The PIDDosome controls cardiomyocyte polyploidization during postnatal heart development

The mammalian adult heart is a post-mitotic organ characterized by mainly polyploid cardiomyocytes (CMs). The post-mitotic status of polyploid adult CMs poses a clear limit to heart regeneration. Thus, understanding how CMs acquire this polyploid status is relevant to heart regeneration therapies. Here, we aim to unveil that the PIDDosome, a multi-protein complex known to limit scheduled and accidental polyploidization events by activating p53, implements a CM-specific genetic program that controls polyploidization during postnatal heart development in mice. Flow cytometric DNA content analysis coupled with image-based 3D volumetric measurements of CM nuclei showed that PIDDosome knockout animals harbor significantly more polyploid CM nuclei compared to WT controls. Remarkably, the increased CM ploidy levels do not interfere with cardiac structure or functional output and is implemented in a cell-autonomous manner. During early heart development, ploidy analyses revealed that the PIDDosome starts to shape CM ploidy at postnatal day 7 (P7), reaching a plateau of activity on postnatal day P14. In line with observations made in the hepatocytes, PIDDosome activation in CMs is dependent on PIDD1 localization to extra centrosomes in cells that fail cytokinesis via the distal appendage protein ANKRD26. Interestingly, but in contrast to prior observations in hepatocytes, the PIDDosome limits CM polyploidization in a p53-independent manner but still involves the CDK-inhibitor, p21/Cdkn1a , a notion supported by nuclear RNA sequencing and genetic analyses. Together these results provide a new mechanism how proliferation of polyploid CMs is restricted during development, adding new mechanistic insight into the tightly regulated program of terminal differentiation events occurring in the postnatal heart.