Essential role of ALKBH5-mediated RNA demethylation modification in bile acid-induced gastric intestinal metaplasia

Bile acid reflux and subsequent caudal-related homeobox 2 (CDX2) activation contribute to gastric intestinal metaplasia (IM), a precursor of gastric cancer; however, the mechanism underlying this phenomenon is unclear. Here, we demonstrate that alkylation repair homolog protein 5 (ALKBH5), a major RNA N⁶-adenosine demethylase, is required for bile acid-induced gastric IM. Mechanistically, we revealed the N⁶-methyladenosine (m⁶A) modification profile in gastric IM for the first time and identified ZNF333 as a novel m⁶A target of ALKBH5. ALKBH5 was shown to demethylate ZNF333 mRNA, leading to enhanced ZNF333 expression by abolishing m⁶A-YTHDF2-dependent mRNA degradation. In addition, ALKBH5 activated CDX2 and downstream intestinal markers by targeting the ZNF333/CYLD axis and activating NF-κB signaling. Reciprocally, p65, the key transcription factor of the canonical NF-κB pathway, enhanced the transcription activity of ALKBH5 in the nucleus, thus forming a positive feedforward circuit. Furthermore, ALKBH5 levels were positively correlated with ZNF333 and CDX2 levels in IM tissues, indicating significant clinical relevance. Collectively, our findings suggest that an m⁶A modification-associated positive feedforward loop between ALKBH5 and NF-κB signaling is involved in generating the IM phenotype of gastric epithelial cells. Targeting the ALKBH5/ZNF333/CYLD/CDX2 axis may be a useful therapeutic strategy for gastric IM in patients with bile regurgitation.