Binding of a 40-kDa protein to the N-myc 3'-untranslated region correlates with enhanced N-myc expression in human neuroblastoma.
Subclones of neuronal (N) and non-neuronal (S) cells established from neuroblastoma tumors cultured in vitro differ in their growth characteristics and N-myc expression. N (W-N) cells derived from the NBL-W cell line express 5-fold higher levels of N-myc mRNA and 10-12-fold higher levels of protein than S cells (W-S), despite having the same N-myc copy number. This study demonstrates that the steady-state levels of N-myc are largely determined by differences in N-myc mRNA stability. The half-life of N-myc mRNA in the W-N cells is approximately 35 min compared with approximately 6 min in the W-S cells. Turnover of labile mRNAs is thought to be mediated in part by the interactions of trans-acting factors with elements within the 3'-untranslated region. RNA UV cross-linking assays using W-N cell lysate demonstrate abundant quantities of a protein complex that is 40 kDa in size (p40) that binds to the N-myc 3'-untranslated region. p40 is barely detectable in W-S cells. We have mapped two distinct regions within the 3'-UTR that specifically bind p40 (base pairs 5694-5715 and 6465-6482). Analysis of nine additional neuroblastoma cell lines shows that p40 activity correlates with enhanced expression of N-myc. p40 activity is also detected in 5 of 19 primary neuroblastomas, and activity is associated with clinically aggressive disease. In the accompanying study, we identify p40 as a member of the embryonic lethal abnormal vision (ELAV)-like family of RNA-binding proteins. Our studies suggest that ELAV-like proteins may play a role in the regulation of N-myc mRNA turnover and thereby modulate the steady-state levels of N-myc expression and tumor cell phenotype.
Chagnovich
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Program in Tumor Cell Biology and Children's Memorial Hospital, Robert H. Lurie Cancer Center, Northwestern University Medical School, Chicago, Illinois 60611, USA.
Cohn
S L
SL
D001665:Binding Sites; D051959:ELAV Proteins; D000067777:ELAV-Like Protein 2; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008970:Molecular Weight; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D009447:Neuroblastoma / Q000378:metabolism*; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D012995:Solubility; D014158:Transcription, Genetic
D051959:ELAV Proteins; D000067777:ELAV-Like Protein 2; C086811:ELAVL2 protein, human; D009419:Nerve Tissue Proteins; D016271:Proto-Oncogene Proteins c-myc; D016601:RNA-Binding Proteins