Abstract: Eicosapentaenoic acid (EPA) is known to influence skeletal muscle protein turnover in mammals, but its effect on teleost and the underlying mechanisms remain unclear. In this study, the impact of EPA on cellular protein accumulation and metabolism of lipid and glucose were examined in turbot muscle cells. The nascent protein synthesis and the activities of the protein kinase B (Akt)/the mechanistic target of rapamycin (mTOR) signaling pathway were enhanced by EPA. Furthermore, EPA repressed the expression of Atrogin-1 and Muscle Ring Finger protein-1 (MuRF-1), two dominant muscle-specific ubiquitin ligases. EPA also shifted cellular metabolism, as reflected by changes in the expression of key enzymes involved in lipid metabolism and glycolysis. These results suggested that EPA might exert beneficial effects on protein turnover by stimulating protein synthesis, repressing ubiquitin-proteasome pathway-mediated protein degradation, as well as reprogramming cellular metabolism. Our findings support that the function of EPA on promoting protein accumulation is conserved in teleost.