title: Inducible HSP70 regulates superoxide dismutase-2 and mitochondrial oxidative stress in the endothelial cells from developing lungs.
abstract: Superoxide dismutase 2 (SOD-2) is synthesized in the cytosol and imported into the mitochondrial matrix, where it is activated and functions as the primary antioxidant for cellular respiration. The specific mechanisms that target SOD-2 to the mitochondria remain unclear. We hypothesize that inducible heat shock protein 70 (iHSP70) targets SOD-2 to the mitochondria via a mechanism facilitated by ATP, and this process is impaired in persistent pulmonary hypertension of the newborn (PPHN). We observed that iHSP70 interacts with SOD-2 and targets SOD-2 to the mitochondria. Interruption of iHSP70-SOD-2 interaction with 2-phenylethylenesulfonamide-μ (PFT-μ, a specific inhibitor of substrate binding to iHSP70 COOH terminus) and siRNA-mediated knockdown of iHSP70 expression disrupted SOD-2 transport to mitochondria. Increasing intracellular ATP levels by stimulation of respiration with CaCl2 facilitated the mitochondrial import of SOD-2, increased SOD-2 activity, and decreased the mitochondrial superoxide (O2(·-)) levels in PPHN pulmonary artery endothelial cells (PAEC) by promoting iHSP70-SOD-2 dissociation at the outer mitochondrial membrane. In contrast, oligomycin, an inhibitor of mitochondrial ATPase, decreased SOD-2 expression and activity and increased O2(·-) levels in the mitochondria of control PAEC. The basal ATP levels and degree of iHSP70-SOD-2 dissociation were lower in PPHN PAEC and lead to increased SOD-2 degradation in cytosol. In normal pulmonary arteries (PA), PFT-μ impaired the relaxation response of PA rings in response to nitric oxide (NO) donor, S-nitroso-N-acetyl-penicillamine. Pretreatment with Mito-Q, a mitochondrial targeted O2(·-) scavenger, restored the relaxation response in PA rings pretreated with PFT-μ. Our observations suggest that iHSP70 chaperones SOD-2 to the mitochondria. Impaired SOD-2-iHSP70 dissociation decreases SOD-2 import and contributes to mitochondrial oxidative stress in PPHN.
authors.affiliation: Div. of Neonatology, Dept. of Pediatrics, Suite C460, Children's Corporate Ctr., 999 N. 92nd St., Wauwatosa, WI. aafolaya@mcw.edu.
authors.forename: Adeleye J
authors.initials: AJ
authors.lastname: Afolayan
authors.forename: Ru-Jeng
authors.initials: RJ
authors.lastname: Teng
authors.forename: Annie
authors.initials: A
authors.lastname: Eis
authors.forename: Ujala
authors.initials: U
authors.lastname: Rana
authors.forename: Katarzyna A
authors.initials: KA
authors.lastname: Broniowska
authors.forename: John A
authors.initials: JA
authors.lastname: Corbett
authors.forename: Kirkwood
authors.initials: K
authors.lastname: Pritchard
authors.forename: Girija G
authors.initials: GG
authors.lastname: Konduri
mesh_terms: D000255:Adenosine Triphosphate / Q000096:biosynthesis / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000201:enzymology*; D018840:HSP70 Heat-Shock Proteins / Q000502:physiology*; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism; D007231:Infant, Newborn; D008168:Lung / Q000473:pathology; D008928:Mitochondria / Q000201:enzymology*; D010085:Oxidative Phosphorylation; D018384:Oxidative Stress; D010547:Persistent Fetal Circulation Syndrome / Q000201:enzymology* / Q000473:pathology; D021381:Protein Transport; D059748:Proteolysis; D011651:Pulmonary Artery / Q000473:pathology; D012756:Sheep
chemical_list: D018840:HSP70 Heat-Shock Proteins; D000255:Adenosine Triphosphate; D006861:Hydrogen Peroxide
keywords: nitric oxide; oxidative stress; persistent pulmonary hypertension of the newborn; pulmonary artery endothelium; vasodilation