Background: Although cardiac c-kit ⁺ cells are being tested in clinical trials, the circumstances that determine lineage differentiation of c-kit ⁺ cells in vivo are unknown. Recent findings suggest that endogenous cardiac c-kit ⁺ cells rarely contribute cardiomyocytes to the adult heart. We assessed whether various pathological stimuli differentially affect the eventual cell fates of c-kit ⁺ cells. Methods: We used single-cell sequencing and genetic lineage tracing of c-kit ⁺ cells to determine whether various pathological stimuli would result in different fates of c-kit ⁺ cells. Results: Single-cell sequencing of cardiac CD45 ^- c-kit ⁺ cells showed innate heterogeneity, indicative of the existence of vascular and mesenchymal c-kit ⁺ cells in normal hearts. Cardiac pressure overload resulted in a modest increase in c-kit-derived cardiomyocytes, with significant increases in the numbers of endothelial cells and fibroblasts. Doxorubicin-induced acute cardiotoxicity did not increase c-kit-derived endothelial cell fates but instead induced cardiomyocyte differentiation. Mechanistically, doxorubicin-induced DNA damage in c-kit ⁺ cells resulted in expression of p53. Inhibition of p53 blocked cardiomyocyte differentiation in response to doxorubicin, whereas stabilization of p53 was sufficient to increase c-kit-derived cardiomyocyte differentiation. Conclusions: These results demonstrate that different pathological stimuli induce different cell fates of c-kit ⁺ cells in vivo. Although the overall rate of cardiomyocyte formation from c-kit ⁺ cells is still below clinically relevant levels, we show that p53 is central to the ability of c-kit ⁺ cells to adopt cardiomyocyte fates, which could lead to the development of strategies to preferentially generate cardiomyocytes from c-kit ⁺ cells.