Abstract The p53 tumor suppressor is a DNA damage/stress response protein that functions as a transcription factor to regulate a large repertoire of genes that prevent proliferation of damaged cells via initiation of cell cycle arrest and senescent programs or via apoptosis and other mechanisms of cell death. However, the molecular, morphological, and physiological consequences of this transcriptional program remain poorly understood in vivo. We have defined the physiological p53 transcriptional program using the mouse as a model system. Global p53 activation causes anomalies in the pancreas (an acinar to ductal metaplasia), intestine (crypt atrophy) and kidney (dilated tubules), and directly activates hundreds of genes many in a tissue-specific manner. Seven genes, however, are universally expressed in the five tissues (pancreas, intestine, kidney, ovary, heart) examined and represent a pan-tissue p53 transcriptional signature which includes Mdm2 (which encodes a potent p53 inhibitor) but not p21 (a cell cycle inhibitor and first identified p53 target gene). The pan-tissue p53 transcriptional signature includes three other genes with central functions in cell survival or cell death for which little is known. Eda2r is a transmembrane receptor and member of the TNF family with the ability to induce apoptosis. Significantly reduced expression of EDA2R is observed in mutant p53 breast, colorectal, and lung cancers and in pancreatic ductal adenocarcinoma and glioblastoma. Gtse1 encodes a cytoplasmic protein that migrates to the nucleus upon DNA damage, and binds and inhibits p53 activity in cells in culture. A review of cancer sequencing data indicates that amplification of GTSE1 is mutually exclusive with TP53 alterations in angiosarcomas further implicating it as an inhibitor of p53 function. Polκ is an error prone DNA polymerase that allows replication of damaged DNA and ensures cell survival, but contributes to an increased mutation burden. We hypothesize that these three transcriptional targets are key effectors of the p53 pathway and functionally contribute to ensure cell survival (via cell cycle arrest or senescence) or initiate cell death. In addition, two of these effectors have opposing properties: Eda2r induces apoptosis, while Gtse1 inhibits p53 activity and ensures cell survival. Single cell analysis in the pancreas upon p53 activation, either by deletion of Mdm2 or inflammation, will be performed to understand the cell-specific and dynamic nature of the p53 transcriptional program. The specific aims are: 1) to determine the roles of Eda2r in p53-dependent processes in vivo; 2) To determine the importance of Gtse1 in inhibiting p53 activity in vivo; 3) To determine the functional significance of p53 regulation of Polκ in homeostasis, inflammation and cancer; and 4) To determine the specificity of the p53 response using single cell analyses of the pancreas. A deep fundamental knowledge of the physiological p53 transcriptional program and ...