Novel Insights into the Regulation of Translation in Skeletal Muscle as Revealed by a New Non‐Radioactive Method for In‐Vivo Measurements of Protein Synthesis

The principles of SUface Sensing of Translation (SUnSET) were used to develop a non‐radioactive method for ex‐vivo and in‐vivo measurements of protein synthesis (PS). Compared with controls, we first demonstrate excellent agreement between SUnSET and a 3H‐ phenylalanine method when detecting synergist ablation (SA)‐induced increases in skeletal muscle PS ex‐vivo. We then show that SUnSET can detect the same SA‐induced increase in PS when employed in‐vivo ( IV ‐SUnSET). Additionally, IV ‐SUnSET detected food deprivation‐ induced decreases in PS in the heart, kidney and skeletal muscles, with similar changes being visualized with an immunohistochemical version of IV ‐SUnSET ( IV HC‐SUnSET). By combining IV HC‐SUnSET with in‐vivo transfection, we demonstrate that constitutively active PKB induces a robust increase in skeletal muscle PS. Furthermore, transfection with Rheb revealed that a PKB‐independent activation of mTOR is also sufficient to induce an increase in skeletal muscle PS. Finally, IV HC‐SUnSET exposed the existence of fiber type‐dependent differences in skeletal muscle PS, with PS in type 2B and 2X fibers being significantly lower than type 2A fibers within the same muscle. Thus, our non‐radioactive method allowed us to accurately visualize and quantify PS under various ex‐vivo and in‐vivo conditions and revealed novel insights into the regulation of PS in skeletal muscle.