DRAM1 activates microglial pyroptosis and neuroinflammation through TMEM9-mediated lysosomal membrane permeabilization after spinal cord injury.

Abstract

Spinal cord injury (SCI) is a devastating condition characterized by severe neuroinflammation and microglial pyroptosis, yet the underlying mechanisms remain incompletely understood. In this study, the role of DRAM1, a lysosomal membrane protein, in SCI pathogenesis was investigated. Single-cell RNA sequencing, an in vivo SCI mouse model, and in vitro LPS-stimulated BV2 microglial cells were employed, combined with histopathological assessments, locomotor function tests, and molecular interaction assays. Our results demonstrated that DRAM1 expression was significantly upregulated in microglia after SCI. Mechanistically, DRAM1 interacted with TMEM9 to regulate the expression of the V-ATPase accessory subunit ATP6AP2, thereby inducing lysosomal membrane permeabilization (LMP) and impairing autophagic flux, and facilitating microglial pyroptosis-mediated neuroinflammation after SCI. Moreover, IGF2BP3 functioned as an m6A reader that recognized and stabilized m6A-modified DRAM1 mRNA, thus enhancing DRAM1 expression in activated microglia. Knockdown of DRAM1 or IGF2BP3 attenuated microglial pyroptosis and inflammatory responses in vitro. Consistently, in vivo experiments showed that AAV-mediated DRAM1 knockdown improved locomotor recovery, reduced spinal cord lesion area, and suppressed neuroinflammatory responses in SCI mice. Collectively, these findings reveal that DRAM1 promotes microglial pyroptosis and neuroinflammation after SCI via the TMEM9-dependent induction of LMP and autophagic flux impairment, with its expression tightly regulated by IGF2BP3 in an m6A-dependent manner. Taken together, our study identifies DRAM1 as a novel therapeutic target for SCI.