AbstractMdm2, an E3 ubiquitin ligase, negatively regulates the tumour suppressor p53. Loss of Mdm2 in mice results in p53‐dependent apoptosis and embryonic lethality. This phenotype was rescued by the p53515C allele, which encodes an apoptosis‐deficient p53R172P protein. However, these mice died within 2 weeks of birth, due to a severe impairment of progenitor cell expansion during postnatal haematopoiesis and cerebellar development, leading to p53‐dependent cell cycle arrest. Loss of Mdm2 led to phosphorylation of the p53R172P protein, p53R172P stability and activation of the cell cycle inhibitor p21 in proliferating cells, but not in differentiated cells, in multiple tissue compartments. Proliferating cells of epithelial origin were not affected. The haematopoietic and neural defects were alleviated in mice lacking Mdm2 and containing one p53515C and one p53‐null allele, but spermatogenesis was arrested. These findings establish a crucial role for the p53–Mdm2 network in regulating proliferation and progenitor expansion in many cell lineages and have important implications for the use of drugs that aim to disrupt the p53–Mdm2 interaction. Copyright © 2007 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.