Hormone-induced assembly and activation of V-ATPase in blowfly salivary glands is mediated by protein kinase A

  • The vacuolar H+-ATPase (V-ATPase) in the apical membrane of blowfly (Calliphora vicina) salivary gland cells energizes the secretion of a KCl-rich saliva in response to the neurohormone serotonin (5-HT). We have shown previously that exposure to 5-HT induces a cAMP-mediated reversible assembly of V-0 and V-1 subcomplexes to V-ATPase holoenzymes and increases V-ATPase-driven proton transport. Here, we analyze whether the effect of cAMP on V-ATPase is mediated by protein kinase A (PKA) or exchange protein directly activated by cAMP (Epac), the cAMP target proteins that are present within the salivary glands. Immunofluorescence microscopy shows that PKA activators, but not Epac activators, induce the translocation of V1 components from the cytoplasm to the apical membrane, indicative of an assembly of V-ATPase holoenzymes. Measurements of transepithelial voltage changes and microfluorometric pH measurements at the luminal surface of cells in isolated glands demonstrate further that PKA-activating cAMP analogs increase cation transport toThe vacuolar H+-ATPase (V-ATPase) in the apical membrane of blowfly (Calliphora vicina) salivary gland cells energizes the secretion of a KCl-rich saliva in response to the neurohormone serotonin (5-HT). We have shown previously that exposure to 5-HT induces a cAMP-mediated reversible assembly of V-0 and V-1 subcomplexes to V-ATPase holoenzymes and increases V-ATPase-driven proton transport. Here, we analyze whether the effect of cAMP on V-ATPase is mediated by protein kinase A (PKA) or exchange protein directly activated by cAMP (Epac), the cAMP target proteins that are present within the salivary glands. Immunofluorescence microscopy shows that PKA activators, but not Epac activators, induce the translocation of V1 components from the cytoplasm to the apical membrane, indicative of an assembly of V-ATPase holoenzymes. Measurements of transepithelial voltage changes and microfluorometric pH measurements at the luminal surface of cells in isolated glands demonstrate further that PKA-activating cAMP analogs increase cation transport to the gland lumen and induce a V-ATPase-dependent luminal acidification, whereas activators of Epac do not. Inhibitors of PKA block the 5-HT-induced V-1 translocation to the apical membrane and the increase in proton transport. We conclude that cAMP exerts its effects on V-ATPase via PKA.show moreshow less

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Metadaten
Author:Julia Rein, Martin Voss, Wolfgang Blenau, Bernd Walz, Otto Baumann
URN:urn:nbn:de:kobv:517-opus-46126
Document Type:Postprint
Language:English
Date of Publication (online):2010/08/27
Year of Completion:2008
Publishing Institution:Universität Potsdam
Release Date:2010/08/27
Tag:Vacuolar h+-atpase; camp binding-sites; cyclic-amp; drosophila-melanogaster; plasma-membrane
Source:American journal of physiology : cell physiology. - 294 (2008), 1, S. C56 - C65, DOI: 10.1152/ajpcell.00041.2007
Organizational units:Mathematisch-Naturwissenschaftliche Fakultät / Institut für Biochemie und Biologie
Dewey Decimal Classification:5 Naturwissenschaften und Mathematik / 57 Biowissenschaften; Biologie / 570 Biowissenschaften; Biologie
Licence (German):License LogoKeine Nutzungslizenz vergeben - es gilt das deutsche Urheberrecht
Notes extern:
The article was originally published by the American Physiological Society:
American Journal of Physiology : Cell Physiology. - 294 (2008), 1, S. C56-C65
ISSN 0363-6143
DOI 10.1152/ajpcell.00041.2007