N⁶ -Methyladenosine-Sensitive RNA-Cleaving Deoxyribozymes

Area of Science:

• Biochemistry
• Molecular Biology
• Synthetic Biology

Background:

• Deoxyribozymes are DNA-based catalysts crucial for RNA biochemistry.
• N⁶ -methyladenosine (m⁶ A) is the most prevalent RNA modification, regulating various cellular processes.
• Biochemical validation of m⁶ A sites is essential for understanding its regulatory roles.

Purpose of the Study:

• To develop novel RNA-cleaving deoxyribozymes for interrogating RNA methylation status.
• To provide an alternative biochemical method for validating N⁶ -methyladenosine sites in RNA.
• To assess the sensitivity of deoxyribozymes to m⁶ A modifications near cleavage sites.

Main Methods:

• Design and synthesis of novel DNA enzymes (deoxyribozymes).
• Assaying the catalytic activity of deoxyribozymes on methylated and unmethylated RNA substrates.
• Testing the applicability of deoxyribozymes on natural RNA sequences, including lncRNA and snoRNAs.

Main Results:

• Developed deoxyribozymes sensitive to the presence of N⁶ -methyladenosine.
• Observed differential activity: some deoxyribozymes showed enhanced cleavage of methylated RNA, while others were inhibited.
• Demonstrated general applicability on natural RNA targets like lncRNA and C/D box snoRNAs.

Conclusions:

• The new deoxyribozymes serve as effective tools for biochemical validation of RNA methylation.
• These DNA enzymes offer a sensitive and versatile approach to study N⁶ -methyladenosine modifications.
• The findings facilitate further research into the regulatory functions of m⁶ A in RNA folding and protein interactions.