Mis-splicing Drives Loss of Function of p53(E224D) Point Mutation

BACKGROUND: Tp53 is the most commonly mutated gene in cancer. Canonical Tp53 DNA damage response pathways are well characterized and classically thought to underlie the tumor suppressive effect of Tp53. Challenging this dogma, mouse models have revealed that p53 driven apoptosis and cell cycle arrest are dispensable for tumor suppression. Here, we investigated the inverse context of a p53 mutation predicted to drive expression of canonical targets, but is detected in human cancer. METHODS: We established a novel mouse model with a single base pair mutation (GAG>GAC, p53(E221D)) in the DNA-Binding domain that has wild-type function in screening assays, but is paradoxically found in human cancer in Li-Fraumeni syndrome. Using mouse p53(E221D) and the analogous human p53(E224D) mutant, we evaluated expression, transcriptional activation, and tumor suppression in vitro and in vivo. RESULTS: Expression of human p53(E224D) from cDNA translated to a fully functional p53 protein. However, p53(E221D/E221D) RNA transcribed from the endogenous locus is mis-spliced resulting in nonsense mediated decay. Moreover, fibroblasts derived from p53(E221D/E221D) mice do not express a detectable protein product. Mice homozygous for p53(E221D) exhibited increased tumor penetrance and decreased life expectancy compared to p53 WT animals. CONCLUSIONS: Mouse p53(E221D) and human p53(E224D) mutations lead to splice variation and a biologically relevant p53 loss of function in vitro and in vivo.