Extracellular ATP formation from ADP and inorganic phosphate, attributed to the activity of a cell surface ATP synthase, has so far only been reported in cultures of some proliferating and tumoral cell lines. We now provide evidence showing the presence of a functionally active ecto-FoF1-ATP synthase on the plasma membrane of normal tissue cells, i.e. isolated rat hepatocytes. Both confocal microscopy and flow cytometry analysis show the presence of subunits of F1 (α/β and γ) and Fo (FoI-PVP(b) and OSCP) moieties of ATP synthase at the surface of rat hepatocytes. This finding is confirmed by immunoblotting analysis of the hepatocyte plasma membrane fraction. The presence of the inhibitor protein IF1 is also detected on the hepatocyte surface. Activity assays show that the ectopic-ATP synthase can work both in the direction of ATP synthesis and hydrolysis. A proton translocation assay shows that both these mechanisms are accompanied by a transient flux of H+ and are inhibited by F1 and Fo-targeting inhibitors. We hypothesise that ecto-FoF1-ATP synthase may control the extracellular ADP/ATP ratio, thus contributing to intracellular pH homeostasis. © 2008 Elsevier B.V. All rights reserved.

Structural and functional characterization of FoF1-ATP synthase on the extracellular surface of rat hepatocytes

Gnoni A.;Gnoni G. V.;Zanotti F.
2008

Abstract

Extracellular ATP formation from ADP and inorganic phosphate, attributed to the activity of a cell surface ATP synthase, has so far only been reported in cultures of some proliferating and tumoral cell lines. We now provide evidence showing the presence of a functionally active ecto-FoF1-ATP synthase on the plasma membrane of normal tissue cells, i.e. isolated rat hepatocytes. Both confocal microscopy and flow cytometry analysis show the presence of subunits of F1 (α/β and γ) and Fo (FoI-PVP(b) and OSCP) moieties of ATP synthase at the surface of rat hepatocytes. This finding is confirmed by immunoblotting analysis of the hepatocyte plasma membrane fraction. The presence of the inhibitor protein IF1 is also detected on the hepatocyte surface. Activity assays show that the ectopic-ATP synthase can work both in the direction of ATP synthesis and hydrolysis. A proton translocation assay shows that both these mechanisms are accompanied by a transient flux of H+ and are inhibited by F1 and Fo-targeting inhibitors. We hypothesise that ecto-FoF1-ATP synthase may control the extracellular ADP/ATP ratio, thus contributing to intracellular pH homeostasis. © 2008 Elsevier B.V. All rights reserved.
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/11586/238639
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