Xanthine oxidase-dependent regulation of hypoxia-inducible factor in cancer cells.
During chemical hypoxia induced by cobalt chloride (CoCl2), hypoxia-inducible factor 1alpha (HIF1-alpha) mediates the induction of a variety of genes including erythropoietin and vascular endothelial growth factor. We used glioma cells with oxidative phosphorylation-dependent (D54-MG) and glycolytic-dependent (U251-MG) phenotypes to monitor HIF1-alpha regulation in association with redox responsiveness to CoCl2 treatment. We showed that CoCl2 increased xanthine oxidase (XO)-derived reactive oxygen species (ROS), which causes accumulation of HIF1-alpha protein in U251-MG cells. Under these conditions, blockade of XO activity by pharmacologic (N-acetyl-L-cysteine or allopurinol) or molecular (by small interfering RNA) approaches significantly attenuated HIF1-alpha expression. Exogenous H2O2 stabilizes HIF1-alpha protein. XO was present in these cells and was the primary source of free radicals. We also showed higher XO activity in cells exposed to CoCl2 compared with cells grown in normoxia. From the experiments shown here, we concluded that ROS were indeed generated in D54-MG cells exposed to CoCl2 but it was unlikely that ROS participated in the hypoxic signal transduction pathways in this cell type. Possibly, cell type-dependent and stimulus-dependent factors may control ROS dependency or redox sensitivity of HIF1-alpha and thus HIF1-alpha activation either directly or by induction of specific signaling cascades. Our findings reveal that XO-derived ROS is a novel and critical component of HIF1-alpha regulation in U251-MG cells, pointing toward a more general role of this transcription factor in tumor progression.
Griguer
Corinne E
CE
Department of Surgery, University of Alabama at Birmingham, Birmingham, Alabama 35294-0005, USA. cgriguer@uab.edu
Oliva
Claudia R
CR
Kelley
Eric E
EE
Giles
Gregory I
GI
Lancaster
Jack R
JR
Gillespie
G Yancey
GY
D000111:Acetylcysteine / Q000494:pharmacology; D045744:Cell Line, Tumor; D003035:Cobalt / Q000494:pharmacology; D005609:Free Radicals / Q000378:metabolism; D005910:Glioma / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000096:biosynthesis* / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics; D017382:Reactive Oxygen Species / Q000378:metabolism*; D014162:Transfection; D014969:Xanthine Oxidase / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
D005609:Free Radicals; C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; D003035:Cobalt; D014969:Xanthine Oxidase; C018021:cobaltous chloride; D000111:Acetylcysteine