title: Ca2+/calmodulin kinase-dependent activation of hypoxia inducible factor 1 transcriptional activity in cells subjected to intermittent hypoxia.
abstract: Intermittent hypoxia (IH) occurs in many pathological conditions. However, very little is known about the molecular mechanisms associated with IH. Hypoxia-inducible factor 1 (HIF-1) mediates transcriptional responses to continuous hypoxia. In the present study, we investigated whether IH activates HIF-1 and, if so, which signaling pathways are involved. PC12 cells were exposed to either to 20% O2 (non-hypoxic control) or to 60 cycles consisting of 30 s at 1.5% O2, followed by 4 min at 20% O2 (IH). Western blot analysis revealed significant increases in HIF-1alpha protein in nuclear extracts of cells subjected to IH. Expression of a HIF-1-dependent reporter gene was increased 3-fold in cells subjected to IH. Although IH induced the activation of ERK1, ERK2, JNK, PKC-alpha, and PKC-gamma, inhibitors of these kinases and of phosphatidylinositol 3-kinase did not block HIF-1-mediated reporter gene expression induced by IH, indicating that signaling via these kinases was not required. In contrast, addition of the intracellular Ca2+ chelator BAPTA-AM or the Ca2+/calmodulin-dependent (CaM) kinase inhibitor KN93 blocked reporter gene activation in response to IH. CaM kinase activity was increased 5-fold in cells subjected to IH. KN 93 prevented IH-induced transactivation mediated by HIF-1alpha, and its coactivator p300, which was phosphorylated by CaM kinase II in vitro. Expression of the HIF-1-regulated gene encoding tyrosine hydroxylase was induced by IH and this effect was blocked by KN93. These observations suggest that IH induces HIF-1 transcriptional activity via a novel signaling pathway involving CaM kinase.
authors.affiliation: Department of Physiology & Biophysics, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44106, USA.
authors.forename: Guoxiang
authors.initials: G
authors.lastname: Yuan
authors.forename: Jayasri
authors.initials: J
authors.lastname: Nanduri
authors.forename: C Raman
authors.initials: CR
authors.lastname: Bhasker
authors.forename: Gregg L
authors.initials: GL
authors.lastname: Semenza
authors.forename: Nanduri R
authors.initials: NR
authors.lastname: Prabhakar
mesh_terms: D000818:Animals; D001216:Asparagine / Q000737:chemistry; D001596:Benzylamines / Q000494:pharmacology; D015152:Blotting, Northern; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D050881:E1A-Associated p300 Protein; D004789:Enzyme Activation; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D005982:Glutathione Transferase / Q000378:metabolism; D000860:Hypoxia; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D015151:Immunoblotting; D020935:MAP Kinase Signaling System; D009687:Nuclear Proteins / Q000378:metabolism; D009841:Oligonucleotides / Q000737:chemistry; D010100:Oxygen / Q000378:metabolism; D016716:PC12 Cells; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D015398:Signal Transduction; D013449:Sulfonamides / Q000494:pharmacology; D013997:Time Factors; D015534:Trans-Activators / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism; D000096926:Benzenesulfonamides
chemical_list: D001216:Asparagine; D001596:Benzylamines; D002118:Calcium; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D004247:DNA; D050881:E1A-Associated p300 Protein; D005982:Glutathione Transferase; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D009687:Nuclear Proteins; D009841:Oligonucleotides; D010100:Oxygen; D019869:Phosphatidylinositol 3-Kinases; D011493:Protein Kinase C; D012333:RNA, Messenger; D013449:Sulfonamides; D015534:Trans-Activators; D014157:Transcription Factors; D014446:Tyrosine 3-Monooxygenase; C072105:KN 93; D000096926:Benzenesulfonamides; C494362:Ep300 protein, rat