The m⁶A Methylation-Regulated AFF4 Promotes Self-Renewal of Bladder Cancer Stem Cells

Area of Science:

• Epigenetics
• Cancer Biology
• Molecular Oncology

Background:

• N6-methyladenosine (m6A) mRNA modification regulates gene expression and cell fate.
• The role of m6A in bladder cancer stem cells (BCSCs) remains largely unexplored.
• m6A modification is a dynamic and reversible process influencing various cellular functions.

Purpose of the Study:

• To investigate the function of m6A mRNA modification in BCSCs.
• To elucidate the molecular mechanisms underlying m6A-mediated regulation in bladder cancer.
• To identify key molecules involved in m6A-driven BCSCs self-renewal and tumorigenicity.

Main Methods:

• Quantification of global RNA m6A abundance and METTL3 expression in BCSCs versus non-CSCs.
• Depletion of METTL3 to assess effects on BCSC self-renewal (ALDH activity, sphere formation).
• Investigation of METTL3-AFF4 interaction and AFF4's role in BCSC tumorigenicity.
• Analysis of AFF4 binding to SOX2 and MYC promoter regions.

Main Results:

• Global RNA m6A abundance and METTL3 expression are elevated in BCSCs compared to non-CSCs.
• METTL3 depletion significantly inhibits BCSC self-renewal and sphere-forming ability.
• METTL3 regulates AFF4 expression; AFF4 knockdown mimics METTL3 depletion effects.
• AFF4 promotes SOX2 and MYC transcription, crucial for BCSC functions and in vivo tumorigenicity.

Conclusions:

• m6A modification, particularly via METTL3, is essential for maintaining BCSCs self-renewal and tumorigenicity.
• A novel signaling axis, METTL3-AFF4-SOX2/MYC, is identified as a key regulator in BCSCs.
• Targeting the m6A pathway presents a potential therapeutic strategy for bladder cancer.