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We determined the effect of protein supplementation on anabolic signaling and rates of myofibrillar and mitochondrial protein synthesis after a single bout of concurrent training.Using a randomized crossover design, eight healthy males were assigned to experimental trials consisting of resistance exercise (8 × 5 leg extension, 80% 1RM) followed by cycling (30 min at approximately 70% V˙O2peak) with either postexercise protein (PRO, 25-g whey protein) or placebo (PLA) ingestion. Muscle biopsies were obtained at rest and at 1 and 4 h after exercise.AktSer473 and mTORSer2448 phosphorylation increased 1 h after exercise with PRO (175%–400%, P < 0.01) and was different from PLA (150%–300%, P < 0.001). Muscle RING finger 1 and atrogin-1 messenger RNA (mRNA) were elevated after exercise but were higher with PLA compared with those in PRO at 1 h (50%–315%, P < 0.05), whereas peroxisome proliferator-activated receptor gamma coactivator 1-alpha mRNA increased 4 h after exercise (620%–730%, P < 0.001), with no difference between treatments. Postexercise rates of myofibrillar protein synthesis increased above rest in both trials (75%–145%, P < 0.05) but were higher with PRO (67%, P < 0.05), whereas mitochondrial protein synthesis did not change from baseline.Our results show that a concurrent training session promotes anabolic adaptive responses and increases metabolic/oxidative mRNA expression in the skeletal muscle. PRO ingestion after combined resistance and endurance exercise enhances myofibrillar protein synthesis and attenuates markers of muscle catabolism and thus is likely an important nutritional strategy to enhance adaptation responses with concurrent training.