title: RNA sequencing of Sleeping Beauty transposon-induced tumors detects transposon-RNA fusions in forward genetic cancer screens.
abstract: Forward genetic screens using Sleeping Beauty (SB)-mobilized T2/Onc transposons have been used to identify common insertion sites (CISs) associated with tumor formation. Recurrent sites of transposon insertion are commonly identified using ligation-mediated PCR (LM-PCR). Here, we use RNA sequencing (RNA-seq) data to directly identify transcriptional events mediated by T2/Onc. Surprisingly, the majority (∼80%) of LM-PCR identified junction fragments do not lead to observable changes in RNA transcripts. However, in CIS regions, direct transcriptional effects of transposon insertions are observed. We developed an automated method to systematically identify T2/Onc-genome RNA fusion sequences in RNA-seq data. RNA fusion-based CISs were identified corresponding to both DNA-based CISs (Cdkn2a, Mycl1, Nf2, Pten, Sema6d, and Rere) and additional regions strongly associated with cancer that were not observed by LM-PCR (Myc, Akt1, Pth, Csf1r, Fgfr2, Wisp1, Map3k5, and Map4k3). In addition to calculating recurrent CISs, we also present complementary methods to identify potential driver events via determination of strongly supported fusions and fusions with large transcript level changes in the absence of multitumor recurrence. These methods independently identify CIS regions and also point to cancer-associated genes like Braf. We anticipate RNA-seq analyses of tumors from forward genetic screens will become an efficient tool to identify causal events.
authors.affiliation: Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota 55455, USA;
authors.forename: Nuri A
authors.initials: NA
authors.lastname: Temiz
authors.affiliation: Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota 55455, USA; Department of Genetics, Cell Biology and Development, University of Minnesota, Minneapolis, Minnesota 55455, USA; Brain Tumor Program, University of Minnesota, Minneapolis, Minnesota 55455, USA; Department of Pediatrics, University of Minnesota, Minneapolis, Minnesota 55455, USA;
authors.forename: Branden S
authors.initials: BS
authors.lastname: Moriarity
authors.affiliation: Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota 55455, USA; Department of Genetics, Cell Biology and Development, University of Minnesota, Minneapolis, Minnesota 55455, USA; Department of Pediatrics, University of Minnesota, Minneapolis, Minnesota 55455, USA;
authors.forename: Natalie K
authors.initials: NK
authors.lastname: Wolf
authors.affiliation: Department of Anatomy and Cell Biology, University of Iowa, Iowa City, Iowa 52242, USA;
authors.forename: Jesse D
authors.initials: JD
authors.lastname: Riordan
authors.affiliation: Department of Anatomy and Cell Biology, University of Iowa, Iowa City, Iowa 52242, USA;
authors.forename: Adam J
authors.initials: AJ
authors.lastname: Dupuy
authors.affiliation: Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota 55455, USA; Department of Genetics, Cell Biology and Development, University of Minnesota, Minneapolis, Minnesota 55455, USA; Brain Tumor Program, University of Minnesota, Minneapolis, Minnesota 55455, USA; Department of Pediatrics, University of Minnesota, Minneapolis, Minnesota 55455, USA; Center for Genome Engineering, University of Minnesota, Minneapolis, Minnesota 55455, USA.
authors.forename: David A
authors.initials: DA
authors.lastname: Largaespada
authors.affiliation: Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota 55455, USA;
authors.forename: Aaron L
authors.initials: AL
authors.lastname: Sarver
mesh_terms: D002874:Chromosome Mapping; D004251:DNA Transposable Elements; D030541:Databases, Genetic; D055088:Early Detection of Cancer / Q000379:methods*; D050939:Gene Fusion; D005820:Genetic Testing / Q000379:methods; D006801:Humans; D016254:Mutagenesis, Insertional; D009369:Neoplasms / Q000175:diagnosis* / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D017423:Sequence Analysis, RNA; D014158:Transcription, Genetic
chemical_list: D004251:DNA Transposable Elements; D012333:RNA, Messenger