Identification and characterization of the hypoxia-responsive element in human stanniocalcin-1 gene

Abstract

In this study, we aimed to identify the hypoxia-inducible factor-1 (HIF-1) binding motif in human STC1 gene promoter and to characterize the associated gene transactivation mechanism. Using normoxic human nasopharyngeal cancer cells (CNE2), we manipulated the stability of HIF-1α protein by overexpressing HIF-1α or the silencing of prolyl hydroxylase-2 (PHD2), to illustrate HIF-1 activation of STC1 promoter-driven luciferase activity. Subsequently luciferase activities of the deletion and mutated STC1 promoter constructs were investigated in HIF-1 overexpressed cells. The data revealed the presence of an authentic HRE motif in STC1 gene. This result was further supported by the chromatin immunoprecipitation (ChIP) assay. Using a similar experimental treatment, however, had no significant effect on the expression level of STC1 mRNA and protein. Moreover the activation of STC1 expression can be restored by the silencing of "factor inhibiting HIF-1" (FIH-1) in either HIF-1 overexpressed or PHD2 silenced cells. The data implied that the HIF-1-mediated STC1 gene expression required the recruitment of p300. This presumption was confirmed by the use of p300 inhibitor, chetomin and HIF-1α/p300 re-ChIP assay. Collectively our data provide the first evidence to show that STC1 is a FIH-inhibited gene with a functional HRE motif located at the upstream region between -2322/-2335. The data support the need for further investigation to reveal if STC1 can be used as a novel tumor marker for HIF-1 induction and for the monitoring of anti-angiogenic therapy. © 2009 Elsevier Ireland Ltd. All rights reserved.