ENO3 encodes enolase 3 (β-enolase), a muscle-specific glycolytic enzyme that catalyzes the conversion of 2-phosphoglycerate to phosphoenolpyruvate in glycolysis and the reverse reaction in gluconeogenesis 1. The ENO3 gene spans approximately 6 kb, contains 12 exons, and encodes a 433-residue protein with regulatory elements including muscle-specific transcription factor binding sites 1. ENO3 expression is regulated by tissue-specific methylation patterns, with unmethylated CpG islands in the promoter region across all tissues but differential methylation within the gene body 2. Beyond its metabolic role, ENO3 appears critical for striated muscle development and regeneration, with expression changes observed during satellite cell activation 3. ENO3 dysfunction contributes to various pathological conditions: it promotes aerobic glycolysis in cholangiocarcinoma through USP21-mediated stabilization 4, enhances colorectal cancer progression by increasing cellular glycolysis, ATP production, and lactate levels 5, and participates in diabetic angiopathy through SIRT6-mediated deacetylation that drives pathological glycolysis reprogramming in vascular smooth muscle cells 6. These findings establish ENO3 as both a fundamental metabolic enzyme and a potential therapeutic target in cancer and diabetic complications.