Microbiologist David Weiss and his colleagues at Emory University used small CRISPR RNA to home in on viral RNA, with the Cas proteins targeting and inhibiting the proper functioning of this RNA within eukaryotic cells.
The targeting resulted in the inhibition of viral protein production, their paper in PNAS reports. The CRISPR-Cas system was discovered in bacteria, where it functions as an adaptive immune system that spots and attacks viral nucleic acid.
In recent years it has revolutionized genomic engineering by allowing scientists to target specific DNA sequences in mammalian cells for splicing. The Emory scientists, including immunologist Arash Grakoui who studies HCV, used the Cas9 machinery from Francisella novicida bacteria.
“We wondered whether this Cas9 protein could be expressed in eukaryotic cells and we reprogrammed it to target an RNA of interest to us. We were keen on targeting RNA without having the potential confounding factor of DNA being targeted,” Weiss explained.
“We got around this by targeting an RNA virus [hepatitis C], and obviously there are many reasons you would like to target a viral genome and interrupt viral replication.”
Weiss had encountered Cas9 in the genome of F. novicida earlier in his career as a research postdoc in Stanford, but at the time this protein was an unknown. The new research shows that this is a versatile and portable system for targeting RNA inside eukaryotic cells, which could be programmed as an antiviral defense.
However, it is not clear if expressing Cas9 in humans would be safe, Weiss says. “This is not something we are going to jump into.”
The Emory researchers write that it is likely that the F. novicida machinery could be used to target diverse viruses, such as poliovirus, flavivirus, filovirus and paramyxovirus.
Weiss said “in theory you could imagine transiently expressing Cas9 and RNA in a person to target a virus, but we are very far away from anything like that. It might be conceivably possible, but there would be delivery and safety concerns.”
However, he believes that it might be possible to incorporate this antiviral machinery into the genome of livestock for public health reasons.
“If pigs are infected with a virus, the virus can in some cases be transmitted to humans. You could make these pigs resistant to infection and that would break the cycle of transmission to people. That is one way it might offer an antiviral benefit,” said Weiss.
Previously, Weiss had reported that Cas9 of F. novicida could target messenger RNA and turn down production of one of its own lipoproteins. This protein could otherwise alert the host immune response, so knocking down its levels allowed the bacteria to evade the host immune system.