PI3K/Akt is required for heat shock proteins to protect hypoxia-inducible factor 1alpha from pVHL-independent degradation.
Hypoxia inducible factor 1 (HIF-1), a heterodimeric transcription factor composed of HIF-1alpha and HIF-1beta subunits, serves as a key regulator of metabolic adaptation to hypoxia. The amount of HIF-1alpha protein is regulated either by attenuating von Hippel-Lindau protein (pVHL)-dependent ubiquitination and subsequent 26 S proteasomal degradation or by enhancing cap-dependent ... mRNA translation, presumably involving a phosphatidyinositol 3-kinase (PI3K)/Akt-regulated pathway. In addition, it became apparent that Hsp90 protects HIF-1alpha from oxygen-independent degradation. Here we present evidence that PI3K/Akt is required for heat shock proteins to stabilize HIF-1alpha. In pVHL-deficient renal cell carcinoma cells, PI3K inhibition by LY294002 and wortmannin or transfection of either a dominant negative PI3K or a kinase-dead Akt mutant substantially lowered constitutively expressed HIF-1alpha without altering HIF-1alpha mRNA. Inhibitors of mitogen-activated protein kinase kinase (MAPKK) such as PD98059 or the p38 MAPK inhibitor SB203580 showed no interference. Considering that PI3K inhibitors down-regulated heat shock protein 90 (Hsp90) as well as Hsp70 expression and moreover attenuated heat- or hypoxia-induced Hsp70 as well as hypoxia-induced Hsp90 up-regulation we conclude that PI3K inhibition promoted degradation of HIF-1alpha indirectly by reducing steady state concentrations of Hsp90 and/or Hsp70. HIF-1alpha co-immunoprecipitated with Hsp90/Hsp70 and direct binding of Hsp70 to the oxygen-dependent degradation domain (ODD) of HIF-1alpha was proven by a pull-down assay and a peptide array. PI3K-mediated degradation of HIF-1alpha was confirmed in HEK 293 cells under hypoxia, suggesting that heat shock proteins constitute an integral component for HIF-1alpha accumulation. We conclude that PI3K/Akt contributes to HIF-1alpha stabilization by provoking expression of heat shock proteins.
Mesh Terms:
Amino Acid Sequence, Cell Hypoxia, Cell Line, Enzyme Inhibitors, Gene Expression, HSP70 Heat-Shock Proteins, HSP90 Heat-Shock Proteins, Humans, Hypoxia-Inducible Factor 1, alpha Subunit, Kidney, Molecular Sequence Data, Phosphatidylinositol 3-Kinases, Protein Structure, Tertiary, Protein-Serine-Threonine Kinases, Proto-Oncogene Proteins, Proto-Oncogene Proteins c-akt, Signal Transduction, Transcription Factors, Tumor Suppressor Proteins, Ubiquitin-Protein Ligases, Von Hippel-Lindau Tumor Suppressor Protein
Amino Acid Sequence, Cell Hypoxia, Cell Line, Enzyme Inhibitors, Gene Expression, HSP70 Heat-Shock Proteins, HSP90 Heat-Shock Proteins, Humans, Hypoxia-Inducible Factor 1, alpha Subunit, Kidney, Molecular Sequence Data, Phosphatidylinositol 3-Kinases, Protein Structure, Tertiary, Protein-Serine-Threonine Kinases, Proto-Oncogene Proteins, Proto-Oncogene Proteins c-akt, Signal Transduction, Transcription Factors, Tumor Suppressor Proteins, Ubiquitin-Protein Ligases, Von Hippel-Lindau Tumor Suppressor Protein
J. Biol. Chem.
Date: Apr. 02, 2004
PubMed ID: 14726529
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