title: Extended ischemia prevents HIF1alpha degradation at reoxygenation by impairing prolyl-hydroxylation: role of Krebs cycle metabolites.
abstract: Hypoxia-inducible factor (HIF) is a heterodimeric transcription factor that activates the cellular response to hypoxia. The HIF1alpha subunit is constantly synthesized and degraded under normoxia, but degradation is rapidly inhibited when oxygen levels drop. Oxygen-dependent hydroxylation by prolyl-4-hydroxylases (PHD) mediates HIF1alpha proteasome degradation. Brain ischemia limits the availability not only of oxygen but also of glucose. We hypothesized that this circumstance could have a modulating effect on HIF. We assessed the separate involvement of oxygen and glucose in HIF1alpha regulation in differentiated neuroblastoma cells subjected to ischemia. We report higher transcriptional activity and HIF1alpha expression under oxygen deprivation in the presence of glucose (OD), than in its absence (oxygen and glucose deprivation, OGD). Unexpectedly, HIF1alpha was not degraded at reoxygenation after an episode of OGD. This was not due to impairment of proteasome function, but was associated with lower HIF1alpha hydroxylation. Krebs cycle metabolites fumarate and succinate are known inhibitors of PHD, while alpha-ketoglutarate is a co-substrate of the reaction. Lack of HIF1alpha degradation in the presence of oxygen was accompanied by a very low alpha-ketoglutarate/fumarate ratio. Furthermore, treatment with a fumarate analogue prevented HIF1alpha degradation under normoxia. In all, our data suggest that postischemic metabolic alterations in Krebs cycle metabolites impair HIF1alpha degradation in the presence of oxygen by decreasing its hydroxylation, and highlight the involvement of metabolic pathways in HIF1alpha regulation besides the well known effects of oxygen.
authors.affiliation: Department of Brain Ischemia and Neurodegeneration, Institute of Biomedical Research of Barcelona, Consejo Superior de Investigaciones Científicas, Institut d'Investigacions Biomèdiques August Pi i Sunyer, 08036 Barcelona, Spain.
authors.forename: Anna
authors.initials: A
authors.lastname: Serra-Pérez
authors.forename: Anna M
authors.initials: AM
authors.lastname: Planas
authors.forename: Analía
authors.initials: A
authors.lastname: Núñez-O'Mara
authors.forename: Edurne
authors.initials: E
authors.lastname: Berra
authors.forename: Judit
authors.initials: J
authors.lastname: García-Villoria
authors.forename: Antònia
authors.initials: A
authors.lastname: Ribes
authors.forename: Tomàs
authors.initials: T
authors.lastname: Santalucía
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D002952:Citric Acid Cycle; D005650:Fumarates / Q000737:chemistry; D005786:Gene Expression Regulation; D005947:Glucose / Q000378:metabolism; D006801:Humans; D006900:Hydroxylation; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism*; D051379:Mice; D008954:Models, Biological; D010100:Oxygen / Q000737:chemistry / Q000378:metabolism*; D011393:Procollagen-Proline Dioxygenase / Q000378:metabolism*; D011392:Proline / Q000737:chemistry*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D019802:Succinic Acid / Q000737:chemistry
chemical_list: D005650:Fumarates; C497442:HIF1A protein, human; C497443:Hif1a protein, mouse; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D011392:Proline; D019802:Succinic Acid; D011393:Procollagen-Proline Dioxygenase; D046988:Proteasome Endopeptidase Complex; D005947:Glucose; D010100:Oxygen