The CRISPR-Cas9 gene editing tool allows the editing of genes with a high level of precision, but advances have been made that allow even greater control of when the edits are made. Researchers at the Karolinska Institute have recently demonstrated it is possible to control when the edits are made using light. The new system allows edits to be made in specific subsets of cells in cell culture at set points in time.
Several approaches can be used to activate gene editing at specific points in time. Gene editing could be initiated by introducing chemicals, but this approach requires those chemicals to be delivered to cells. The use of light is advantageous as it is minimally invasive, and it works much more rapidly.
The use of light to activate CRISPR-Cas9 is not new, as several researchers have developed Cas9 proteins that are activated by light. While this approach can be effective, creating proteins that are activated by light does not allow upgrades to be made to the system to improve efficiency or selectivity.
A better approach is to make the guide RNA (gRNA) component of the system light sensitive. The gRNA pairs with a specific part of the DNA and guides the Cas9 protein to the precise location where an edit needs to be performed. This approach makes it far easier to tweak and edit the system over time.
Researchers at the Karolinska Institute have developed a gRNA that includes a light sensitive component, which ‘cages’ the gRNA until it is activated by light. Without light activation, it cannot pair with the DNA so no edits will be performed.
The gRNA will only be released when it is exposed to a specific wavelength of light, which cleaves off the molecules that prevent binding with the DNA, thus initiating the gene editing process. The researchers tested their new gRNA in human cell cultures and zebrafish embryos. Once introduced, there was no sign of DNA editing until the cells were illuminated, after which the editing took place.
The researchers compared the level of RNA activity in the light activated system to a non-light activated RNA and found they could achieve 70% RNA activation with the light-activated system. The researchers used the light-activated RNA to make edits in just one eye of a zebrafish embryo by illuminating one eye and not the other.
It should also be possible to develop a system that allows multicolor activation of several gRNAs, which would allow specific genes to be edited simultaneously or at different times using different wavelengths of light
You can read more about the study in the paper – Advanced spatiotemporal control of CRISPR-Cas9 activity is demonstrated through the use of chemically modified photoactivatable guide RNA – which was recently published in the journal ASC Central Science. DOI: 10.1021/acscentsci.0c00350