Co-amplification of a novel gene, NAG, with the N-myc gene in neuroblastoma

Substantial evidence implicates amplification of the N-myc gene with aggres sive tumor growth and poor outcome in neuroblastoma. However some evidence suggests that this gene alone is not the sole determinant of outcome in N-m yc amplified tumors. We have searched for genes that co-amplify with N-myc in neuroblastoma by means of two-dimensional analysis of genomic restrictio n digests. Using this approach, we have identified and cloned a novel genom ic fragment which is co-amplified with N-myc in neuroblastomas, This fragme nt was mapped in close vicinity to N-myc on chromosome arm 2p24, It was amp lified in 5/8 N-myc amplified neuroblastoma cell lines and in 9/13 N-myc am plified tumors. Using a PCR-based approach we isolated a 4.5 kb c-DNA seque nce that is partly contained in the genomic fragment, The open reading fram e of the cDNA encodes a predicted protein of 1353 amino acids (aa), The hom ology of the predicted protein, which we designated NAG (neuroblastoma ampl ified gene), to a C, elegans protein of as yet unknown function, and its ub iquious expression suggest that NAG may serve an essential function, By Nor thern blot analysis we showed that amplification of the cloned gene correla tes with over-expression in neuroblastoma cell lines. Amplification and con sequent over-expression of NAG may, therefore, contribute to the phenotype of a subset of neuroblastomas.