FTO promotes post-stroke neuroprotection by m⁶A demethylation of c-Jun

N⁶-methyladenosine (m⁶A) is a critical epitranscriptomic regulator of neuronal function. Cerebral ischemia induces m⁶A hypermethylation due to decreased expression of m⁶A demethylase fat mass and obesity-associated (FTO) protein. Previously, we showed that cerebral overexpression of FTO with an adeno-associated virus (AAV) 9 protects the post-stroke brain. We presently evaluated the mechanistic basis for FTO-dependent m⁶A demethylation in post-ischemic neuroprotection using the mice transient middle cerebral artery occlusion model of experimental stroke. Based on the bioinformatic predictions and m⁶A abundance, pro-apoptotic transcription factor Jun proto-oncogene (c-Jun) with 19 m⁶A sites was chosen as an exemplary target. FTO overexpression normalized the post-stroke m⁶A hypermethylation of c-Jun without altering its transcript levels. FTO-dependent m⁶A demethylation suppressed translation of c-Jun. Consequently, several c-Jun target genes are transcriptionally repressed, and the post-ischemic neuronal apoptosis is decelerated, as seen by decreased cleaved caspase-3 levels and TUNEL⁺ neurons in the FTO AAV9 treated group compared to the control AAV9 treated group. Moreover, replenishing c-Jun precluded the FTO-mediated post-stroke neuroprotection and functional recovery. Collectively, this study demonstrated that the FTO/m⁶A/c-Jun axis ameliorates post-stroke neuronal apoptosis and brain damage, leading to better functional outcomes.