IF(1) distribution in HepG2 cells in relation to ecto-F(0)F (1)ATPsynthase and calmodulin.
F(0)F(1)ATPsynthase is now known to be expressed as a plasma membrane receptor for several extracellular ligands. On hepatocytes, ecto-F(0)F(1)ATPsynthase binds apoA-I and triggers HDL endocytosis concomitant with ATP hydrolysis. Considering that inhibitor protein IF(1) was shown to regulate the hydrolytic activity of ecto-F(0)F(1)ATPsynthase and to interact with calmodulin (CaM) in vitro, we investigated the subcellular distributions of IF(1), calmodulin (CaM), OSCP and beta subunits of F(0)F(1)ATPsynthase in HepG2 cells. Using immunofluorescence and Western blotting, we found that around 50% of total cellular IF(1) is localized outside mitochondria, a relevant amount of which is associated to the plasma membrane where we also found Ca(2+)-CaM, OSCP and beta. Confocal microscopy showed that IF(1) colocalized with Ca(2+)-CaM on plasma membrane but not in mitochondria, suggesting that Ca(2+)-CaM may modulate the cell surface availability of IF(1) and thus its ability to inhibit ATP hydrolysis by ecto-F(0)F(1)ATPsynthase. These observations support a hypothesis that the IF(1)-Ca(2+)-CaM complex, forming on plasma membrane, functions in the cellular regulation of HDL endocytosis by hepatocytes.
Contessi
Stefania
S
Department of Biomedical Sciences and Technologies, MATI Centre of Excellence, University of Udine, Piazzale Kolbe 4, 33100 Udine, Italy.
Comelli
Marina
M
Cmet
Sara
S
Lippe
Giovanna
G
Mavelli
Irene
I
D015153:Blotting, Western; D002147:Calmodulin / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D004705:Endocytosis; D022781:Hepatocytes / Q000378:metabolism; D006801:Humans; D008075:Lipoproteins, HDL / Q000378:metabolism; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000378:metabolism; D011506:Proteins / Q000378:metabolism*; D006180:Proton-Translocating ATPases / Q000378:metabolism*; D013347:Subcellular Fractions / Q000378:metabolism; D000097795:ATPase Inhibitory Protein
D002147:Calmodulin; D008075:Lipoproteins, HDL; D011506:Proteins; D006180:Proton-Translocating ATPases; D000097795:ATPase Inhibitory Protein