The functional -443T/C osteopontin promoter polymorphism influences osteopontin gene expression in melanoma cells via binding of c-Myb transcription factor.

Abstract

In the present report, the possible role of a recently described functional polymorphism of the osteopontin (OPN) promoter at position -443 (-443T/C) for OPN expression in melanoma cells was addressed. As shown by real-time PCR analysis, melanoma metastases that were homozygous for the -443C allele expressed significantly higher levels of OPN mRNA compared with those that were either heterozygous (-443T/C) or homozygous for the -443T allele. In line with this, immunoblotting showed significantly enhanced baseline and bFGF-induced OPN protein expression in melanoma cell lines which were homozygous for the -443C allele, compared with cell lines with other allelic variants. Similar results were obtained in in vitro luciferase assays. Chromatin immunoprecipitation (ChIP) demonstrated binding of c-Myb to the -443 OPN promoter region, and binding could significantly be enhanced after bFGF stimulation. Moreover, as shown by electrophoretic mobility shift assays (EMSA), recombinant DNA-binding domain of c-Myb bound in a sequence-specific manner to this region. Finally, the role of c-Myb for OPN gene regulation via binding to the -443 promoter region could be further substantiated by ectopic overexpression of c-Myb in melanoma cells, using different reporter gene constructs. Taken together, it is demonstrated that the -443 promoter region exerts influence on OPN gene expression in melanoma cells, and differential binding of c-Myb transcription factor appears to play a major role in this process. These findings might be a feasible explanation for different OPN expression levels in metastatic tumors and may also have prognostic and therapeutic relevance.

DOI: 10.1002/mc.20452
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@article{Schultz2009TheF, title={The functional -443T/C osteopontin promoter polymorphism influences osteopontin gene expression in melanoma cells via binding of c-Myb transcription factor.}, author={Julia Schultz and Peter Lorenz and Saleh M. Ibrahim and G{\"{u}nther Kundt and Gerd E. Gross and Manfred Kunz}, journal={Molecular carcinogenesis}, year={2009}, volume={48 1}, pages={14-23} }