Differential Roles of Hypoxia-Inducible Factor 1α (HIF-1α) and HIF-2α in Hypoxic Gene Regulation

TLDR

Hypoxia‑induced transcription is governed by HIF, where HIF‑1α and HIF‑2α share DNA‑binding domains but differ in transactivation, leading to conflicting reports on whether HIF‑2α is dispensable or regulates specific genes. The study aimed to resolve this discrepancy by identifying HIF‑2α‑specific target genes. Researchers engineered HEK293 cells to express stabilized HIF‑1α or HIF‑2α under a tetracycline‑regulated promoter and performed DNA microarray analysis on cells expressing only HIF‑2α to assess hypoxic gene induction. The results showed that HIF‑2α regulates a broad set of hypoxia‑inducible genes beyond endothelial cells, while HIF‑1α uniquely activates glycolytic genes, demonstrating distinct target repertoires.

Abstract

Transcriptional responses to hypoxia are primarily mediated by hypoxia-inducible factor (HIF), a heterodimer of HIF-alpha and the aryl hydrocarbon receptor nuclear translocator subunits. The HIF-1alpha and HIF-2alpha subunits are structurally similar in their DNA binding and dimerization domains but differ in their transactivation domains, implying they may have unique target genes. Previous studies using Hif-1alpha(-/-) embryonic stem and mouse embryonic fibroblast cells show that loss of HIF-1alpha eliminates all oxygen-regulated transcriptional responses analyzed, suggesting that HIF-2alpha is dispensable for hypoxic gene regulation. In contrast, HIF-2alpha has been shown to regulate some hypoxia-inducible genes in transient transfection assays and during embryonic development in the lung and other tissues. To address this discrepancy, and to identify specific HIF-2alpha target genes, we used DNA microarray analysis to evaluate hypoxic gene induction in cells expressing HIF-2alpha but not HIF-1alpha. In addition, we engineered HEK293 cells to express stabilized forms of HIF-1alpha or HIF-2alpha via a tetracycline-regulated promoter. In this first comparative study of HIF-1alpha and HIF-2alpha target genes, we demonstrate that HIF-2alpha does regulate a variety of broadly expressed hypoxia-inducible genes, suggesting that its function is not restricted, as initially thought, to endothelial cell-specific gene expression. Importantly, HIF-1alpha (and not HIF-2alpha) stimulates glycolytic gene expression in both types of cells, clearly showing for the first time that HIF-1alpha and HIF-2alpha have unique targets.