A cytosine deaminase for programmable single-base RNA editing.

Area of Science:

• Molecular Biology
• Genetic Engineering
• Biotechnology

Background:

• Programmable RNA editing offers reversible control over RNA sequences for research and therapeutic applications.
• Previous work established adenosine-to-inosine (A-to-I) RNA editing using dCas13 fused with ADAR2.

Purpose of the Study:

• To develop a novel RNA editing system for cytidine-to-uridine (C-to-U) conversion.
• To expand the scope of targetable mutations and enable precise modulation of RNA.
• To demonstrate the application of the new system in cellular processes.

Main Methods:

• Engineered ADAR2 into a cytidine deaminase to create the RESCUE system.
• Applied RESCUE for targeted C-to-U RNA editing.
• Investigated RESCUE's ability to modulate phosphosignaling pathways.
• Demonstrated multiplexed C-to-U and A-to-I editing.

Main Results:

• Developed RESCUE, a novel RNA editor for specific C-to-U exchange.
• Doubled the number of mutations addressable by RNA editing.
• Successfully modulated phosphosignaling-relevant residues and drove β-catenin activation.
• Achieved multiplexed RNA editing by combining C-to-U and A-to-I activities.

Conclusions:

• RESCUE significantly expands the capabilities of programmable RNA editing.
• This technology holds promise for diverse applications in biological research and the treatment of genetic diseases.
• The ability to perform multiplexed editing opens new avenues for complex genetic manipulations.