Abstract : Breast cancer is the most frequent cancer among women in the United States. A full term pregnancy early in reproductive life can reduce breast cancer incidence in women by up to 50% and p53, an important tumor suppressor gene, was shown to be a major effector for this protection effect. We hypothesized that p53 may negatively regulate the proliferation and self-renewal of mammary stem/progenitor cells and that the increased p53 in parous gland may limit mammary stem/progenitor cells population and reduce the transformation risk. Mammosphere assays showed that decreased p53 dosage in Trp53+/- and Trp53-/- mice led to increased numbers and size of mammospheres. Meanwhile, the number of secondary and tertiary mammospheres was not affected by (IR) regardless of their genotype. The cell cycle analysis also showed that the Trp53-/- mammospheres have higher proliferation rate than Trp53+/+ spheres. Serial dilution and transplantation experiments also showed that the Trp53-/- epithelium had significantly increased frequency of long-term regenerative MaSCs compared to Trp53+/+ epithelium. The BrdU labeling experiment showed that Trp53-/- mammary gland contains less label-retaining epithelial cells (LRECs) than Trp53+/+ epithelium. Our data suggest that p53 negatively regulates the self-renewal of mammary stem cells. The MaSCs pool expands with decreased p53 dosage, which may result in a higher transformation risk. Meanwhile, the p53-mediated apoptosis pathway is compromised in the mammosphere-initiating cells.