Broiler chickens are characterized by fast muscle growth and high protein deposition, most likely subsequent to a high protein synthesis. However, the regulation of protein synthesis in chicken muscle is still unknown. In contrast, it has been clearly demonstrated in mammals that S6K1 is a key regulator of protein synthesis. In the present study, S6K1 was characterized in both pectoralis and gastrocnemius muscles in chickens. A 133-bp fragment of chicken S6K1 cDNA had 84% identity to mammalian S6K1. We investigated in vivo the effects of refeeding and insulin treatment after 16 h starvation. S6K1 enzyme activity was significantly increased in both pectoralis and gastrocnemius muscles by refeeding (two- to threefold greater than in food-deprived chickens, P < 0.05). Optimal activation occurred 30 min after refeeding following 16 h starvation. S6K1 activation was associated with its phosphorylation on serine and Thr 389 residues, which occurred within the first 5 min of refeeding. S6K1 was also significantly stimulated in both pectoralis and gastrocnemius muscles after a single insulin injection (nine- to 12-fold greater than in control chickens, P < 0.001). Our results indicate that S6K1 is expressed in chickens muscles and activated by refeeding and insulin treatment.