METTL14 Promotes Ischemic Stroke-induced Brain Injury by Stabilizing HDAC3 Expression in an m6A-IGF2BP3 Mechanism.

Abstract

N6-methyladenosine (m6A) modification is the most abundant post-transcriptional modification of mRNAs and has been identified to play critical roles in ischemic stroke (IS). Herein, this study aimed to investigate the function and mechanism of Methyltransferase-like 14 (METTL14) methylase in cerebral IS. Murine BV-2 microglial cell OGD/R models and rat middle cerebral artery occlusion (MCAO) models were established to mimic IS-induced neuronal damage in vitro and brain injury in vivo. Levels of METTL14, Histone Deacetylase 3 (HDAC3) and cGAS-STING axis-related proteins were detected using qRT-PCR or western blotting. Cell proliferation and inflammation were assessed by CCK-8 assay, EdU assay and ELISA. Flow cytometry detected microglia polarization. Cell pyroptosis was analyzed by detecting related-protein markers by western blotting. The m6A modification was determined by methylated RNA immunoprecipitation assay. Brain injury was analyzed by evaluating infarct volume and neurologic score. METTL14 levels were higher in OGD/R-induced microglial cells, primary microglia and infarct brain tissues of rat MCAO models. Functionally, METTL14 silencing reversed OGD/R-induced proliferation inhibition, inflammation and pyroptosis in microglial cells and primary microglia in vitro, and ameliorated cerebral ischemic injury in rat MCAO models. Mechanistically, METTL14 induced HDAC3 m6A modification in an IGF2BP3-dependent manner, and could activate cGAS-STING pathway through HDAC3. Moreover, HDAC3 overexpression reversed the neuroprotective effects of METTL14 silencing. METTL14 silencing reversed ischemic stroke-induced brain injury by inducing HDAC3 m6A modification in an IGF2BP3-dependent mechanism, recommending a novel insight for ameliorating cerebral ischemic stroke.