High mTORC1 signaling is maintained, while protein degradation pathways are perturbed in old murine skeletal muscles in the fasted state

Abstract

This study investigated age-associated changes to protein synthesis and degradation pathways in the quadriceps muscles of male C57BL/6J mice at 5 ages, between 4 and 24 months (m). Sarcopenia was evident by 18m and was accompanied by hyper-phosphorylation of S6K1, indicating increased mTORC1 signaling. Proteasomal and autophagosomal degradation pathways were also impacted by aging. In the 1% NP40 insoluble protein fraction, the abundance of MuRF1 increased at 24m, while p62 increased at 15m, and remained elevated at older ages. In addition, we investigated how protein synthesis and degradation pathways are modulated by fasting in young (4m) and old (24m) muscles, and showed that old mice respond to fasting less robustly compared with young. Overnight fasting for 16h caused de-phosphorylation of AKT and molecules downstream of mTORC1 (S6K1, rpS6 and 4E-BP1) in young, but not old muscles. A longer time of fasting (24h) was required to reduce phosphorylation of these molecules in old mice. Induction of MuRF1 and Fbxo32 mRNA was also more robust in young compared with old muscles following fasting for 16h. In addition, a 16h fast reduced ULK1 phosphorylation at the mTORC1 specific site Ser757 only in young muscles. The striking accumulation of insoluble p62 protein in muscles of all old male mice (fed or fasted), suggests age-related dysregulation of autophagy and protein aggregation. These data provide an insight into the mechanisms of metabolic responses that affect protein homeostasis in old skeletal muscles, with applications to design of clinical interventions that target sarcopenia.

Keywords: Aging; Autophagy; Muscle; Protein degradation; mTORC1; p62.