“Switch Mechanism” in CRISPR Could Prevent Unwanted Side Effects
A new study published in Cell magazine and co-authored by CRISPR pioneer Jennifer Doudna has suggested a potential solution to the unwanted side-effects of using CRISPR in the body. The study details using a “switch” mechanism which could keep the Cas9 enzyme turned off until it reaches its target site.
Normally, CRISPR Cas9 can stay on after it’s intended use is complete, cutting genes unnecessarily. Using an engineering technique called circular permutation, the scientists created Cas9 variants called “ProCas9s” which can ensure CRISPR is only turned on when it gets to the targeted cells. These variants understand the cells they’re in based on proteases, enzymes which cut proteins.
The new technique rearranges the primary sequence of proteins, connecting the protein’s termini with a peptide linker, then cutting the sequence at a different position to create new termini. Not only is Cas9 highly malleable to circular permutation, but the enzyme was found to have several hotspots letting it be re-opened without affecting its functions.
The study found that tailored ProCas9s, when paired with the right guide RNA, detect viral proteases and launch a defence against viruses such as Zika and the West Nile Virus to kill infected host cells.
The researchers from Berkeley believe that ProCas9 could aid molecular screening and drug discovery. They intend to develop Cas9 future proteins for base editing and epigenetic modifications, and look to test whether the ProCas9s can be built into an entire synthetic immune system, or are sensitive to endogenous proteases so they can be used in cancer cells.