GSE132551
Analysis of the transcriptomic changes in murine small and large intestinal organoids after the sudden expression of BRAFV600E and/or p53R172H
BRAFV600E confers poor prognosis and a distinct molecular type of colorectal cancer, which is often associated with TP53 alterations. In order to understand how BRAFV600E and p53R172H promote tumorigenesis in the intestinal epithelium, we generated murine organoids harboring conditional BraffloxV600E and/or Trp53LSL-R172H alleles and conducted RNA sequencing analysis after sudden oncogene induction. As expected from their phenotypes, organoids expressing BRAFV600E, either singly or in combination with p53R172H, displayed strong changes in their transcriptome, while induction of mutant p53 alone had only little impact on gene expression. Our transcriptome analyses revealed there are profound differences in the transcriptomes of SI and COL organoids, in their oncogene naïve ground state and upon BRAFV600E expression. We observed many transcripts positively associated with proliferation, invasion and metastasis to be upregulated in both organoid types expressing BRAFV600E or BRAFV600E together with p53R172H. However, several of them were only differentially regulated in COL organoids, or displayed a more pronounced fold change in COL than in SI organoids. Interestingly, COL organoids more closely recapitulate human CRC patterns and p53R172H cooperates with mutant BRAFV600E in the regulation of several transcripts. Overall design: Small intestinal (SI) and colonic (COL) organoids of three donor mice per tissue (Trp53LSL-R172H/+;Villin::CreERT2, BraffloxV600E/+;Villin::CreERT2, BraffloxV600E/+;Trp53LSL-R172H/+;Villin::CreERT2) were cultured. Cre recombination was induced with 4-Hydroxytamoxifen (0.5 µM for SI organoids, 3.5 µM for COL organoids) or organoids were treated with ethanol as controls in duplicates in-vitro for 24 hours. Isolated mRNA (for SI organoids 8 days after Cre recombination, for COL organoids 3 days after Cre recombination) was sequenced on an Illumina HiSeq4000.