Metal ions-stimulated iron oxidation in hydroxylases facilitates stabilization of HIF-1 alpha protein.
The exposure of cells to several metal ions stabilizes HIF-1 alpha protein. However, the molecular mechanisms are not completely understood. They may involve inhibition of hydroxylation by either substitution of iron by metal ions or by iron oxidation in the hydroxylases. Here we provide evidence supporting the latter mechanism. We show that HIF-1 alpha stabilization in human lung epithelial cells occurred following exposure to various metal and metalloid ions, including those that cannot substitute for iron in the hydroxylases. In each case addition of the reducing agent ascorbic acid (AA)* abolished HIF-1 alpha protein stabilization. To better understand the role of iron oxidation in hydroxylase inhibition and to define the role of AA in the enzyme recovery we applied molecular modeling techniques. Our results indicate that the energy required for iron substitution by Ni(II) in the enzyme is high and unlikely to be achieved in a biological system. Additionally, computer modeling allowed us to identify a tridentate coordination of AA with the enzyme-bound iron, which explains the specific demand for AA as the iron reductant. Thus, the stabilization of HIF-1 alpha by numerous metal ions that cannot substitute for iron in the enzyme, the alleviation of this effect by AA, and our computer modeling data support the hypothesis of iron oxidation in the hydroxylases following exposure to metal ions.
Kaczmarek
Monika
M
Laboratory of Comparative Carcinogenesis, National Cancer Institute at Frederick, Frederick, Maryland 21702, USA.
Cachau
Raul E
RE
Topol
Igor A
IA
Kasprzak
Kazimierz S
KS
Ghio
Andy
A
Salnikow
Konstantin
K
D000975:Antioxidants / Q000494:pharmacology; D001205:Ascorbic Acid / Q000494:pharmacology; D015153:Blotting, Western; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D017930:Genes, Reporter / Q000187:drug effects; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000737:chemistry / Q000378:metabolism*; D007202:Indicators and Reagents; D007501:Iron / Q000378:metabolism*; D007700:Kinetics; D008156:Luciferases / Q000235:genetics; D009682:Magnetic Resonance Spectroscopy; D008670:Metals / Q000633:toxicity*; D006899:Mixed Function Oxygenases / Q000378:metabolism*; D008958:Models, Molecular; D008968:Molecular Conformation; D010084:Oxidation-Reduction; D013268:Stimulation, Chemical
D000975:Antioxidants; C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007202:Indicators and Reagents; D008670:Metals; D007501:Iron; D006899:Mixed Function Oxygenases; D008156:Luciferases; D001205:Ascorbic Acid