Molecular mechanisms of transcription activation by HLF and HIF1alpha in response to hypoxia: their stabilization and redox signal-induced interaction with CBP/p300.
Hypoxia-inducible factor 1 alpha (HIF1alpha) and its related factor, HLF, activate expression of a group of genes such as erythropoietin in response to low oxygen. Transfection analysis using fusion genes of GAL4DBD with various fragments of the two factors delineated two transcription activation domains which are inducible in response to hypoxia and are localized in the C-terminal half. Their sequences are conserved between HLF and HIF1alpha. One is designated NAD (N-terminal activation domain), while the other is CAD (C-terminal activation domain). Immunoblot analysis revealed that NADs, which were rarely detectable at normoxia, became stabilized and accumulated at hypoxia, whereas CADs were constitutively expressed. In the mammalian two-hybrid system, CAD and NAD baits enhanced the luciferase expression from a reporter gene by co-transfection with CREB-binding protein (CBP) prey, whereas CAD, but not NAD, enhanced beta-galactosidase expression in yeast by CBP co-expression, suggesting that NAD and CAD interact with CBP/p300 by a different mechanism. Co-transfection experiments revealed that expression of Ref-1 and thioredoxin further enhanced the luciferase activity expressed by CAD, but not by NAD. Amino acid replacement in the sequences of CADs revealed a specific cysteine to be essential for their hypoxia-inducible interaction with CBP. Nuclear translocation of thioredoxin from cytoplasm was observed upon reducing O2 concentrations.
Ema
M
M
Department of Chemistry, Graduate School of Science, Tohoku University, Sendai 980-77, Japan.
Hirota
K
K
Mimura
J
J
Abe
H
H
Yodoi
J
J
Sogawa
K
K
Poellinger
L
L
Fujii-Kuriyama
Y
Y
D000595:Amino Acid Sequence; D001483:Base Sequence; D051792:Basic Helix-Loop-Helix Proteins; D019757:Carbon-Oxygen Lyases / Q000378:metabolism; D015687:Cell Hypoxia / Q000235:genetics* / Q000502:physiology*; D002460:Cell Line; D017931:DNA Primers / Q000235:genetics; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004355:Drug Stability; D006367:HeLa Cells; D006801:Humans; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D008954:Models, Biological; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D010084:Oxidation-Reduction; D015398:Signal Transduction; D013879:Thioredoxins / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors; D015533:Transcriptional Activation; D014162:Transfection; D000099162:Endothelial PAS Domain-Containing Protein 1
D051792:Basic Helix-Loop-Helix Proteins; D019757:Carbon-Oxygen Lyases; D017931:DNA Primers; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D004268:DNA-Binding Proteins; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D009687:Nuclear Proteins; D013879:Thioredoxins; D015534:Trans-Activators; D014157:Transcription Factors; D000099162:Endothelial PAS Domain-Containing Protein 1; C497442:HIF1A protein, human; C085108:APEX1 protein, human