title: Important role of PLC-γ1 in hypoxic increase in intracellular calcium in pulmonary arterial smooth muscle cells.
abstract: An increase in intracellular calcium concentration ([Ca(2+)](i)) in pulmonary arterial smooth muscle cells (PASMCs) induces hypoxic cellular responses in the lungs; however, the underlying molecular mechanisms remain incompletely understood. We report, for the first time, that acute hypoxia significantly enhances phospholipase C (PLC) activity in mouse resistance pulmonary arteries (PAs), but not in mesenteric arteries. Western blot analysis and immunofluorescence staining reveal the expression of PLC-γ1 protein in PAs and PASMCs, respectively. The activity of PLC-γ1 is also augmented in PASMCs following hypoxia. Lentiviral shRNA-mediated gene knockdown of mitochondrial complex III Rieske iron-sulfur protein (RISP) to inhibit reactive oxygen species (ROS) production prevents hypoxia from increasing PLC-γ1 activity in PASMCs. Myxothiazol, a mitochondrial complex III inhibitor, reduces the hypoxic response as well. The PLC inhibitor U73122, but not its inactive analog U73433, attenuates the hypoxic vasoconstriction in PAs and hypoxic increase in [Ca(2+)](i) in PASMCs. PLC-γ1 knockdown suppresses its protein expression and the hypoxic increase in [Ca(2+)](i). Hypoxia remarkably increases inositol 1,4,5-trisphosphate (IP(3)) production, which is blocked by U73122. The IP(3) receptor (IP(3)R) antagonist 2-aminoethoxydiphenyl borate (2-APB) or xestospongin-C inhibits the hypoxic increase in [Ca(2+)](i). PLC-γ1 knockdown or U73122 reduces H(2)O(2)-induced increase in [Ca(2+)](i) in PASMCs and contraction in PAs. 2-APB and xestospongin-C produce similar inhibitory effects. In conclusion, our findings provide novel evidence that hypoxia activates PLC-γ1 by increasing RISP-dependent mitochondrial ROS production in the complex III, which causes IP(3) production, IP(3)R opening, and Ca(2+) release, playing an important role in hypoxic Ca(2+) and contractile responses in PASMCs.
authors.affiliation: Center for Cardiovascular Sciences, Albany Medical College, Albany, NY 12208, USA. WangY@mail.amc.edu
authors.forename: Vishal R
authors.initials: VR
authors.lastname: Yadav
authors.forename: Tengyao
authors.initials: T
authors.lastname: Song
authors.forename: Leroy
authors.initials: L
authors.lastname: Joseph
authors.forename: Lin
authors.initials: L
authors.lastname: Mei
authors.forename: Yun-Min
authors.initials: YM
authors.lastname: Zheng
authors.forename: Yong-Xiao
authors.initials: YX
authors.lastname: Wang
mesh_terms: D000818:Animals; D001896:Boron Compounds / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D014450:Electron Transport Complex III / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D004963:Estrenes / Q000494:pharmacology; D015870:Gene Expression; D000860:Hypoxia / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D015544:Inositol 1,4,5-Trisphosphate / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D053496:Inositol 1,4,5-Trisphosphate Receptors / Q000037:antagonists & inhibitors / Q000378:metabolism; D016086:Lentivirus / Q000235:genetics; D047028:Macrocyclic Compounds / Q000494:pharmacology; D008689:Methacrylates / Q000494:pharmacology; D051379:Mice; D009119:Muscle Contraction / Q000187:drug effects; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D010080:Oxazoles / Q000494:pharmacology; D051966:Phospholipase C gamma / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D011651:Pulmonary Artery / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology; D011760:Pyrrolidinones / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000235:genetics; D017382:Reactive Oxygen Species / Q000819:agonists / Q000037:antagonists & inhibitors / Q000378:metabolism; D013844:Thiazoles / Q000494:pharmacology; D014661:Vasoconstriction / Q000187:drug effects
chemical_list: D001896:Boron Compounds; D004963:Estrenes; D053496:Inositol 1,4,5-Trisphosphate Receptors; D047028:Macrocyclic Compounds; D008689:Methacrylates; D010080:Oxazoles; D011760:Pyrrolidinones; D034741:RNA, Small Interfering; D017382:Reactive Oxygen Species; C037989:Rieske iron-sulfur protein; D013844:Thiazoles; C511704:xestospongin C; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; C030517:myxothiazol; D015544:Inositol 1,4,5-Trisphosphate; C109986:2-aminoethoxydiphenyl borate; D051966:Phospholipase C gamma; D014450:Electron Transport Complex III; D002118:Calcium