Integration of genomic and proteomic data to identify MYCN-regulated genes, proteins, and interaction networks in neuroblastoma cells.

Abstract

9584 Background: Neuroblastoma is the most common extracranial solid tumor found in children. The most common marker prognostic of poor outcome is amplification of the MYCN oncogene, yet the biological programs by which MYCN affects its aggressive phenotype are largely unknown. In order to identify biological pathways affected by MYCN amplification, we performed global analysis of genes and proteins in the Tet 21/N neuroblastoma cell line. Methods: MYCN expression in Tet21/N cells is regulated by tetracycline. Gene arrays were performed on MYCN-high and MYCN-low Tet21/N cells and SH-EP parental cells using Affymetrix GeneChip Human Exon 1.0 ST. For proteomic analysis, the cells were labeled with stable isotopes for relative quantitation, enriched for phosphoproteins to enhance detection of signaling proteins, and analyzed by GeLC-MS/MS analysis. Integrative pathway analysis was performed on the genomic and proteomic datasets using DAVE and GeneGo. Results: Integrating genomic and proteomic data in MYCN-high and MYCN-low expressing cells identified 88 proteins and 300 genes that change in abundance. We compare these results with previous studies and find both novel and previously identified genes and proteins. Integrating proteomic and genomic data identified over 50 gene products that are present in both analyses and are altered in abundance in response to modulating MYCN expression. The majority of these have discordant abundance changes, with protein levels more frequently altered with no change in gene expression. We have validated changes in protein levels using Western blots including NPM1 and MATR3. We present interaction networks and pathways that correlate with MYCN expression. Conclusions: We showed a significant discordance between gene and protein expression, underscoring the need for integrative genomic and proteomic analysis to describe complex systems. Our analysis identified previously reported and novel genes and proteins and networks regulated by MYCN expression that may provide new insights into the biology of MYCN-amplified tumors.