Our objective was to determine the impact of carbohydrate and/or protein ingestion before and after exercise on ribosomal protein S6 kinase (S6K1) and S6 phosphorylation status in human skeletal muscle tissue. Seven healthy, untrained men (22.5 +/- 0.9 y) were randomly assigned to 2 cross-over experiments. Before, immediately after, and 1 h after a single bout of resistance exercise, subjects consumed 0.3 g.kg(-1) carbohydrate with or without 0.3 g.kg(-1) protein hydrolysate (CHO+PRO and CHO, respectively). Muscle biopsies were taken before and immediately after exercise and after 1 and 4 h of postexercise recovery to determine 4E-BP1, S6K1 (both T(421)/S(424) and T(389)), and S6 phosphorylation status. Following resistance exercise, 4E-BP1 phosphorylation was reduced to a greater extent in the CHO treatment (-48 +/- 7%) than in the CHO+PRO treatment (-15 +/- 14%, P < 0.01). During recovery, 4E-BP1 phosphorylation increased in both experiments (P < 0.01), and tended to be higher in the CHO+PRO test (P = 0.08). S6K1 phosphorylation at T(421)/S(424) substantially increased following exercise and remained elevated during recovery with no differences between treatments. In contrast to the CHO treatment (-4 +/- 2%), S6K1 phosphorylation at T(389) was higher following exercise in the CHO+PRO treatment only (+78 +/- 2%, P < 0.01). During recovery, S6K1 phosphorylation at T(389) remained higher in CHO+PRO than in CHO (P < 0.05). S6 phosphorylation was substantially higher following exercise in the CHO+PRO (1.69 +/- 0.35) than in the CHO experiment (0.45 +/- 0.07, P < 0.01) and remained elevated during recovery (P < 0.05). We conclude that the availability of dietary protein further enhances phosphorylation of S6K1 during recovery from resistance type exercise. AD - Departments of Movement Sciences and 3Human Biology, Nutrition and Toxicology Research Institute Maastricht (NUTRIM), Maastricht University, Maastricht 6200 MD, The Netherlands.