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MDM2 regulates MYCN mRNA stabilization and translation in human neuroblastoma cells.

ABSTRACT: The MYCN gene has a critical role in determining the clinical behavior of neuroblastoma. Although it is known that genomic amplification occurs in high-risk subsets, it remains unclear how MYCN expression is regulated in the pathogenesis of neuroblastomas. Here, we report that MYCN expression was regulated by the oncoprotein MDM2 at the post-transcriptional level and was associated with neuroblastoma cell growth. Increasing MDM2 by ectopic overexpression in the cytoplasm enhanced both mRNA and protein expression of MYCN. Mechanistic studies found that the C-terminal RING domain of the MDM2 protein bound to the MYCN mRNA's AREs within the 3'UTR and increased MYCN 3'UTR-mediated mRNA stability and translation. Conversely, MDM2 silencing by specific siRNA rendered the MYCN mRNA unstable and reduced the abundance of the MYCN protein in MYCN-amplified neuroblastoma cell lines. Importantly, this MDM2 silencing resulted in a remarkable inhibition of neuroblastoma cell growth and induction of cell death through a p53-independent pathway. Our results indicate that MDM2 has a p53-independent role in the regulation of both MYCN mRNA stabilization and its translation, suggesting that MDM2-mediated MYCN expression is one mechanism associated with growth of MYCN-associated neuroblastoma and disease progression.

SUBMITTER: Gu L

PROVIDER: S-EPMC3213308 | biostudies-literature | 2012 Mar

REPOSITORIES: biostudies-literature

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MDM2 regulates MYCN mRNA stabilization and translation in human neuroblastoma cells.

Gu L L Zhang H H He J J Li J J Huang M M Zhou M M

Oncogene 20110808 11

The MYCN gene has a critical role in determining the clinical behavior of neuroblastoma. Although it is known that genomic amplification occurs in high-risk subsets, it remains unclear how MYCN expression is regulated in the pathogenesis of neuroblastomas. Here, we report that MYCN expression was regulated by the oncoprotein MDM2 at the post-transcriptional level and was associated with neuroblastoma cell growth. Increasing MDM2 by ectopic overexpression in the cytoplasm enhanced both mRNA and p ...[more]

PMID: 21822304

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Project description:BackgroundMYCN is amplified in small cell lung cancers and several pediatric tumors, including alveolar rhabdomyosarcomas and neuroblastomas. MYCN protein is known to play a key oncogenic role in both alveolar rhabdomyosarcomas and neuroblastomas. MYCN opposite strand (MYCNOS) is a gene located on the antisense strand to MYCN that encodes alternatively spliced transcripts, two of which (MYCNOS-01 and MYCNOS-02) are known to be expressed in neuroblastoma and small cell lung cancer with reciprocal regulation between MYCNOS-02 and MYCN reported for neuroblastomas. We sought to determine a functional role for MYCNOS-01 in alveolar rhabdomyosarcoma and neuroblastoma cells and identify any associated regulatory effects between MYCN and MYCNOS-01.MethodsMYCNOS-01, MYCNOS-02 and MYCN expression levels were assessed in alveolar rhabdomyosarcoma and neuroblastoma cell lines and tumor samples from patients using Affymetrix microarray data and quantitative RT-PCR. Following MYCNOS-01 or MYCN siRNA knockdown and MYCNOS-01 overexpression, transcript levels were assayed by quantitative RT-PCR and MYCN protein expression assessed by Western blot and immunofluorescence. Additionally, effects on cell growth, apoptosis and cell cycle profiles were determined by a metabolic assay, caspase activity and flow cytometry, respectively.ResultsMYCNOS-01 transcript levels were generally higher in NB and RMS tumor samples and cell lines with MYCN genomic amplification. RNA interference of MYCNOS-01 expression did not alter MYCN transcript levels but decreased MYCN protein levels. Conversely, MYCN reduction increased MYCNOS-01 transcript levels, creating a negative feedback loop on MYCN protein levels. Reduction of MYCNOS-01 or MYCN expression decreased cell growth in MYCN-amplified alveolar rhabdomyosarcoma and neuroblastoma cell lines. This is consistent with MYCNOS-01-mediated regulation of MYCN contributing to the phenotype observed.ConclusionsAn alternative transcript of MYCNOS, MYCNOS-01, post-transcriptionally regulates MYCN levels and affects growth in MYCN-amplified rhabdomyosarcoma and neuroblastoma cells.

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MDM2 regulates MYCN mRNA stabilization and translation in human neuroblastoma cells.

Publications

MDM2 regulates MYCN mRNA stabilization and translation in human neuroblastoma cells.

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