Sigma-1 receptor counteracts non-cell-autonomous poly-PR-induced astrocytic oxidative stress in C9orf72 ALS.

C9orf72-associated amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are characterized by the accumulation of toxic dipeptide repeat proteins (DPRs) generated from G4C2 hexanucleotide repeat expansions. Among these, the arginine-rich poly-PR (proline-arginine) species is the most neurotoxic, eliciting glial activation and neuroinflammation via non-cell-autonomous mechanisms. Although growing evidence implicates glial cells, particularly astrocytes, in disease progression, the molecular pathways linking neuron-derived poly-PR to astrocyte-mediated oxidative stress remain poorly understood. We demonstrate that exogenous poly-PR induces robust NOX4 expression and hydrogen peroxide (H2O2) production in astrocytes through activation of the IKK/IκB/NF-κB p65 signaling pathway. Mechanistically, poly-PR promotes nuclear translocation of p65 and enhances its binding to the NOX4 promoter, thereby amplifying astrocytic oxidative stress. Overexpression of the Sigma-1 receptor (Sigma-1R), an endoplasmic reticulum-resident chaperone, significantly attenuates poly-PR-induced NOX4 transcription and reactive oxygen species (ROS) production by interacting with p65 and blocking its nuclear translocation, independently of upstream p65 phosphorylation. Notably, clemastine, a clinically approved Sigma-1R agonist, suppresses astrocytic NOX4 expression by disrupting p65 binding to the NOX4 promoter. In a mouse model of C9orf72 ALS, Sigma-1R deficiency exacerbates poly-PR-induced neurodegeneration, astrogliosis, and NOX4 upregulation, whereas Sigma-1R sufficiency confers neuroprotection and anti-inflammatory effects. This study identifies Sigma-1R as a critical modulator of non-cell-autonomous poly-PR toxicity and establishes its activation as a potent suppressor of astrocyte-derived oxidative stress. Our findings uncover a previously unrecognized glial mechanism driving C9orf72 ALS pathogenesis and support Sigma-1R activation, via clemastine, as a promising therapeutic strategy to mitigate neuroinflammation and disease progression.