Structural and functional characterization of F(o)F(1)-ATP synthase on the extracellular surface of rat hepatocytes.
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-F(o)F(1)-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 F(1) (alpha/beta and gamma) and F(o) (F(o)I-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 IF(1) 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 F(1) and F(o)-targeting inhibitors. We hypothesise that ecto-F(o)F(1)-ATP synthase may control the extracellular ADP/ATP ratio, thus contributing to intracellular pH homeostasis.
Mangiullo
Roberto
R
Department of Medical Biochemistry, Biology and Physics, University of Bari, Italy.
Gnoni
Antonio
A
Leone
Antonella
A
Gnoni
Gabriele V
GV
Papa
Sergio
S
Zanotti
Franco
F
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002462:Cell Membrane / Q000737:chemistry / Q000201:enzymology; D002478:Cells, Cultured; D005110:Extracellular Space / Q000378:metabolism*; D022781:Hepatocytes / Q000166:cytology / Q000201:enzymology*; D008297:Male; D053078:Membrane Potential, Mitochondrial / Q000502:physiology; D025261:Mitochondrial Proton-Translocating ATPases / Q000737:chemistry / Q000378:metabolism; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar
D021122:Protein Subunits; D000255:Adenosine Triphosphate; D025261:Mitochondrial Proton-Translocating ATPases