title: Intracellular signaling for vasoconstrictor coupling factor 6: novel function of beta-subunit of ATP synthase as receptor.
abstract: Coupling factor 6 (CF6), a component of adenosine triphosphate (ATP) synthase, is circulating and functions as an endogenous vasoconstrictor by inhibiting cytosolic phospholipase A2. We showed a high plasma level of CF6 in human hypertension. The present study focused on the identification and characterization of a receptor for CF6 and its post-receptor signaling pathway. Incubation of human umbilical vein endothelial cells (HUVECs) with an excess of free CF6 reduced by 50% the immunoreactivity for the antibody to beta-subunit of ATP synthase at the cell surface, but unaffected that for the alpha-subunit antibody. A significant displacement of radioligand was observed at 3x10(-9) through 10(-7) M unlabeled CF6, and the Kd was 7.6 nM. Adenosine diphosphate (ADP) at 10(-7) M and beta-subunit antibody suppressed the binding of (125)I-CF6 by 81.3+/-9.7% and 32.0+/-2.0%, respectively, whereas the alpha-subunit antibody unaffected it. The hydrolysis activity of ATP to ADP was increased by 1.6-fold by CF6 at 10(-7) M, and efrapeptin at 10(-5) M, an inhibitor of ATP synthase, blocked it. CF6 at 10(-7) M decreased intracellular pH in 2',7'-bis(carboxyethyl-5 (6))-carboxyfluorescein-loaded HUVEC. Amyloride at 10(-4) M augmented the pH decrease in response to CF6, whereas efrapeptin at 10(-5) M blocked it. Arachidonic acid release was suppressed by CF6, and it was reversed by efrapeptin at 10(-5) M or beta-subunit antibody or ADP at 10(-7) M. The beta-subunit antibody suppressed coupling factor 6-induced increase in blood pressure. These indicate that membrane-bound ATP synthase functions as a receptor for CF6 and may have a previously unsuspected role in the genesis of hypertension by modulating the concentration of intracellular hydrogen.
authors.affiliation: The Second Department of Internal Medicine, Hirosaki University School of Medicine, Hirosaki, 036-8562, Japan. osanait@cc.hirosaki-u.ac.jp
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Osanai
authors.forename: Koji
authors.initials: K
authors.lastname: Magota
authors.forename: Makoto
authors.initials: M
authors.lastname: Tanaka
authors.forename: Michiko
authors.initials: M
authors.lastname: Shimada
authors.forename: Reiichi
authors.initials: R
authors.lastname: Murakami
authors.forename: Satoko
authors.initials: S
authors.lastname: Sasaki
authors.forename: Hirofumi
authors.initials: H
authors.lastname: Tomita
authors.forename: Naotaka
authors.initials: N
authors.lastname: Maeda
authors.forename: Ken
authors.initials: K
authors.lastname: Okumura
mesh_terms: D016718:Arachidonic Acid / Q000378:metabolism; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D006801:Humans; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D047468:Immunoprecipitation; D007425:Intracellular Membranes / Q000378:metabolism*; D007527:Isoenzymes / Q000378:metabolism; D025261:Mitochondrial Proton-Translocating ATPases / Q000378:metabolism* / Q000494:pharmacology; D010086:Oxidative Phosphorylation Coupling Factors / Q000378:metabolism* / Q000494:pharmacology; D011956:Receptors, Cell Surface / Q000378:metabolism*; D015398:Signal Transduction; D014471:Umbilical Veins; D014661:Vasoconstriction / Q000502:physiology*
chemical_list: C506526:ATP5F1B protein, human; D007527:Isoenzymes; D010086:Oxidative Phosphorylation Coupling Factors; D011956:Receptors, Cell Surface; D016718:Arachidonic Acid; C000628217:F(6) ATPase; D025261:Mitochondrial Proton-Translocating ATPases