Intravesicular acidification correlates with binding of ADP-ribosylation factor to microsomal membranes.
The ADP-ribosylation factor (ARF), a highly conserved low molecular weight GTP-binding protein, has been implicated to function in intracellular protein transport to and within the Golgi complex. In pancreatic acinar cells the ARF is confined to the cytoplasmic faces of trans-Golgi stack membranes, a compartment known to maintain a low intravesicular pH, which is established by a chloride-dependent MgATP-driven proton pump. The present study shows that MgATP (2mM), but neither adenosine 5'-[gamma-thio]triphosphate in the presence of Mg2+ nor ATP in the absence of Mg2+, increases transfer of ARF from the surrounding medium into the vesicle membranes. The specific vacuolar-type proton pump inhibitor bafilomycin B1 (10 nM), the protonophore carbonylcyanide m-chlorophenylhydrazone (10 microM), and replacement of chloride in the incubation buffer by acetate or nitrate resulted in an almost complete inhibition of the MgATP-dependent association of ARF to the vesicle membranes. The results demonstrate that redistribution of ARF to the vesicle membrane correlates with the intravesicular pH established by a vacuolar-type H(+)-ATPase. The intravesicular pH appears to be one mechanism by which certain low molecular weight GTP-binding proteins become relocated from the cytosol to their specific membrane vesicles.
Zeuzem
S
S
Max-Planck-Institut für Biophysik, Frankfurt, Federal Republic of Germany.
Feick
P
P
Zimmermann
P
P
Haase
W
W
Kahn
R A
RA
Schulz
I
I
D020727:ADP-Ribosylation Factors; D000136:Acid-Base Equilibrium; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D002451:Cell Compartmentation; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D007425:Intracellular Membranes / Q000378:metabolism; D018942:Macrolides; D008854:Microscopy, Electron; D008861:Microsomes / Q000378:metabolism*; D010179:Pancreas / Q000648:ultrastructure; D011485:Protein Binding; D006180:Proton-Translocating ATPases / Q000378:metabolism*; D051381:Rats
D000900:Anti-Bacterial Agents; D018942:Macrolides; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; C052501:bafilomycin B1; D000255:Adenosine Triphosphate; D019204:GTP-Binding Proteins; D006180:Proton-Translocating ATPases; D020727:ADP-Ribosylation Factors