ENO1 from Mycoplasma bovis Disrupts Host Glycolysis and Inflammation by Binding ACTB.

Mycoplasma bovis is an important pathogen that is associated with respiratory diseases, mastitis, and arthritis in cattle, leading to significant economic losses in the global cattle industry. Most notably in this study, we pioneer the discovery that its secreted effector ENO1 (α-enolase) directly targets host cytoskeletal proteins for metabolic-immune regulation. Using an innovative GST pull-down/mass spectrometry approach, we made the seminal discovery of β-actin (ACTB) as the primary host target of ENO1-the first reported bacterial effector-cytoskeleton interaction mediating metabolic reprogramming. ENO1-ACTB binding depends on a hydrogen bond network involving ACTB's 117Glu and 372Arg residues. This interaction triggers (1) glycolytic activation via Glut1 upregulation, establishing Warburg effect characteristics (lactic acid accumulation/ATP inhibition), and (2) ROS-mediated activation of dual inflammatory axes (HIF-1α/IL-1β and IL-6/TNF-α). This work establishes three groundbreaking concepts: (1) the first evidence of a pathogen effector hijacking host ACTB for metabolic manipulation, (2) a novel 'glycolysis-ACTB-ROS-inflammation' axis, and (3) the first demonstration of bacterial proteins coordinating a Warburg effect with cytokine storms. These findings provide new targets for anti-infection therapies against Mycoplasma bovis.