Hypoxia response element of the human vascular endothelial growth factor gene mediates transcriptional regulation by nitric oxide: control of hypoxia-inducible factor-1 activity by nitric oxide.
Nitric oxide (NO) regulates production of vascular endothelial growth factor (VEGF) by normal and transformed cells. We demonstrate that NO donors may up-regulate the activity of the human VEGF promoter in normoxic human glioblastoma and hepatoma cells independent of a cyclic guanosine monophosphate-mediated pathway. Deletion and mutation analysis of the VEGF promoter indicates that the NO-responsive cis-elements are the hypoxia-inducible factor-1 (HIF-1) binding site and an adjacent ancillary sequence that is located immediately downstream within the hypoxia-response element (HRE). This work demonstrates that the HRE of this promoter is the primary target of NO. In addition, VEGF gene regulation by NO, as well as by hypoxia, is potentiated by the AP-1 element of the gene. Our study also reveals that NO and hypoxia induce an increase in HIF-1 binding activity and HIF-1alpha protein levels, both in the nucleus and the whole cell. These results suggest that there are common features of the NO and hypoxic pathways of VEGF induction, while in part, NO mediates gene transcription by a mechanism distinct from hypoxia. This is demonstrated by a difference in sensitivity to guanylate cyclase inhibitors and a different pattern of HIF-1 binding. These results show that there is a primary role for NO in the control of VEGF synthesis and in cell adaptations to hypoxia. (Blood. 2000;95:189-197)
Kimura
H
H
Investigative Treatment Division, National Cancer Center Research Institute East, Kashiwa, Chiba, Japan.
Weisz
A
A
Kurashima
Y
Y
Hashimoto
K
K
Ogura
T
T
D'Acquisto
F
F
Addeo
R
R
Makuuchi
M
M
Esumi
H
H
D001483:Base Sequence; D015687:Cell Hypoxia / Q000502:physiology*; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000378:metabolism*; D016228:Endothelial Growth Factors / Q000235:genetics*; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007700:Kinetics; D008156:Luciferases / Q000096:biosynthesis / Q000235:genetics; D008222:Lymphokines / Q000235:genetics*; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009569:Nitric Oxide / Q000502:physiology*; D020030:Nitric Oxide Donors / Q000494:pharmacology*; D009687:Nuclear Proteins / Q000378:metabolism*; D010396:Penicillamine / Q000031:analogs & derivatives* / Q000494:pharmacology; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D012045:Regulatory Sequences, Nucleic Acid; D026423:S-Nitroso-N-Acetylpenicillamine; D017384:Sequence Deletion; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
D004268:DNA-Binding Proteins; D016228:Endothelial Growth Factors; C497442:HIF1A protein, human; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D008222:Lymphokines; D020030:Nitric Oxide Donors; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D009569:Nitric Oxide; D026423:S-Nitroso-N-Acetylpenicillamine; D008156:Luciferases; D010396:Penicillamine