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Osteopontin (OPN) is a secreted phosphoglycoprotein that has been linked to tumor progression and survival in several solid tumors, including head and neck cancers. Previous studies showed that OPN expression is induced by tumor hypoxia, and its plasma levels can serve as a surrogate marker for tumor hypoxia and treatment outcome in head and neck cancer patients. In this study, we investigate the transcriptional mechanism by which hypoxia enhances OPN expression. We found that OPN is induced in head and neck squamous cell carcinoma (HNSCC) cell lines and in NIH3T3 cells by hypoxia at both mRNA and protein levels in a time-dependent manner. Actinomycin D chase experiments showed that hypoxic induction of OPN was not due to increased mRNA stability. Deletion analyses of the mouse OPN promoter regions indicated that a ras-activated enhancer (RAE) located at -731 to -712 relative to the transcription start site was essential for hypoxia-enhanced OPN transcription. Using electrophoretic mobility shift assays with the RAE DNA sequence, we found that hypoxia induced sequence-specific DNA-binding complexes. Furthermore, hypoxia and ras exposure resulted in an additive induction of OPN protein and mRNA levels that appeared to be mediated by the RAE. Induction of OPN through the RAE element by hypoxia is mediated by an Akt-kinase signaled pathway as decreasing Akt levels with dominant negative constructs resulted in inhibition of OPN induction by hypoxia. Taken together, these results have identified a new hypoxia responsive transcriptional enhancer that is regulated by Akt signaling.

Original publication

DOI

10.1038/sj.onc.1208800

Type

Journal article

Journal

Oncogene

Publication Date

29/09/2005

Volume

24

Pages

6555 - 6563

Keywords

3T3 Cells, Animals, Carcinoma, Squamous Cell, Cell Hypoxia, Dactinomycin, Enhancer Elements, Genetic, Gene Expression Regulation, Genes, ras, Head and Neck Neoplasms, Mice, Osteopontin, Protein-Serine-Threonine Kinases, Proto-Oncogene Proteins, Proto-Oncogene Proteins c-akt, Proto-Oncogene Proteins p21(ras), RNA Stability, RNA, Messenger, Response Elements, Sialoglycoproteins, Transcription, Genetic