Researchers at University of California San Francisco have modified the architecture of the CRISPR gene editing tool to target the epigenome: The proteins and small molecules that are used to switch genes off and on.
Until now, CRIPSR has only been used by researchers to edit or replace faulty genes. The novel CRIPSR system does not make any changes to the genome, instead it can be used to switch off virtually any gene in human cells. The new tool has two complimentary parts: CRISPRoff is used to switch off specific genes and CRISPRon is used to reverse that process. Once switched off, a gene will remain inert in all of the cell’s descendants until the gene is switched back on again.
Conventional CRISPR therapy does allow highly accurate edits to be made to the genome, but there is potential for off-target edits to be made which could be harmful. The new CRISPR system would avoid these problems as no changes are made to the genome.
Being able to switch genes off and on has important therapeutic benefits. Viral infections and cancer can affect the epigenome. For instance, tumor cells often have mutations that encode regulators of the epigenome which are involved in resistance to immune surveillance and chemotherapy. Having control over the epigenome could see the new CRISPR tools used alone or in combination with other conventional therapies to improve their effectiveness.
“Though genetic and cellular therapies are the future of medicine, there are potential safety concerns around permanently changing the genome, which is why we’re trying to come up with other ways to use CRISPR to treat disease,” said Luke Gilbert, PhD, a professor at the UCSF Helen Diller Family Comprehensive Cancer Center and co-senior author of the new paper.
The conventional CRISPR gene editing tool consists of two main parts, one which cuts the DNA to allow edits to be made and the other is the part that can be programmed to home in on a very specific section of DNA to make the cut. The researchers removed the DNA cutting component but kept the part that allows a specific section of DNA to be targeted. An enzyme was then attached in the place of the DNA cutting molecule that targets the epigenome.
The new tool acts on a feature of the epigenome called DNA methylation. DNA methylation occurs naturally and involves adding a methyl group to DNA, which silences nearby genes. CRISPRoff adds a methyl group tag to silence genes, and CRISPRon removes the methyl group to reactivate the gene. Any gene that is switched off can have that process easily reversed. The system can be used to switch off single genes or multiple genes in the same treatment.
One problem with their approach is it is only possible to methylate two of the four nucleobases in DNA – C and G. Since it was thought that it would only be possible to silence genes using methylation if there are high concentrations of CG sequences – termed CpG islands – it was thought that the tool could only be used to silence around 70% of genes, as around 30% do not have CpG islands. That proved not to be the case.
“What was thought before this work was that the 30 percent of genes that do not have CpG islands were not controlled by DNA methylation,” said Gilbert. “But our work clearly shows that you don’t require a CpG island to turn genes off by methylation. That, to me, was a major surprise.”
The researchers demonstrated the genes that were switched off remained inert for as many as 450 generations, even in maturing stem cells, which saw the methylation inherited in a significant fraction of differentiated adult cells. That means that one treatment would have long lasting effects.
The researchers suggest their tool could be used in the treatment of cancers caused by the activity of a single damaged gene, and for treating rare genetic disorders such as Job’s syndrome and Marfan syndrome.
You can read more about the new CRIPR tool in the paper – Genome-wide programmable transcriptional memory by CRISPR-based epigenome editing – which was recently published in the journal Cell. DOI: 10.1016/j.cell.2021.03.025