He soon found one that was resistant, and then some. It’s called phi-kappa-zeta (or phiKZ)—a name that it coincidentally shares with a sorority. Unusually large for a virus, phiKZ typically infects a bacterium called Pseudomonas aeruginosa. Unsurprisingly, it could resist the version of CRISPR used by its host. Unexpectedly, it also resisted every other version of CRISPR that the team tried, including those from bacteria that it would never have naturally encountered. Its armor seemed to work against every possible weapon. No anti-CRISPR protein should work in such a universal way. “It didn’t make any sense,” Bondy-Denomy says.
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He slowly realized what was happening after chatting with David Agard, who works in the same building. In 2017, Agard, along with Joe Pogliano and other colleagues, discovered that another phage does something that viruses are not meant to do. It encapsulates its DNA inside a shell of protein, which it suspends inside itself with thin filaments. That’s exceptionally odd. The cells of animals and plants also house their DNA within a special compartment—the nucleus. But such compartments aren’t meant to exist in simpler cells, like those of bacteria. And they’re certainly not meant to exist in viruses, which some scientists don’t even regard as alive. And yet, here was a phage, packaging its DNA in something akin to a nucleus. Why?
Agard told Bondy-Denomy about the phage that surrounds its DNA in a shell. Bondy-Denomy told Agard about the phage that’s resistant to all forms of CRISPR. It slowly dawned on the duo that the viruses they were studying were closely related, and that the weird phenomena they had found were linked. CRISPR can’t destroy what it can’t reach, and the shell stops it from getting at the phage’s DNA. The phiKZ phage and its relatives don’t need to evolve countermeasures against each and every form of CRISPR when they have ways of excluding them all. “Proving it was the hard part,” Bondy-Denomy says. “That took a couple of years.”
His student Eliza Nieweglowska confirmed, using a microscope, that the CRISPR scissors really are blocked by the shell. Meanwhile, another student, Senén Mendoza, showed that once the phage’s DNA was removed from the shell, the CRISPR scissors were perfectly capable of cutting it up. Mendoza also managed to smuggle the scissors into the shell by fusing them with proteins that are normally allowed to pass. When that happened, the phages were destroyed. “This work definitively shows that the structure protects against CRISPR,” says Benjamin Chan, who studies phages at Yale University. The similarity to a nucleus “is fascinating,” he adds.
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Finding new forms of CRISPR, or new defenses against it, could lead to ways of controlling gene-editing technologies more carefully or efficiently. But on a more basic level, Bondy-Denomy’s discovery might hint at a bit of evolutionary history, of our own cells. If viruses can protect their DNA from bacterial enemies using a nucleus-like structure, perhaps the nucleus itself evolved as a way for cells to protect their DNA from viruses? To explore that idea, Bondy-Denomy and his team are trying to understand more about how the mysterious shell functions. It’s incredibly selective, blocking everything except the proteins that the virus uses to copy its DNA and switch on its genes. “It’s not clear how it works,” Bondy-Denomy says. “But we’re really in love with it now.”