Regulation of HIF prolyl hydroxylases by hypoxia-inducible factors.
Hypoxia and induction of hypoxia-inducible factors (HIF-1alpha and HIF-2alpha) is a hallmark of many tumors. Under normal oxygen tension HIF-alpha subunits are rapidly degraded through prolyl hydroxylase dependent interaction with the von Hippel-Lindau (VHL) tumor suppressor protein, a component of E3 ubuiquitin ligase complex. Using microarray analysis of VHL mutated and re-introduced cells, we found that one of the prolyl hydroxylases (PHD3) is coordinately expressed with known HIF target genes, while the other two family members (PHD1 and 2) did not respond to VHL. We further tested the regulation of these genes by HIF-1 and HIF-2 and found that siRNA targeted degradation of HIF-1alpha and HIF-2alpha results in decreased hypoxia-induced PHD3 expression. Ectopic overexpression of HIF-2alpha in two different cell lines provided a much better induction of PHD3 gene than HIF-1alpha. In contrast, we demonstrate that PHD2 is not affected by overexpression or downregulation of HIF-2alpha. However, induction of PHD2 by hypoxia has HIF-1-independent and -dependent components. Short-term hypoxia (4 h) results in induction of PHD2 independent of HIF-1, while PHD2 accumulation by prolonged hypoxia (16 h) was decreased by siRNA-mediated degradation of HIF-1alpha subunit. These data further advance our understanding of the differential role of HIF factors and putative feedback loop in HIF regulation.
Aprelikova
Olga
O
Laboratory of Biosystems and Cancer, Center for Cancer Research, National Cancer Institute, Bethesda, Maryland 20892, USA.
Chandramouli
Gadisetti V R
GV
Wood
Matthew
M
Vasselli
James R
JR
Riss
Joseph
J
Maranchie
Jodi K
JK
Linehan
W Marston
WM
Barrett
J Carl
JC
D051792:Basic Helix-Loop-Helix Proteins; D015687:Cell Hypoxia / Q000235:genetics; D045744:Cell Line, Tumor; D004268:DNA-Binding Proteins / Q000235:genetics*; D049308:Dioxygenases; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D064799:Hypoxia-Inducible Factor-Proline Dioxygenases; D017874:Immediate-Early Proteins / Q000235:genetics*; D009154:Mutation; D020411:Oligonucleotide Array Sequence Analysis; D011393:Procollagen-Proline Dioxygenase / Q000235:genetics*; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D015534:Trans-Activators / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D014162:Transfection; D025521:Tumor Suppressor Proteins / Q000235:genetics / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism; D051794:Von Hippel-Lindau Tumor Suppressor Protein; D000099162:Endothelial PAS Domain-Containing Protein 1
D051792:Basic Helix-Loop-Helix Proteins; D004268:DNA-Binding Proteins; D049308:Dioxygenases; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D064799:Hypoxia-Inducible Factor-Proline Dioxygenases; D017874:Immediate-Early Proteins; D011393:Procollagen-Proline Dioxygenase; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D015534:Trans-Activators; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; D044767:Ubiquitin-Protein Ligases; D051794:Von Hippel-Lindau Tumor Suppressor Protein; D000099162:Endothelial PAS Domain-Containing Protein 1; C497442:HIF1A protein, human; C490190:EGLN1 protein, human; C488894:EGLN3 protein, human; C497446:VHL protein, human