Regulation of hypoxia-inducible factor-1alpha by nitric oxide through mitochondria-dependent and -independent pathways

Biochem J. 2003 Dec 1;376(Pt 2):537-44. doi: 10.1042/BJ20031155.


Nitric oxide (NO) has been reported both to promote and to inhibit the activity of the transcription factor hypoxia-inducible factor-1 (HIF-1). In order to avoid the pitfalls associated with the use of NO donors, we have developed a human cell line (Tet-iNOS 293) that expresses the inducible NO synthase (iNOS) under the control of a tetracycline-inducible promoter. Using this system to generate finely controlled amounts of NO, we have demonstrated that the stability of the alpha-subunit of HIF-1 is regulated by NO through two separate mechanisms, only one of which is dependent on a functional respiratory chain. HIF-1alpha is unstable in cells maintained at 21% O(2), but is progressively stabilized as the O(2) concentration decreases, resulting in augmented HIF-1 DNA-binding activity. High concentrations of NO (>1 microM) stabilize HIF-1alpha at all O(2) concentrations tested. This effect does not involve the respiratory chain, since it is preserved in cells lacking functional mitochondria (rho(0)-cells) and is not reproduced by other inhibitors of the cytochrome c oxidase. By contrast, lower concentrations of NO (<400 nM) cause a rapid decrease in HIF-1alpha stabilized by exposure of the cells to 3% O(2). This effect of NO is dependent on the inhibition of mitochondrial respiration, since it is mimicked by other inhibitors of mitochondrial respiration, including those not acting at cytochrome c oxidase. We suggest that, although stabilization of HIF-1alpha by high concentrations of NO might have implications in pathophysiological processes, the inhibitory effect of lower NO concentrations is likely to be of physiological relevance.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Hypoxia
  • Cell Line
  • Cell Respiration
  • Humans
  • Hypoxia-Inducible Factor 1, alpha Subunit
  • Mitochondria / metabolism*
  • Mitochondria / physiology
  • Nitric Oxide / biosynthesis
  • Nitric Oxide / physiology*
  • Nitric Oxide Synthase / metabolism
  • Nitric Oxide Synthase Type II
  • Oxygen / pharmacology
  • Signal Transduction
  • Transcription Factors / metabolism*


  • HIF1A protein, human
  • Hypoxia-Inducible Factor 1, alpha Subunit
  • Transcription Factors
  • Nitric Oxide
  • NOS2 protein, human
  • Nitric Oxide Synthase
  • Nitric Oxide Synthase Type II
  • Oxygen