Enhanced or reversible RNA N6-methyladenosine editing by red/far-red light induction

Area of Science:

• Molecular Biology
• Optogenetics
• RNA Epigenetics

Background:

• N6-methyladenosine (m6A) is a crucial RNA modification regulating biological processes.
• Achieving precise and dynamic control over m6A levels is a significant challenge in RNA biology.

Purpose of the Study:

• To develop a red/far-red light-inducible system for efficient and reversible m6A editing.
• To enable targeted modulation of m6A deposition and removal on specific RNA transcripts.

Main Methods:

• Engineered CRISPR dCas13 and sgRNA with light-inducible heterodimerizing proteins (ΔphyA/FHY1 and Bphp1/PspR2).
• Utilized optogenetics for targeted recruitment of m6A effectors.
• Demonstrated cyclic modulation of m6A at a single target site.

Main Results:

• Achieved efficient and reversible modulation of m6A levels with minimal off-target effects.
• Successfully regulated m6A deposition and removal on target transcripts (e.g., SOX2, ACTB).
• Showcased influence on mRNA expression and human embryonic stem cell differentiation.

Conclusions:

• The developed optogenetic platform provides precise and versatile control over m6A.
• This system has broad implications for RNA biology research and potential therapeutic applications.