CRISPR is most commonly associated with making changes to the DNA; however, researchers at the Salk Institute have developed a new use for the gene editing tool. They hope to use their new CRISPR RNA editing technique to correct protein imbalances in cells of patients with dementia.
The researchers have named their new CRISPR RNA editing technique CasRx. Senior author of the paper and Helmsley Salk Fellow, Patrick Hsu, said “CRISPR has revolutionized genome engineering, and we wanted to expand the toolbox from DNA to RNA.”
There has been considerable research into the use of CRISPR to eliminate or replace genes known to cause a wide range of genetic diseases. The Cas9 component of the technology is the enzyme used to make the cuts to double stranded DNA allowing defective genes to be removed or new genes to be inserted.
In order to develop the technique for use on RNA, the researchers needed to identify a bacterial enzyme that targets RNA instead of DNA. The researchers developed a program to search through bacterial DNA databases to look for the signatures of CRISPR enzymes that targeted RNA. They found one suitable candidate which they named Cas13d.
In contrast to CRISPR-Cas9 which is used by Streptococcus pyogenes, CRISPR-Cas13d is used by the gut bacterium Ruminococcus flavefaciens XPD3002. The researchers engineered their CRISPR-Cas13d based CasRx tool to work in human cells and investigated its potential for making changes to RNA.
“We began the project with the hypothesis that different CRISPR systems may have been specialized throughout an evolutionary arms race between bacteria and their viruses, potentially giving them the ability to target viral RNA,” said Salk Research Associate Silvana Konermann.
The researchers demonstrated that CasRx could be used to splice RNA in different ways to make different proteins. Splicing problems can lead to diseases such as atypical cystic fibrosis and frontotemporal dementia (FTD). In the latter there is an imbalance in two forms of the tau protein in neurons. The researchers used CasRx to address that imbalance and attempted to return the proteins to the correct ratio.
The team packed CasRx into a virus and introduced it into extracted neuron cells of patients with FTD. The team reports that their CasRx technique was 80% effective at balancing the levels of tau proteins and returning them to normal levels, with no detectable off-target effects.
The research is detailed in the paper –Transcriptome Engineering with RNA-Targeting Type VI-D CRISPR Effectors – recently published in the journal Cell.