FOXO6, Stabilized by METTL3-Mediated m6A Modification, Accelerates Polycystic Ovary Syndrome Progression by Transcriptional Activation of TXNIP

Polycystic ovary syndrome (PCOS) is a prevalent endocrine disorder with complex pathophysiology and limited therapeutic options. To elucidate potential biomarkers and molecular mechanisms underlying PCOS, we analyzed gene expression profiles in ovarian tissues from dehydroepiandrosterone (DHEA)-induced PCOS mice and control mice by conducting mRNA sequencing. Transcription factor forkhead box O6 (FOXO6) drew our focus, as it exhibited higher expression in PCOS ovarian tissues (fold change ≈1.71, p < 0.05). Functional validation revealed that FOXO6 knockdown alleviated ovarian dysfunction accompanied by attenuated mitochondrial damage and reduced apoptosis in the ovarian tissues of PCOS mice. Meanwhile, high expression of FOXO6 was also observed in dihydrotestosterone (DHT)-treated granulosa cells. Post-FOXO6 downregulation, DHT-induced apoptosis and mitochondrial dysfunction were suppressed. Notably, the RNA level of FOXO6 was upregulated by the methyltransferase METTL3, which inhibited the decay of FOXO6 RNA by increasing the m6A modification of FOXO6 RNA. The inhibitory effects of METTL3 knockdown on apoptosis and mitochondrial dysfunction in granulosa cells were weakened by FOXO6 overexpression. Furthermore, transcriptomic profiling of DHT-treated granulosa cells with or without FOXO6 overexpression was conducted. This analysis yielded 386 upregulated genes and 582 downregulated genes, among which thioredoxin-interacting protein (TXNIP) was prominently upregulated in FOXO6-overexpressing cells (fold change ≈4.5, p < 0.05). FOXO6 transcriptionally activated TXNIP by binding to its promoter, and TXNIP knockdown reversed the effects of FOXO6 overexpression. Collectively, our findings highlight the METTL3/FOXO6/TXNIP axis as a vital regulator of ovarian dysfunction in PCOS, offering novel insights for therapeutic intervention. Graphical To elucidate PCOS biomarkers and mechanisms, mRNA-sequencing of ovarian tissues from DHEA-induced PCOS and control mice identified highly expressed FOXO6 (FC ≈1.71, p < 0.05). FOXO6 knockdown alleviated PCOS ovarian dysfunction, mitochondrial damage, and apoptosis, and suppressed DHT-induced granulosa cell defects. METTL3 upregulated FOXO6 via m6A modification to inhibit its RNA decay. FOXO6 activated TXNIP transcription by binding its promoter. This highlights the METTL3/FOXO6/TXNIP axis as a key PCOS ovarian dysfunction regulator, offering new insights for PCOS therapy.