PGK1 phosphorylates NLRP3 and mediates inflammasome activation independent of its glycolytic activity.

Glycolysis is a critical player in the inflammatory response. Phosphoglycerate kinase 1 (PGK1) is an essential enzyme in the glycolysis pathway. However, little is known about its role in inflammatory response. In this study, we report PGK1 as a kinase that directly regulates NLRP3 inflammasome activation in response to lipopolysaccharide (LPS) stimulation via non-glycolytic function. We identified a novel phosphorylation modification of PGK1 at S271, mediated by protein kinase CK2. Importantly, phosphorylation at S271 serves as a molecular switch that activates PGK1's kinase function, activating it to phosphorylate NLRP3. This PGK1-mediated phosphorylation at S448/S449 of NLRP3 subsequently recruits USP14 to facilitate NLRP3 deubiquitination, thereby promoting NLRP3 inflammasome activation. Using PGK1S271A/S271A transgenic mouse model, we further demonstrated that blocking S271 phosphorylation conferred significant protection against LPS-induced endotoxemia and alum-induced peritonitis. Our findings reveal a novel regulatory role of PGK1 in inflammation and provide potential therapeutic strategies for NLRP3-driven diseases.