Copper-dependent activation of hypoxia-inducible factor (HIF)-1: implications for ceruloplasmin regulation.
Cellular oxygen partial pressure is sensed by a family of prolyl-4-hydroxylase domain (PHD) enzymes that modify hypoxia-inducible factor (HIF)alpha subunits. Upon hydroxylation under normoxic conditions, HIFalpha is bound by the von Hippel-Lindau tumor suppressor protein and targeted for proteasomal destruction. Since PHD activity is dependent on oxygen and ferrous iron, HIF-1 mediates not only oxygen- but also iron-regulated transcriptional gene expression. Here we show that copper (CuCl(2)) stabilizes nuclear HIF-1alpha under normoxic conditions, resulting in hypoxia-response element (HRE)-dependent reporter gene expression. In in vitro hydroxylation assays CuCl(2) inhibited prolyl-4-hydroxylation independently of the iron concentration. Ceruloplasmin, the main copper transport protein in the plasma and a known HIF-1 target in vitro, was also induced in vivo in the liver of hypoxic mice. Both hypoxia and CuCl(2) increased ceruloplasmin (as well as vascular endothelial growth factor [VEGF] and glucose transporter 1 [Glut-1]) mRNA levels in hepatoma cells, which was due to transcriptional induction of the ceruloplasmin gene (CP) promoter. In conclusion, our data suggest that PHD/HIF/HRE-dependent gene regulation can serve as a sensory system not only for oxygen and iron but also for copper metabolism, regulating the oxygen-, iron- and copper-binding transport proteins hemoglobin, transferrin, and ceruloplasmin, respectively.
Martin
Falk
F
Carl-Ludwig-Institute of Physiology, University of Leipzig, Germany.
Linden
Tobias
T
Katschinski
Dörthe M
DM
Oehme
Felix
F
Flamme
Ingo
I
Mukhopadhyay
Chinmay K
CK
Eckhardt
Katrin
K
Tröger
Juliane
J
Barth
Sandra
S
Camenisch
Gieri
G
Wenger
Roland H
RH
D000818:Animals; D016466:CHO Cells; D006528:Carcinoma, Hepatocellular / Q000378:metabolism; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D049109:Cell Proliferation; D002570:Ceruloplasmin / Q000378:metabolism*; D004396:Coloring Agents / Q000494:pharmacology; D003300:Copper / Q000378:metabolism*; D006224:Cricetinae; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D051272:Glucose Transporter Type 1; D006367:HeLa Cells; D006801:Humans; D000860:Hypoxia; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D015151:Immunoblotting; D007501:Iron / Q000378:metabolism*; D008099:Liver / Q000378:metabolism; D008156:Luciferases / Q000378:metabolism; D051379:Mice; D008856:Microscopy, Fluorescence; D009004:Monosaccharide Transport Proteins / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism*; D010100:Oxygen / Q000378:metabolism*; D011393:Procollagen-Proline Dioxygenase / Q000378:metabolism; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D013778:Tetrazolium Salts / Q000494:pharmacology; D013844:Thiazoles / Q000494:pharmacology; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D014162:Transfection; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism
D004396:Coloring Agents; D004268:DNA-Binding Proteins; D051272:Glucose Transporter Type 1; C497442:HIF1A protein, human; C497443:Hif1a protein, mouse; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D009004:Monosaccharide Transport Proteins; D009687:Nuclear Proteins; D012333:RNA, Messenger; C495677:SLC2A1 protein, human; C495678:Slc2a1 protein, mouse; D013778:Tetrazolium Salts; D013844:Thiazoles; D014157:Transcription Factors; D042461:Vascular Endothelial Growth Factor A; D003300:Copper; D007501:Iron; D008156:Luciferases; D011393:Procollagen-Proline Dioxygenase; D002570:Ceruloplasmin; C022616:thiazolyl blue; C029892:cupric chloride; D010100:Oxygen