Co-amplification and concomitant high levels of expression of a DEAD box gene with MYCN in human neuroblastoma

Abstract

MYCN gene amplification is strongly correlated with poor prognosis in neuroblastoma (NB), the second most common solid pediatric tumor. However, increased MYCN expression seen in tumors that lack MYCN amplification does not correlate with aggressive clinical behavior. Whereas the MYCN gene spans only 7 kb, the MYCN amplicon has been shown to range in size from 350 kb to more than 1 Mb. Given the large size of the amplicon, it is possible that additional genes are co-amplified in NBs whose expression may contribute to the aggressive phenotype associated with MYCN-amplified tumors. We isolated a cDNA clone from a human NB library that is identical to DDXI, a gene recently reported to be preferentially expressed in two retinoblastoma cell lines that also express high levels of MYCN. DDXI belongs to a family of genes that encode DEAD (Asp-Glu-Ala-Asp) box proteins, putative ATP-dependent RNA helicases implicated in a number of cellular processes involving alterations of RNA secondary structure. We examined the frequency of DDXI amplification in 15 NB cell lines, 1 neuroepithelioma cell line, and 122 NB tumors by Southern blot analyses, and we found that 7 of 10 MYCN-amplified cell lines and 27 of 40 (68%) MYCN-amplified tumors also harbored multiple copies of the DDXI gene. Amplification of DDXI was associated with high levels of DDXI mRNA expression in the NB cell lines and tumors as examined by Northern analysis. Neither DDXI gene amplification nor enhanced expression was observed in tumors or cell lines that lacked MYCN amplification. Because RNA helicases play important roles in both post-transcriptional and translational gene regulation, high levels of DDXI expression consequent to genomic amplification may contribute to the malignant phenotype of a subset of NBs.