Regulation of Autophagy in Chick Skeletal Muscle: Effect of mTOR Inhibition.

Abstract

Autophagy in the skeletal muscle increases under catabolic conditions resulting in muscle atrophy. This study investigated the effect of inhibition of mechanistic target of rapamycin (mTOR) on autophagy in chick skeletal muscle. We examined the effects of Torin1, an mTOR inhibitor, on autophagy. Chick myotubes were incubated with Torin1 (100 nM) for 3 h. It was observed that Torin1 inhibited the phosphorylation of AKT (Ser473), p70 ribosomal S6 kinase 1 (S6K1, Thr389), S6 ribosomal protein (Ser235/236), and eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1, Thr37/46), which are used for measurement of mTOR activity. Torin1 significantly (P< 0.01) increased the LC3-II/LC3-I ratio, an index for autophagosome formation, while it did not influence the expression of autophagy-related genes (LC3B, GABARAPL1, and ATG12). In addition, Torin1 increased atrogin-1/MAFbx (a muscle-specific ubiquitin ligase) mRNA expression. Fasting for 24 h inhibited the phosphorylation of AKT (Ser473), S6K1 (Ther389), S6 ribosomal protein (Ser235/236), and 4E-BP1 (Thr37/46) in chick skeletal muscle and significantly (P<0.01) increased the LC3-II/LC3-I ratio. Fasting also increased GABARAPL1 and atrogin-1/MAFbx mRNA expression but not LC3B or ATG12 mRNA expression. These results indicate that mTOR signaling regulates autophagy and the ubiquitin-proteasome proteolytic pathway in chick skeletal muscle.