Scientists manipulate CRISPR technology to alter individual RNA nucleotides within human cell structures
Scientists from the Broad Institute and MIT have engineered a new system for efficiently editing RNA in human cells, a breakthrough that could significantly advance research and disease treatment. The system, called RNA Editing for Programmable A to I Replacement, or REPAIR, allows for precise edits to single RNA nucleotides.
In a paper published today in Science, lead author Feng Zhang and his team detail the new CRISPR-based system. REPAIR can alter RNA without modifying the genome, offering a more flexible and potentially safer method for correction within cells.
REPAIR targets individual RNA letters, converting adenosines to inosines, and has the ability to reverse disease-causing mutations at the RNA level. The system can effectively reverse mutations with a G-to-A change, which are common in human diseases such as focal epilepsy, Duchenne muscular dystrophy, and Parkinson's disease.
Unlike DNA editing, which involves permanent changes to the genome, RNA editing offers a reversible fix, as RNA naturally degrades. Co-first author David Cox explains, "REPAIR can fix mutations without tampering with the genome."
To create REPAIR, the researchers screened the CRISPR-Cas13 enzyme family for potential "editor" candidates, selecting the most effective enzyme from Prevotella bacteria, called PspCas13b. They deactivated this enzyme while retaining its ability to bind to specific stretches of RNA, and fused it with a protein called ADAR2, which changes the letters A to I in RNA transcripts.
REPAIR seeks out a target sequence of RNA, and the ADAR2 element performs the base conversion without cutting the transcript or relying on the cell's native machinery. The team improved the editing system's specificity, reducing detectable off-target edits from 18,385 to 20 in the whole transcriptome. The upgraded version, REPAIRv2, consistently achieved the desired edit in 20 to 40 percent - and up to 51 percent - of a targeted RNA without signs of significant off-target activity.
To demonstrate REPAIR's therapeutic potential, the team corrected pathogenic mutations that cause Fanconi anemia and X-linked nephrogenic diabetes insipidus in human cells. The researchers plan to improve REPAIRv2's efficiency and package it into a delivery system for use in animal models.
In addition to REPAIR, the team is working on further tools for other types of nucleotide conversions. According to co-first author Jonathan Gootenberg, "There's immense natural diversity in these enzymes. We're always looking to harness the power of nature to carry out these changes."
The research, which was funded, in part, by the National Institutes of Health and the Poitras Center for Affective Disorders Research, will be made widely available for academic research through the Zhang lab's page on the plasmid-sharing website Addgene.
While the Enrichment Data does not specifically mention the REPAIR system, it highlights the broader potential of CRISPR technology for RNA editing and delivery, including applications in treating neurological disorders, genetic diseases, and cancer.
Students from various departments, particularly those focusing on science, engineering, and neuroscience, will find the tool called REPAIR interesting. This new CRISPR-based system allows for precise editing of single RNA nucleotides, a breakthrough in health sciences and technology. REPAIR targets RNA without modifying the genome, offering a more flexible and potentially safer method for correction within cells.
Researchers at MIT and the Broad Institute have discovered that REPAIR can reverse disease-causing mutations at the RNA level, effects that could significantly impact conditions like focal epilepsy, Duchenne muscular dystrophy, and Parkinson's disease. The system can effectively correct G-to-A mutations, which are common in numerous medical conditions.
Engineered from the CRISPR-Cas13 enzyme family, the most effective enzyme selected for REPAIR is PspCas13b from Prevotella bacteria. The enzyme is deactivated while retaining its ability to bind to specific stretches of RNA, then fused with a protein called ADAR2 that changes A to I in RNA transcripts.
The REPAIR system will be made widely available for academic research by the Zhang lab through Addgene, a plasmid-sharing website. This move encourages further development in education and self-development, as students and researchers can improve the system and explore other nucleotide conversions.
In addition to REPAIR, the team is working on additional tools for other types of nucleotide conversions, emphasizing the immense natural diversity in these enzymes. These advancements could lead to new therapies and treatments for various health and medical conditions, enhancing health-and-wellness and contributing to medical-condition research.
By leveraging the power of nature to carry out these changes, the research could open new therapeutic possibilities for genetic diseases, neurological disorders, and cancer, ultimately benefiting the wider scientific community and individual graduate students working in these fields.
This research underlines the significant impact that technology and education-and-self-development can have on the quality of life for millions affected by genetic disorders, ultimately advancing the state of health, technology, and science for the better.
