Transposon mutagenesis identifies genetic drivers of BrafV600E melanoma

Abstract

Although nearly half of human melanomas harbor oncogenic BRAFV600E mutations, the genetic events that cooperate with these mutations to drive melanogenesis are still largely unknown. Here we show that Sleeping Beauty (SB) transposon-mediated mutagenesis drives melanoma progression in BrafV600E mutant mice and identify 1,232 recurrently mutated candidate cancer genes (CCGs) from 70 SB-driven melanomas. CCGs are enriched in Wnt, PI3K, MAPK and netrin signaling pathway components and are more highly connected to one another than predicted by chance, indicating that SB targets cooperative genetic networks in melanoma. Human orthologs of >500 CCGs are enriched for mutations in human melanoma or showed statistically significant clinical associations between RNA abundance and survival of patients with metastatic melanoma. We also functionally validate CEP350 as a new tumor-suppressor gene in human melanoma. SB mutagenesis has thus helped to catalog the cooperative molecular mechanisms driving BRAFV600E melanoma and discover new genes with potential clinical importance in human melanoma.

Transposon mutagenesis identifies genes that cooperate with mutant Pten in breast cancer progression

“…In addition, in many cases, only a single transposon insertion was present at a CCG in tumor cells, suggesting that many CCGs are functioning as haploinsufficient TSGs. This is similar to what has been observed in other SB mutagenesis screens performed in solid tumors (18)(19)(20)(21)(22)(23)(24)(25)(26).…”

“…Identification of Trunk Driver Genes. Most CCGs identified by SB mutagenesis are thought to function during late stages of tumor progression (18)(19)(20)(21)(22)(23)(24)(25)(26). To identify the CCGs that function early in mammary tumor development, we selected transposon insertion sites represented by the highest number of sequencing reads (SB insertions in the CCG in ≥3 tumors with ≥9 reads per tumor), arguing that these insertions would be present in the largest number of tumor cells.…”

“…New technologies that would provide a more complete understanding of the genetics of TNBC are still needed to deconvolute the complexity of this deadly cancer. Our laboratory and others have pioneered the use of transposon mutagenesis in mice as a tool for cancer gene discovery (18)(19)(20)(21)(22)(23)(24)(25)(26). Transposons induce cancer by randomly inserting into the mouse genome, mutating, and disrupting potential cancer genes.…”

Transposon mutagenesis identifies genes and cellular processes driving epithelial-mesenchymal transition in hepatocellular carcinoma

“…Transposon insertion sites were sequenced using splinkerette PCR to produce barcoded PCR products, which were then pooled and sequenced (22)(23)(24). To obtain the maximum amount of sequencing reads and avoid PCR bias (29), we used Illumina sequencing and acoustic shearing of genomic DNA (gDNA).…”

“…Our laboratory has successfully modeled more than 20 different types of human cancers in mice using Sleeping Beauty (SB) transposon mutagenesis and has identified hundreds of candidate cancer genes (CCGs), which show striking overlaps with genes mutated or deregulated in human cancers (22)(23)(24). We also have developed in vitro cell-based SB mutagenesis screens that have made it possible to identify genes involved in the malignant transformation of neural stem cells into glioma-initiating Significance Epithelial-mesenchymal transition (EMT) contributes to metastasis and chemoresistance in patients with hepatocellular carcinoma (HCC), but the genes driving EMT are poorly understood.…”