Characterization of a complex genomic alteration on chromosome 2p that leads to four alternatively spliced fusion transcripts in the neuroblastoma cell lines IMR-5, IMR-5/75 and IMR-32.
Genetic aberrations in neuroblastoma (NB) have been extensively characterized over the last years. Alterations of the short arm of chromosome 2 (2p) have been of particular interest, since amplification of the MYCN oncogene on 2p24 is associated with an adverse outcome in NB patients. Here, we report on the characterization of a novel genomic rearrangement involving genetic material from 2p13 and 2p24 in NB cell lines that was discovered based on a serial analysis of gene expression (SAGE) profile of the MYCN-amplified NB cell line IMR-5. By analysis of a highly expressed SAGE tag not matching a Unigene cluster we identified four alternatively spliced corresponding transcripts, each of which consisted of the first 14 exons of the anthrax toxin receptor 1 gene (2p13.1) and varying combinations of exons of an unidentified gene located 1.3 Mb telomeric of MYCN (2p24.3) that was termed novel neuroblastoma gene 1. By Southern Blotting, Fluorescent In Situ Hybridization and Long Distance Inverse-PCR we disclosed that these transcripts result from a genomic alteration including material from distinct regions of chromosome 2p and four genomic breakpoints that are joined by short sequences of unknown origin. Furthermore, we show that this rearrangement lies within the homogeneous staining regions (HSR) in IMR-32 cells and is prevalent in both IMR-32 cells and their sub-clones IMR-5 and IMR-5/75, but not in a panel of 70 primary NB tumors. Our work is the first study discovering a fusion transcript based on a SAGE profile and for the first time precisely describes the DNA sequence of amplified breakpoint regions in NB.
Oberthuer
André
A
Children's Hospital, Department of Pediatric Oncology and Hematology and Center for Molecular Medicine Cologne, University of Cologne, Kerpener Strasse 62, D-50924 Köln, Cologne, Germany. andre.oberthuer@uk-koeln.de
Skowron
Matthias
M
Spitz
Rüdiger
R
Kahlert
Yvonne
Y
Westermann
Frank
F
Mehler
Kathrin
K
Berthold
Frank
F
Fischer
Matthias
M
D017398:Alternative Splicing; D015152:Blotting, Northern; D015139:Blotting, Southern; D045744:Cell Line, Tumor; D002889:Chromosomes, Human, Pair 2; D020869:Gene Expression Profiling; D016259:Genes, myc; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D008565:Membrane Proteins / Q000235:genetics; D008840:Microfilament Proteins; D009363:Neoplasm Proteins / Q000235:genetics; D009447:Neuroblastoma / Q000235:genetics* / Q000473:pathology; D012333:RNA, Messenger / Q000235:genetics*; D011956:Receptors, Cell Surface / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction
C473287:ANTXR1 protein, human; D008565:Membrane Proteins; D008840:Microfilament Proteins; D009363:Neoplasm Proteins; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D011993:Recombinant Fusion Proteins