see CRISPR, but he was probably the first to be smitten by it. He remembers the day in 1992 when he got his first glimpse of the microbial immune system that would launch a biotechnology revolution. He was reviewing genome-sequence data from the salt-loving microbe Haloferax mediterranei and noticed 14 unusual DNA sequences, each 30 bases long. They read roughly the same backwards and forwards, and they repeated every 35 bases or so. Soon, he saw more of them. Mojica was entranced, and made the repeats a focus of his research at the University of Alicante in Spain.
It wasn't a popular decision. His lab went years without funding. At meetings, Mojica would grab the biggest bigwigs he could find and ask what they thought of the strange little repeats. “Don't care about repeats so much,” he says that they would warn him. “There are many repeats in many organisms — we've known about them for years and still don't know how many of them work.”
Today, much more is known about the clustered, regularly interspaced short palindromic repeats that give CRISPR its name and help the CRISPR–Cas microbial immune system to destroy invading viruses. But although most in biomedicine have come to revere the mechanics of the system — particularly of a version called CRISPR–Cas9 — for the ways in which it can be harnessed to edit genes, Mojica and other microbiologists are still puzzling over some basic questions about the system and how it works. How did it evolve, and how did it shape microbial evolution? Why do some microbes use it, whereas others don't? And might it have other, yet-to-be-appreciated roles in their basic biology?
“A lot of the attention paid to CRISPR systems in the media has really been around its use as a technology — and with good reason. That's where we're seeing incredible impact and opportunities,” says Jennifer Doudna, a molecular biologist at the University of California, Berkeley, and one of the first scientists to reveal CRISPR–Cas's agility as a gene-editing tool. “At the same time, there's a lot of interesting fundamental biology research to be done.”
Where did it come from?
The biological advantages of something like CRISPR–Cas are clear. Prokaryotes — bacteria and less-well-known single-celled organisms called archaea, many of which live in extreme environments — face a constant onslaught of genetic invaders. Viruses outnumber prokaryotes by ten to one and are said to kill half of the world's bacteria every two days. Prokaryotes also swap scraps of DNA called plasmids, which can be parasitic — draining resources from their host and forcing it to self-destruct if it tries to expel its molecular hitch-hiker. It seems as if nowhere is safe: from soil to sea to the most inhospitable places on the planet, genetic invaders are present.
Prokaryotes have evolved a slew of weapons to cope with these threats. Restriction enzymes, for example, are proteins that cut DNA at or near a specific sequence. But these defences are blunt. Each enzyme is programmed to recognize certain sequences, and a microbe is protected only if it has a copy of the right gene. CRISPR–Cas is more dynamic. It adapts to and remembers specific genetic invaders in a similar way to how human antibodies provide long-term immunity after an infection. “When we first heard about this hypothesis, we thought that would be way too sophisticated for simple prokaryotes,” says microbiologist John van der Oost of Wageningen University in the Netherlands.
Mojica and others deduced the function of CRISPR–Cas when they saw that DNA in the spaces between CRISPR's palindromic repeats sometimes matches sequences in viral genomes. Since then, researchers have worked out that certain CRISPR-associated (Cas) proteins add these spacer sequences to the genome after bacteria and archaea are exposed to specific viruses or plasmids. RNA made from those spacers directs other Cas proteins to chew up any invading DNA or RNA that matches the sequence (see 'Lasting protection').
How did bacteria and archaea come to possess such sophisticated immune systems? That question has yet to be answered, but the leading theory is that the systems are derived from transposons — 'jumping genes' that can hop from one position to another in the genome. Evolutionary biologist Eugene Koonin of the US National Institutes of Health in Bethesda, Maryland, and his colleagues have found1 a class of these mobile genetic elements that encodes the protein Cas1, which is involved in inserting spacers into the genome. These 'casposons', he reasons, could have been the origin of CRISPR–Cas immunity. Researchers are now working to understand how these bits of DNA hop from one place to another — and then to track how that mechanism may have led to the sophistication of CRISPR–Cas.
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