FIG. 5.

Analysis of HIF-1 interaction with the putative HREs in the hTERT promoter. (a and b) ³²P end-labeled TERT/HRE1 or ³²P end-labeled TERT/HRE2 oligonucleotide was used as a probe. For the competition assays, a 50-fold or 200-fold molar excess of the HRE was used. For the supershift assay, anti-HIF-1α antibody (0.5 μg) was added to the binding reaction. The thick arrow indicates HIF-1/DNA complexes, and the thin arrows indicate supershifted bands. (c) The HIF-1 consensus oligonucleotide was end-labeled with ³²P. For the competition assays, a 50-fold or 200-fold molar excess of the TERT/HRE1 and of the TERT/HRE2 oligonucleotides were added to the binding reaction, respectively. For the supershift assay, anti-HIF-1α antibody (0.5 μg) was added to the binding reaction. The thick arrow indicates HIF-1/DNA complexes, and the thin arrows indicate supershifted bands. NS, nonspecific.