A team from Montana State University published research showing how RNA, the close chemical cousin of DNA, can be edited using CRISPR. The work reveals a new process in human cells that has the potential to treat a wide variety of genetic diseases.
Postdoctoral researchers Artem Nemudryi and Anna Nemudraia conducted the research along with Blake Wiedenheft, a professor in the Department of Microbiology and Cell Biology in the MSU College of Agriculture. The paper, titled “CRISPR-guided RNA break repair enables site-specific RNA cleavage in human cells,” was published online in the journal. Science and marks the latest advance in the team’s ongoing exploration of CRISPR applications for programmable genetic engineering.
CRISPR, which stands for clustered regularly interspaced short palindromic repeats, is a type of immune system that bacteria use to recognize and fight viruses. Wiedenheft, one of the country’s leading CRISPR researchers, said the system has been used for years to cut and edit DNA, but applying similar technology to RNA is unprecedented. DNA editing uses a CRISPR-associated protein called Cas9, while RNA editing requires the use of a different CRISPR system, called type III.
“In our previous work, we used type III CRISPR to edit viral RNA in a test tube,” Nemudryi said. “But we asked ourselves: can we program RNA manipulation into a living human cell?”
To explore that question, the team programmed CRISPR type III proteins to cut RNA containing a mutation that causes cystic fibrosis, restoring cellular function.
“We were confident that we could use these CRISPR systems to cut RNA in a programmable way, but we were all surprised when we sequenced the RNA and realized that the cell had rejoined the RNA in a way that eliminated the mutation,” Wiedenheft said.
Nemudryi observed that RNA is transient within the cell; It is constantly being destroyed and replaced.
“The general belief is that it doesn’t make much sense to repair RNA,” he said. “We speculated that the RNA would be repaired in living human cells and it turned out to be true.”
Wiedenheft has been a mentor to the two postdoctoral researchers since their arrival at MSU nearly six years ago and said the impact of their scientific contributions will lead to significant and continued advances.
“The work done by Artem and Anna suggests that RNA repair could be a fundamental aspect of biology and that harnessing this activity can lead to new, life-saving cures,” Wiedenheft said. “Artem and Anna are two of the most brilliant scientists I have ever met and I am confident that their work will have a lasting impact on humanity.”
RNA editing has important applications in finding treatments for genetic diseases, Nemudryi said. RNA is a temporary copy of a cell’s DNA, which serves as a template. Manipulating the template by editing the DNA could cause unwanted and potentially irreversible collateral changes, but because the RNA is a temporary copy, she said, the edits made are essentially reversible and carry much less risk.
“People used Cas9 to break DNA and study how cells repair these breaks. Then, based on these patterns, they improved Cas9 editors,” Nemudraia said. “Here we hope the same thing will happen with RNA editing. We created a tool that allows us to study how cells repair their RNA and we hope to use this knowledge to make RNA editors more efficient.”
In the new publication, the team shows that a mutation that causes cystic fibrosis can be successfully removed from RNA. But this is just one of thousands of known disease-causing mutations. The question of how many of them could be addressed with this new RNA editing technology will guide Nemudryi and Nemudraia’s future work as they finish their postdoctoral training at MSU and prepare for faculty positions at the University of Florida this fall. Both recognized Wiedenheft as a mentor who changed lives.
“Blake taught us not to be afraid to try any idea,” Nemudraia said. “As a scientist, you have to be brave and not be afraid to fail. RNA editing and repair is terra incognita. It’s scary but it’s also exciting. You feel like you’re working at the edge of science, pushing the limits where no one else has made”. been before.”