Sleeping Beauty transposon mutagenesis in mouse intestinal organoids identifies genes involved in tumor progression and metastasis [Atxn1]
To identify genes important for colorectal cancer (CRC) development and metastasis, we established a new metastatic mouse organoid model using Sleeping Beauty (SB) transposon mutagenesis. Intestinal organoids derived from mice carrying actively mobilizing SB transposons, an activating KrasG12D, and an inactivating Apc?716 allele, were transplanted to immunodeficient mice. Analysis of SB insertion sites in tumors identified numerous candidate cancer genes (CCGs) identified previously in intestinal SB screens performed in vivo, in addition to new CCGs, such as Atxn1. To provide functional validation, we knocked out Atxn1 in mouse tumor organoids carrying an activating KrasG12D, and an inactivating Apc?716 allele, and transplanted to mice. Tumor development was promoted when these gene were knocked out, demonstrating that these are potent tumor suppressors. Overall design: Atxn1 was knocked out by CRISPR-Cas9 in mouse colon tumor organoids carrying an activating KrasG12D, and an inactivating Apc?716 allele, and RNA was collected to perform RNA-seq.