Imaging of existing and newly translated proteins elucidates mechanisms of sarcomere turnover

Abstract

How the sarcomeric complex is continuously turned-over in long-living cardiomyocytes is unclear. Here, we imaged the exchange of old and new Halo-tagged sarcomeric proteins. We disprove the prevailing ‘protein pool’ model and instead show an ordered mechanism in which only newly translated proteins enter the sarcomeric complex while older ones are removed and degraded. We also show that degradation is independent of protein age, and that proteolytic extraction is a rate limiting step in the turnover. We imaged and quantified the turnover of expressed and endogenous sarcomeric proteins, including the giant protein titin, in cardiomyocytes in culture and in vivo, at the single cell and at the single sarcomere level. We show that replacement occurs at a similar rate within cells and across the heart and is markedly slower in adult cells. Our findings imply a new model of sarcomere turnover and sarcomeric subunit replacement.