Structural Control of RNA Demethylation: G-Quadruplex Proximity Suppresses ALKBH5 Activity.

Area of Science:

• Molecular Biology
• RNA Biology
• Epigenetics

Background:

• RNA modifications, like N6-methyladenosine (m6A), and RNA structures, such as G-quadruplexes, are crucial regulatory layers in RNA metabolism.
• The interplay between RNA modifications and higher-order structures, particularly their impact on regulatory protein activity, is not well understood.
• m6A is the most abundant internal mRNA modification, regulated by writer, reader, and eraser proteins, including ALKBH5 and FTO.

Purpose of the Study:

• To investigate how G-quadruplex RNA structures influence the demethylation efficiency of m6A eraser proteins.
• To determine if G-quadruplex formation affects the activity of ALKBH5 and FTO differently.

Main Methods:

• Synthesized RNA molecules with m6A modifications positioned adjacent to G-quadruplex cores or within flexible loops.
• Assessed the demethylation activity of ALKBH5 and FTO on these modified RNAs.
• Utilized the G-quadruplex-specific ligand BRACO19 to stabilize G-quadruplex structures and observe its effect on ALKBH5 activity.

Main Results:

• G-quadruplex formation significantly suppressed ALKBH5 activity compared to FTO, particularly when m6A was located near the G-quadruplex core.
• ALKBH5 activity was maintained when m6A was situated within a flexible loop of the G-quadruplex structure.
• Stabilization of G-quadruplex structures with BRACO19 confirmed the structural sensitivity of ALKBH5.

Conclusions:

• This study demonstrates a direct link between G-quadruplex structures and m6A eraser protein activity, highlighting the structural sensitivity of ALKBH5.
• RNA structural context plays a critical role in regulating the efficiency of m6A demethylation.
• G-quadruplexes may function as regulatory elements in epitranscriptomic control by modulating m6A eraser activity.