N-MYC regulates focal adhesion kinase expression in human neuroblastoma.

Abstract

N-MYC is a transcription factor that plays an important role in cellular survival in neuroblastoma, and amplification of the N-MYC oncogene is the primary adverse prognostic indicator for neuroblastoma. Focal adhesion kinase (FAK) is a survival factor that has been shown to be overexpressed in many types of human cancers. In this study, we investigated the role of N-MYC regulation of FAK expression in neuroblastoma. We first found a correlation between N-MYC and FAK expression in neuroblastoma. Real time quantitative PCR demonstrated an increase in FAK mRNA abundance in the N-MYC-amplified IMR-32 compared with the nonamplified SK-N-AS neuroblastoma cell lines. FAK protein expression also correlated positively with N-MYC expression in the N-MYC-amplified IMR-32 versus nonamplified SK-N-AS neuroblastoma cell lines. The same results were seen with the isogenic N-MYC(+) (Tet(-)) and N-MYC(-) (Tet(+)) neuroblastoma cell lines. Promoter-reporter assays showed that activity of the FAK promoter was increased in the N-MYC-amplified IMR-32 cell line, in the N-MYC-transfected SK-N-AS nonamplified cell line, and in the isogenic N-MYC(+) (Tet(-)) neuroblastoma cell lines compared with the nonamplified and N-MYC-nonexpressing cell lines. We also identified two N-MYC binding sites in the FAK promoter sequence and showed binding of N-MYC transcription factor to the FAK promoter through electrophoretic mobility shift, chromatin immunoprecipitation, and dual luciferase assays. Finally down-regulation of FAK expression in N-MYC-inducible neuroblastoma cell lines with FAK small interfering RNA or a dominant-negative FAK inhibitor (AdFAK-CD) significantly decreased viability and increased apoptosis in the N-MYC(+) (Tet(-)) cells compared with the isogenic N-MYC(-) (Tet(+)) cells, demonstrating the biological significance of FAK overexpression in the N-MYC-expressing cell lines. This is the first report linking N-MYC and FAK in neuroblastoma, and it clearly demonstrates that N-MYC induces FAK expression. The results indicate that N-MYC regulation of FAK expression can control cellular functions in isogenic N-MYC(-/+) (Tet(+/-)) neuroblastoma cell lines.

• Authors: Elizabeth A Beierle, Angelica Trujillo, Abhilasha Nagaram, Elena V Kurenova, Richard Finch, Xiaojie Ma, Jennifer Vella, William G Cance, Vita M Golubovskaya
• Journal: The Journal of biological chemistry (J Biol Chem) Vol. 282 Issue 17 Pg. 12503-16 (Apr 27 2007) ISSN: 0021-9258 [Print] United States
• PMID: 17327229 (Publication Type: Comparative Study, Journal Article)

Chemical References

• Protein Kinase Inhibitors
• Proto-Oncogene Proteins c-myc
• RNA, Small Interfering
• Focal Adhesion Kinase 1
• PTK2 protein, human

Topics

• Cell Line, Tumor
• Focal Adhesion Kinase 1 (antagonists & inhibitors, biosynthesis, genetics)
• Gene Amplification
• Gene Expression Regulation, Enzymologic (drug effects)
• Gene Expression Regulation, Neoplastic (drug effects)
• Humans
• Neuroblastoma (enzymology, genetics)
• Protein Kinase Inhibitors (pharmacology)
• Proto-Oncogene Proteins c-myc (metabolism)
• RNA, Small Interfering (pharmacology)
• Response Elements