Following birth, mammalian cardiomyocytes become polyploid by replicating their DNA without completing cell division. Different ploidy classes can arise depending on whether cells undergo nuclear division, and this variation may affect regeneration; for example, a higher proportion of mononuclear diploid cardiomyocytes (MNDCMs) is associated with greater regenerative competence. Michaela Patterson and colleagues had previously shown that prevalence of various ploidy classes varies between different inbred mouse strains. Here, they focus on A/J mice and show that the cardiomyocyte ploidy is more dynamic than studies of other strains had suggested. Specifically, they show the proportion of MNDCMs in the A/J cardiomyocyte population increases during postnatal weeks 3-6 as a result of delayed completion of cardiomyocyte cell division. A/J mice carry a mutation in Tnni3k, a gene that has been previously linked to ploidy regulation, but knocking out Tnni3k in wild-type mice only partially reproduces the A/J phenotype. Genome-wide association studies identify Runx1 as an additional candidate for ploidy regulation, and they show that overexpressing Runx1 is sufficient to induce A/J phenotypes. Runx1 overexpression can also stimulate cardiomyocyte proliferation following myocardial infarction. Taken together, this work identifies a role for Runx1 in the regulation of cardiomyocyte division and polyploidy.