ALKBH5 demethylates the m <sup>6</sup> A modification of SOCS3 in microglia/macrophages and alleviates neuroinflammation after brain injury
Lin Cai, Yuqing Liang, Xiaoyu Li, Runxi Fu, Xingyu Niu, Yuxiao Jin, Yuxin Li, Yuheng Zhang, Pei Ouyang, Chen Wang, Qiuyuan Gong, Yang Yang, Wei Li, Jing Yao, Dianxu Yang, Zhiming Xu, Fang Yuan, Jun Ding, Hao Chen, Bo Peng, Yanxia Rao, Heng-Li Tian
Abstract
Microglia/macrophage-induced neuroinflammation plays a crucial role in the progression of traumatic brain injury (TBI). However, the involvement of N6-methyladenosine (m 6 A) RNA modifications in this process remains elusive. Single-cell RNA sequencing (scRNA-seq) and m 6 A RNA immunoprecipitation sequencing (MeRIP-seq) across multiple time points postinjury revealed a strong correlation between m 6 A modifications and genes enriched in microglia/macrophages. Furthermore, the m 6 A demethylase ALKBH5 was identified as a key regulator of dynamic m 6 A patterns at the injury site. ALKBH5 suppression in microglia/macrophages exacerbated neuroinflammation in vitro and worsened neurological deficits in controlled cortical impact (CCI) models. MeRIP-qPCR and RNA pull-down assays revealed SOCS3 was a downstream target of ALKBH5-mediated m 6 A demethylation. This demethylation stabilized Socs3 mRNA and enhanced its protein expression, which in turn suppressed neuroinflammation via inhibiting the JAK2-STAT3 pathway. Conversely, SOCS3 depletion impaired functional recovery after injury. These findings unveiled a critical ALKBH5–m 6 A–SOCS3 regulatory axis that mitigated microglia/macrophage-driven neuroinflammation after TBI, underscoring its potential as a therapeutic intervention target for TBI progression.