What is the central question of this study? The adaptations to protein synthesis and degradation that occur in skeletal muscle in response to HIIT are not well understood. The magnitudes of the changes in response to HIIT, compared to MICT, as well as the mechanisms underlying these changes, are also unclear. What is the main finding and its importance? HIIT is more effective than MICT in altering the expression of MuRF-1and MAFbx, and enhancing the autophagic flux in rat soleus muscle. In addition, HIIT could activate the mTOR pathway. These findings suggest that HIIT might be an effective exercise strategy for health promotion in skeletal muscle.This study aimed to investigate the impact of high-intensity interval training (HIIT) on the proteins involved in protein synthesis, the ubiquitin-proteasome system (UPS) and autophagy in skeletal muscle of middle-aged rats. Nine-month old male Wistar rats (N = 56) were randomly divided into three groups: a control group (C group), a moderate-intensity continuous training (MICT) group, and a HIIT group. Rats in the training groups ran on treadmills 5 days/week for 8 weeks. The MICT group ran for 50 min at 60% VO max, while the HIIT group ran for 3 minutes at 80% of VO max six times separated by 3-minute periods at 40% VO max. Aerobic endurance, number of autophagosome and expression of proteins involved in protein synthesis and degradation in the soleus muscle were measured at three time points: before training, after 4 weeks and after 8 weeks of training. Compared to the C group, HIIT and MICT increased the expression of phosphorylated mTOR after 8 weeks (P < 0.05, P < 0.01). HIIT increased the expression of MuRF-1 after 4 weeks (P < 0.01), and decreased its expression after 8 weeks (P < 0.01). Both HIIT and MICT decreased the expression of MAFbx after 4 weeks (P < 0.05). HIIT improved the expression of LC3II (P < 0.05), and decreased the P62 content (P < 0.01) after 4 weeks. The LC3II/LC3I ratio was increased after 8 weeks (P < 0.01). This study demonstrated that HIIT could activate the mTOR pathway, alter the expression of MuRF1 and MAFbx proteins, and enhance the autophagic flux in soleus muscle of middle-aged rats. This article is protected by copyright. All rights reserved.