Meclofenamic acid represses spermatogonial proliferation through modulating m⁶A RNA modification

Area of Science:

• Epigenetics and Gene Regulation
• Reproductive Biology
• Molecular Mechanisms of Cell Division

Background:

• N6-Methyladenosine (m⁶A) is a prevalent mRNA modification crucial for biological processes.
• While several m⁶A-binding proteins regulate germ cell development, the role of the m⁶A demethylase FTO in germ cells is largely unknown.
• Understanding FTO's function in germ cells is vital for insights into mammalian reproduction.

Purpose of the Study:

• To investigate the role of the m⁶A demethylase FTO in spermatogonial proliferation.
• To elucidate the molecular mechanisms underlying FTO's regulation of germ cell fate.

Main Methods:

• Inhibition of FTO demethylase activity in mouse GC-1 spg cells using meclofenamic acid (MA2).
• Quantification of cellular m⁶A and m⁶A_m levels via HPLC/MS-MS.
• Assessment of cell apoptosis, proliferation, mRNA stability, and cyclin-dependent kinases (CDKs) expression.

Main Results:

• MA2 treatment significantly increased cellular m⁶A levels and downregulated CDK1, CDK2, CDK6, and CdC25a expression.
• FTO inhibition led to G1/S phase arrest, decreased cell proliferation, and reduced CDK2 mRNA stability.
• Mutation of predicted m⁶A sites in the CDK2 3'UTR partially rescued mRNA degradation following MA2 treatment.

Conclusions:

• MA2 affects CDK expression via an m⁶A-dependent mRNA degradation pathway.
• FTO inhibition represses spermatogonial proliferation by modulating CDK expression and mRNA stability.
• This study highlights FTO's critical role in regulating spermatogonial proliferation through epigenetic mechanisms.