Wild type and tumor-derived mutant p53 share a non-canonical mode of gene occupancy [RNA-seq]

To directly examine the interplay between mutant p53 or Mdm2 and wild type p53 in gene occupancy and expression, an integrated RNA-seq and ChIP-seq analysis was performed in vivo using isogenically matched mouse strains. Response to radiation was used as an endpoint to place findings in a biologically relevant context. Unexpectedly, mutant p53 and Mdm2 only inhibit a subset of wild type p53 gene expression. In contrast to a dominant-negative or inhibitory role, the presence of either mutant p53 or Mdm2 actually enhances the occupancy of wild type p53 on many canonical targets. The C-terminal 19 amino acids of wild type p53 suppresses the p53 response allowing for survival at sublethal doses of radiation. Further, the mutant 172H p53 is shown to occupy genes and regulate their expression via a non-canonical means that is shared with wild type p53. This results in the heterozygous 172H/+ genotype having an expanded transcriptome compared to wild type p53+/+. Expression profiling by high-throughput sequencing of bone marrow in wt p53 mice and mice with a deletion of the C-terminus of p53. Bone marrow from the indicated genotype mice was treated as indicated (9.5 or 6 Gy) from 8-week-old mice (8w). Tissue were immediately placed in RNAlater during dissection and stored at 4oC until RNA is extracted using RNeasy Mini Kit (Qiagen) and following manufacturer’s instructions. The isolated RNA is quantitated and either stored at -80o in preparation for RNA-seq. Sequencing libraries prepared with the TruSeq Stranded Total RNA kit (Illumina Inc), from 1ug total RNA.