Interaction between vacuolar H(+)-ATPase and microfilaments during osteoclast activation.
Vacuolar H(+)-ATPases (V-ATPases) are multisubunit enzymes that acidify compartments of the vacuolar system of all eukaryotic cells. In osteoclasts, the cells that degrade bone, V-ATPases, are recruited from intracellular membrane compartments to the ruffled membrane, a specialized domain of the plasma membrane, where they are maintained at high densities, serving to acidify the resorption bay at the osteoclast attachment site on bone (Blair, H. C., Teitelbaum, S. L., Ghiselli, R., and Gluck, S. L. (1989) Science 249, 855-857). Here, we describe a new mechanism involved in controlling the activity of the bone-resorptive cell. V-ATPase in osteoclasts cultured in vitro was found to form a detergent-insoluble complex with actin and myosin II through direct binding of V-ATPase to actin filaments. Plating bone marrow cells onto dentine slices, a physiologic stimulus that activates osteoclast resorption, produced a profound change in the association of the V-ATPase with actin, assayed by coimmunoprecipitation and immunocytochemical colocalization of actin filaments and V-ATPase in osteoclasts. Mouse marrow and bovine kidney V-ATPase bound rabbit muscle F-actin directly with a maximum stoichiometry of 1 mol of V-ATPase per 8 mol of F-actin and an apparent affinity of 0.05 microM. Electron microscopy of negatively stained samples confirmed the binding interaction. These findings link transport of V-ATPase to reorganization of the actin cytoskeleton during osteoclast activation.
Lee
B S
BS
Department of Internal Medicine, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Gluck
S L
SL
Holliday
L S
LS
D008841:Actin Cytoskeleton / Q000378:metabolism* / Q000648:ultrastructure; D000199:Actins / Q000378:metabolism; D000818:Animals; D002417:Cattle; D003599:Cytoskeleton / Q000737:chemistry; D003804:Dentin / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D051379:Mice; D008854:Microscopy, Electron; D009218:Myosins / Q000378:metabolism; D017830:Octoxynol; D010010:Osteoclasts / Q000166:cytology / Q000378:metabolism*; D011233:Precipitin Tests; D011485:Protein Binding; D006180:Proton-Translocating ATPases / Q000378:metabolism* / Q000648:ultrastructure; D011817:Rabbits; D025262:Vacuolar Proton-Translocating ATPases
D000199:Actins; D017830:Octoxynol; D025262:Vacuolar Proton-Translocating ATPases; D006180:Proton-Translocating ATPases; D009218:Myosins