title: Induction of HIF-1alpha expression by intermittent hypoxia: involvement of NADPH oxidase, Ca2+ signaling, prolyl hydroxylases, and mTOR.
abstract: Sleep-disordered breathing with recurrent apnea (periodic cessation of breathing) results in chronic intermittent hypoxia (IH), which leads to cardiovascular and respiratory pathology. Molecular mechanisms underlying IH-evoked cardio-respiratory co-morbidities have not been delineated. Mice with heterozygous deficiency of hypoxia-inducible factor 1alpha (HIF-1alpha) do not develop cardio-respiratory responses to chronic IH. HIF-1alpha protein expression and HIF-1 transcriptional activity are induced by IH in PC12 cells. In the present study, we investigated the signaling pathways associated with IH-evoked HIF-1alpha accumulation. PC12 cells were exposed to aerobic conditions (20% O(2)) or 60 cycles of IH (30 sec at 1.5% O(2) followed by 5 min at 20% O(2)). Our results show that IH-induced HIF-1alpha accumulation is due to increased generation of ROS by NADPH oxidase. We further demonstrate that ROS-dependent Ca(2+) signaling pathways involving phospholipase Cgamma (PLCgamma) and protein kinase C activation are required for IH-evoked HIF-1alpha accumulation. IH leads to activation of mTOR and S6 kinase (S6K) and rapamycin partially inhibited IH-induced HIF-1alpha accumulation. IH also decreased hydroxylation of HIF-1alpha protein and anti-oxidants as well as inhibitors of Ca(+2) signaling prevented this response. Thus, both increased mTOR-dependent HIF-1alpha synthesis and decreased hydroxylase-dependent HIF-1alpha degradation contribute to IH-evoked HIF-1alpha accumulation. Following IH, HIF-1alpha, and phosphorylated mTOR levels remained elevated during 90 min of re-oxygenation despite re-activation of prolyl hydroxylase. Rapamycin or cycloheximide, blocked increased HIF-1alpha levels during re-oxygenation indicating that mTOR-dependent protein synthesis is required for the persistent elevation of HIF-1alpha levels during re-oxygenation.
authors.affiliation: The Center for Systems Biology, Department of Medicine, University of Chicago, Illinois 60637, USA.
authors.forename: Guoxiang
authors.initials: G
authors.lastname: Yuan
authors.forename: Jayasri
authors.initials: J
authors.lastname: Nanduri
authors.forename: Shakil
authors.initials: S
authors.lastname: Khan
authors.forename: Gregg L
authors.initials: GL
authors.lastname: Semenza
authors.forename: Nanduri R
authors.initials: NR
authors.lastname: Prabhakar
mesh_terms: D000818:Animals; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects; D015687:Cell Hypoxia / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D006900:Hydroxylation / Q000187:drug effects; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism*; D051379:Mice; D008954:Models, Biological; D019255:NADPH Oxidases / Q000378:metabolism*; D010100:Oxygen / Q000494:pharmacology; D016716:PC12 Cells; D051966:Phospholipase C gamma / Q000378:metabolism; D011393:Procollagen-Proline Dioxygenase / Q000378:metabolism*; D014176:Protein Biosynthesis / Q000187:drug effects; D011493:Protein Kinase C / Q000378:metabolism; D011494:Protein Kinases / Q000378:metabolism*; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism; D019893:Ribosomal Protein S6 Kinases / Q000378:metabolism; D020123:Sirolimus / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases; D013816:Thermodynamics
chemical_list: D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D017382:Reactive Oxygen Species; D011393:Procollagen-Proline Dioxygenase; D019255:NADPH Oxidases; D011494:Protein Kinases; C546843:mTOR protein, mouse; D019893:Ribosomal Protein S6 Kinases; D058570:TOR Serine-Threonine Kinases; D011493:Protein Kinase C; D051966:Phospholipase C gamma; D010100:Oxygen; D002118:Calcium; D020123:Sirolimus