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Hypoxia-inducible factor 2alpha binds to cobalt in vitro
Y Yuan1, D Beitner-Johnson, D E Millhorn
1Department of Molecular Physiology, Genome Research Institute, University of Cincinnati, 45267-0576, USA.
Biochemical and Biophysical Research Communications
|November 2, 2001
Summary
Cobalt mimics hypoxia by binding to HIF-2alpha, stabilizing this transcription factor. This interaction, dependent on oxygen levels and specific protein domains, reveals potential roles for transition metals in regulating hypoxia-inducible factors.
Area of Science:
- Biochemistry
- Molecular Biology
- Cellular Signaling
Background:
- Hypoxia-inducible factor (HIF) regulates gene expression via hypoxia response elements (HRE).
- HIF-alpha subunits are degraded under normoxia but stabilized during hypoxia.
- Cobalt is known to induce HIF-alpha accumulation, mimicking hypoxic conditions.
Purpose of the Study:
- To elucidate the mechanism by which cobalt stabilizes HIF-2alpha.
- To investigate the direct interaction between cobalt and HIF-2alpha.
- To identify the role of specific HIF-2alpha domains in cobalt-mediated stabilization.
Main Methods:
- In vitro binding assays to assess cobalt-HIF-2alpha interaction.
- Use of proteasome inhibitors and hypoxic conditions to stabilize HIF-2alpha.
- Site-directed mutagenesis of the oxygen-dependent degradation domain (ODDD) of HIF-2alpha.
Main Results:
- Cobalt directly binds to HIF-2alpha in vitro with high affinity and oxygen-dependently.
- Cobalt binding to HIF-2alpha is prevented when HIF-2alpha is stabilized by hypoxia.
- Mutations in the ODDD of HIF-2alpha abolish cobalt binding and cause HIF-2alpha accumulation under normoxia.
Conclusions:
- Cobalt mimics hypoxia by directly interacting with HIF-2alpha, independent of the canonical hypoxic pathway.
- The oxygen-dependent degradation domain of HIF-2alpha is critical for cobalt binding.
- Transition metals, potentially iron, may regulate HIF-2alpha stability in vivo.