Mxd1-mediated transcriptional repression of FTO induces NLRP3-dependent pyroptosis in retinal microglia

Pyroptosis of retinal microglial cells represents a key pathological event in degenerative disorders of the retina, yet the molecular drivers of this process remain incompletely understood. This study investigates the molecular drivers of retinal microglial pyroptosis using primary murine retinal microglial cells and HMC3 cells exposed to lipopolysaccharide (LPS) and adenosine triphosphate (ATP). Transmission electron microscopy (TEM) assessed ultrastructural changes, while cell viability, pyroptosis-associated protein markers, and inflammatory cytokine secretion were evaluated via CCK-8, western blotting, and ELISA, respectively. The results showed that LPS/ATP treatment reduced FTO protein levels in both cell types, and enforced FTO expression reversed LPS/ATP-induced pyroptosis. LPS/ATP elevated Mxd1 levels, and Mxd1 protein bound to the FTO promoter to repress its activity. Mechanistically, FTO decreased both m⁶A abundance and stability of NLRP3 transcripts, thereby lowering NLRP3 expression and suppressing pyroptosis, and this effect required IGF2BP2 as an m⁶A reader that bound to the coding sequence (CDS) of NLRP3 mRNA to promote its stability. In conclusion, transcriptional repression of FTO driven by Mxd1 facilitates retinal microglial pyroptosis through modulation of NLRP3 in an m⁶A- and IGF2BP2-dependent manner.