Inhibiting HIF-1 Transcriptional Activity

Abstract

Hypoxia-inducible factor 1 (HIF-1) is a transcription factor that regulates genes important for cellular adaptation to low O 2 conditions. The HIF-1α subunit is regulated under normoxia conditions by its interaction with the E3 ligase von Hippel-Lindau protein (VHL), which leads to ubiquitination and degradation. Mahon et al. used the yeast two-hybrid system to identify a protein, factor-inhibiting HIF-1 (FIH-1), that interacts with the transcriptional transactivation domain of HIF-1α. FIH-1 inhibits HIF-1 reporter gene activity under low O 2 conditions, and in a test that eliminates the proline hydroxylation-dependent VHL-mediated degradation of HIF-1, inhibition under normoxia conditions could also be detected. Glutathione- S -transferase fusion proteins were used to demonstrate that FIH-1, HIF-1α, and VHL could interact through nonoverlapping regions, indicating that a ternary complex could form. The effect of expression of VHL, FIH-1, or both, on reporter gene expression stimulated by fusion proteins of the GAL4 DNA binding domain and fragments of the COOH-terminus of HIF-1α representing the binding sites for VHL, FIH-1, or both, was tested. The fragment with only the VHL-binding site was only able to inhibit transcription when both VHL and FIH-1 were expressed, suggesting that VHL recruits FIH-1 and this is required for transcriptional inhibition. Finally, VHL interacted with histone deactylases in vitro; thus, recruitment of these corepressors of transcription may be one mechanism by which the transcriptional activity of HIF-1 is inhibited. P. C. Mahon, K. Hirota, G. L. Semenza, FIH-1: A novel protein that interacts with HIF-1α and VHL to mediate repression of HIF-1 transcriptional activity. Genes Dev. 15 , 2675-2686 (2001). [Abstract] [Full Text]