Protein kinase C activation accelerates proton extrusion by vacuolar-type H(+)-ATPases in murine peritoneal macrophages.
The role of protein kinase C in the regulation of vacuolar-type H(+)-ATPase (V-ATPase) activity was studied in thioglycolate-elicited mouse peritoneal macrophages. Acid-loaded macrophages suspended in a Na(+)- and HCO(3-)-free K(+)-medium containing Zn2+, a H(+)-conductance blocker, exhibited an initial intracellular pH recovery rate of 0.33 +/- 0.04 pH/min (n = 9). Pretreatment with 12-O-tetradecanoyl phorbol 13-acetate (TPA) or mezerein for as little as 3 min induced a marked (82%) increase in the initial pH recovery rate. Stimulation was prevented by the V-ATPase inhibitor, bafilomycin A1 (200 nM) indicating that the effect of the protein kinase C agonist was via augmentation of proton pump activity. The protein kinase C inhibitor, staurosporine (100 nM) completely blocked the stimulatory effects of TPA and mezerein, suggesting involvement of protein kinase C. In keeping with this notion, the inactive analogue of TPA, 4-phorbol didecanoate did not stimulate recovery from an acid load. Extracellular pH determinations revealed that the observed increase in cytosolic pH recovery rate by the protein kinase C agonists was due to increased extrusion of protons from the cells, likely through V-ATPases located in the plasma membrane. Considered together, these data demonstrate regulation of plasmalemmal V-ATPase-mediated proton extrusion by protein kinase C.
Nordström
T
T
Division of Cell Biology, Hospital for Sick Children, Toronto, Ontario, Canada.
Grinstein
S
S
Brisseau
G F
GF
Manolson
M F
MF
Rotstein
O D
OD
D000818:Animals; D004789:Enzyme Activation; D005260:Female; D006863:Hydrogen-Ion Concentration; D017737:Macrophages, Peritoneal / Q000187:drug effects / Q000201:enzymology*; D051379:Mice; D011493:Protein Kinase C / Q000378:metabolism*; D006180:Proton-Translocating ATPases / Q000378:metabolism*; D011522:Protons; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013864:Thioglycolates / Q000494:pharmacology; D014617:Vacuoles / Q000201:enzymology*
D011522:Protons; D013864:Thioglycolates; D011493:Protein Kinase C; D006180:Proton-Translocating ATPases; D013755:Tetradecanoylphorbol Acetate