Sarcomere Function Inhibits Cardiomyocyte Replication and Promotes Polyploidization Through a DNA Damage Response that Limits <i>In vivo</i> Cell Engraftment

Abstract

Human cardiac regeneration is limited by low cardiomyocyte replicative rates and progressive polyploidization by unclear mechanisms. To study this process, we engineered a human cardiomyocyte model to track replication and polyploidization using fluorescently tagged cyclin B1 and cardiac troponin T. Using time-lapse imaging, in vitro cardiomyocyte replication patterns recapitulated the progressive mononuclear polyploidization and replicative arrest observed in vivo. Single-cell transcriptomics and epigenetic analyses revealed that polyploidization was preceded by sarcomere formation, enhanced oxidative metabolism, a DNA damage response, and p53 activation. CRISPR knockout screening revealed p53 as a driver of cell cycle arrest and polyploidization. Inhibiting contractile function with troponin knockout, or scavenging ROS, inhibited cell cycle arrest and polypoloidization. Finally, we showed that cardiomyocyte engraftment in infarcted rat hearts could be enhanced 4-fold by the increased proliferation of troponin-knockout cardiomyocytes. Thus, sarcomere assembly inhibits cell division through a DNA damage response that can be targeted to improve cardiomyocyte replacement strategies.