Evidence for a Lack of a Direct Transcriptional Suppression of the Iron Regulatory Peptide Hepcidin by Hypoxia-Inducible Factors

Abstract

BackgroundHepcidin is a major regulator of iron metabolism and plays a key role in anemia of chronic disease, reducing intestinal iron uptake and release from body iron stores. Hypoxia and chemical stabilizers of the hypoxia-inducible transcription factor (HIF) have been shown to suppress hepcidin expression. We therefore investigated the role of HIF in hepcidin regulation.Methodology/Principal FindingsHepcidin mRNA was down-regulated in hepatoma cells by chemical HIF stabilizers and iron chelators, respectively. In contrast, the response to hypoxia was variable. The decrease in hepcidin mRNA was not reversed by HIF-1α or HIF-2α knock-down or by depletion of the HIF and iron regulatory protein (IRP) target transferrin receptor 1 (TfR1). However, the response of hepcidin to hypoxia and chemical HIF inducers paralleled the regulation of transferrin receptor 2 (TfR2), one of the genes critical to hepcidin expression. Hepcidin expression was also markedly and rapidly decreased by serum deprivation, independent of transferrin-bound iron, and by the phosphatidylinositol 3 (PI3) kinase inhibitor LY294002, indicating that growth factors are required for hepcidin expression in vitro. Hepcidin promoter constructs mirrored the response of mRNA levels to interleukin-6 and bone morphogenetic proteins, but not consistently to hypoxia or HIF stabilizers, and deletion of the putative HIF binding motifs did not alter the response to different hypoxic stimuli. In mice exposed to carbon monoxide, hypoxia or the chemical HIF inducer N-oxalylglycine, liver hepcidin 1 mRNA was elevated rather than decreased.Conclusions/SignificanceTaken together, these data indicate that hepcidin is neither a direct target of HIF, nor indirectly regulated by HIF through induction of TfR1 expression. Hepcidin mRNA expression in vitro is highly sensitive to the presence of serum factors and PI3 kinase inhibition and parallels TfR2 expression.

Hepatic hypoxia-inducible factor-2 down-regulates hepcidin expression in mice through an erythropoietin-mediated increase in erythropoiesis

“…8 However, other studies in HepG2 cells showed that overexpression or knockdown of HIF-1 did not affect hepcidin expression 18 and have suggested indirect pathways including 2-oxoglutarate- 16 or ROS-dependent pathways. Moreover, HIF-2 but not HIF-1 seems to play a major role in iron metabolism and erythropoiesis.…”

“…Whether hepcidin is regulated directly or through alternative pathways by HIF has been the subject of intense research in vitro. 10,[16][17][18] Our previous in vitro studies suggested a direct regulation of hepcidin by HIF. 8 However, our in vivo data could not exclude indirect alternative pathways.…”

Alterations in the iron homeostasis network: A driving force for macrophage-mediated hepatitis C virus persistency

“…25,26 Secreted protein expression was also increased throughout infection, albeit not as dramatically as mRNA. The latter may be due to post-transcriptional regulation of HAMP gene expression, including mRNA destabilisation 27 and secretion modulation. regulator in hepatocytes, 29 FPN was reduced during early infection.…”

Hepatitis C virus core+1/ARF protein decreases hepcidin transcription through an AP1 binding site

“…2a). In agreement with published literature, these included hypoxia-inducible factor (HIF) (Volke et al, 2009) The AP1, C/EBP and STAT3 were mutated in the HAMPluc construct using the QuikChange kit (Agilent). Huh7 cells were co-transfected with each mutated reporter and core or core+1/ARFP expression vectors.…”