PIM1-induced Drp1 phosphorylation disrupts microglial mitophagy and aggravates neuroinflammation.

Current disease-modifying therapies for multiple sclerosis (MS) primarily target peripheral immune responses but exhibit limited efficacy in mitigating the compartmentalized neuroinflammation driven by central nervous system (CNS)-resident microglia. By integrating clinical sample analysis with experimental autoimmune encephalomyelitis (EAE) model studies, we have demonstrated that the proviral integration site for Moloney murine leukemia virus 1 (PIM1) is significantly upregulated, particularly in microglia, in both MS patients and the spinal cords of EAE mice. This upregulation positively correlates with disease severity and levels of proinflammatory cytokines such as IL-1β, TNF-α, and IL-6. Utilizing a multimodal research approach-including pharmacological inhibition (SMI-4a), genetic knockdown, RNA sequencing, and HIS-SIM super-resolution imaging-we confirmed that PIM1 inhibition effectively attenuates neuroinflammatory responses, improves clinical symptoms in EAE mice, and promotes activation of the mitophagy pathway while suppressing inflammation-related molecules. Mechanistically, PIM1 enhances the phosphorylation of dynamin-related protein 1 (Drp1) at Ser616 while suppressing its phosphorylation at Ser637, which disrupts LC3-mitochondria colocalization and autophagosome-lysosome fusion. This leads to mitophagy dysfunction, loss of mitochondrial membrane potential, and accumulation of reactive oxygen species. Notably, the combined administration of PIM1 and Drp1 inhibitors did not yield synergistic therapeutic effects, suggesting that PIM1 likely functions as an upstream master regulator of Drp1. These findings not only elucidate the molecular mechanism by which PIM1 interacts with Drp1 to regulate microglial activation and mitophagy but also establish PIM1 as a promising CNS-intrinsic therapeutic target for restoring mitochondrial homeostasis in MS.