The tumor suppressor p53 is a transcription factor that coordinates the cellular response to DNA damage. Here we provide an integrated analysis of p53 genomic occupancy and p53-dependent gene regulation in the splenic B and non-B cell compartments of mice exposed to whole-body ionizing radiation, providing insight into general principles of p53 activity in vivo. In unstressed conditions, p53 bound few genomic targets; induction of p53 by ionizing radiation increased the number of p53 bound sites, leading to highly overlapping profiles in the different cell types. Comparison of these profiles with chromatin features in unstressed B cells revealed that, upon activation, p53 localized at active promoters, distal enhancers, and a smaller set of unmarked distal regions. At promoters, recognition of the canonical p53 motif as well as binding strength were associated with p53-dependent transcriptional activation, but not repression, indicating that the latter was most likely indirect. p53-activated targets constituted the core of a cell type-independent response, superimposed onto a cell type-specific program. Core response genes included most of the known p53-regulated genes, as well as many new ones. Our data represent a unique characterization of the p53-regulated response to ionizing radiation in vivo.