Tailoring and reversing m6A editing with sequential RNA bioorthogonal chemistry

Area of Science:

• Molecular Biology
• Chemical Biology
• Epigenetics

Background:

• Current methods for N6-methyladenosine (m6A) RNA modification are often one-step processes.
• This limits the ability to dynamically and reversibly control m6A methylation at specific RNA sites.
• Existing bioorthogonal chemistry approaches for RNA modification are typically irreversible.

Purpose of the Study:

• To develop a novel multi-step system for iterative and reversible RNA modifications in living cells.
• To enable precise, site-specific control over m6A methylation.
• To overcome the limitations of conventional one-way bioorthogonal RNA modification methods.

Main Methods:

• Development of a sequential RNA bioorthogonal chemistry system.
• Utilizing an azide-based reagent (NAI-N3) for both cleavage and ligation reactions.
• Integration of the system within CRISPR-based frameworks for targeted RNA editing.

Main Results:

• Demonstration of iterative and reversible m6A modifications in living cells.
• Successful application in CRISPR frameworks for tailored m6A editing at targeted RNA sites.
• Overcoming the one-way restriction of traditional bioorthogonal methods.

Conclusions:

• The developed sequential protocol enables fine-tuned RNA regulation.
• Provides a versatile platform for exploring dynamic m6A function.
• Advances genetic and epigenetic research through precise RNA modification control.