Heart disease is invariably accompanied by the accumulation of resident cardiac fibroblasts (CF) that secrete extracellular matrix and drive the progression of fibrosis and cardiac dysfunction in heart failure. We recently reported that CF exhibit a unique transcriptional response in physiological and pathological remodeling that may underlie the cardioprotective benefits of exercise, and that p53 targets represent the most dysregulated family between the two. To further investigate the role of p53 in pathological cardiac remodeling, we deleted floxed p53 alleles in adult CF using the tamoxifen-inducible Tcf21 MerCreMer mouse (p53-CFKO). p53-CFKO mice initially retain normal cardiac function through 14 days of pressure overload, but eventually progress to heart failure more rapidly than do p53-CFWT, which show gradual functional decline throughout. CF accumulation and fibrosis at these time points match the functional results closely, with p53-CFKO mice protected from both at 14 days relative to p53-CFWT. Conversely, deletion of p53 in myofibroblasts (MF) using the tamoxifen-inducible Postn MerCreMer mouse (p53-MFKO) resulted in no delay in functional loss nor in CF accumulation and fibrosis, suggesting a distinct temporal window of p53 activity in CF exists in pressure overload. In vitro experimentation confirmed that adenovirally-mediated deletion of p53 could delay, but not reverse, myofibroblast activation - and that this irreversible activation is due to induction of Cdkn2a -mediated cell cycle arrest. Furthermore, unbiased single-cell transcriptomic analysis of CF from pressure-overloaded Tcf21 MerCreMer mice at the 14-day functional inflection point revealed p53-CFKO mice have a unique CF cluster defined primarily by a mitotic signature, and this cluster comes at the direct expense of the myofibroblast population. Clusters representing migrating and quiescent CF were unchanged in response to p53. This work describes an inverse correlation of proliferative capacity and myofibroblast activation that indicates p53 exerts master control of the timing of CF accumulation and pathological fibrosis in heart disease.