title: Phosphatidylinositol 3-kinase association with the osteoclast cytoskeleton, and its involvement in osteoclast attachment and spreading.
abstract: Osteoclast activation involves attachment to the mineralized bone matrix and reorganization of the cytoskeleton, leading to polarization of the cell. Signaling molecules, PI3-kinase, rho A, and pp60c-src, were shown to be essential for osteoclastic bone resorption. In this study we have focused on the involvement of these signaling molecules in the early event of osteoclast activation: attachment, spreading, and organization of the cytoskeleton. Highly purified osteoclasts were fractionated into Triton X-100-soluble or cytosolic and Triton X-100-insoluble or cytoskeletal fractions, and the distribution of above-mentioned signaling molecules between the two fractions was examined. PI3-kinase, rho A, and pp60c-src all showed translocation to the cytoskeletal fraction upon osteoclast attachment to plastic. However, PI3-kinase and rho A, but not pp60c-src, showed further translocation of 2.4- and 3.2-fold, respectively, upon attachment of osteoclasts to bone. PI3-kinase translocation to the cytoskeleton was inhibited by either cytochalasin B or colchicine. Furthermore, treatment of osteoclasts with the PI3-kinase inhibitor wortmannin decreased its translocation, suggesting that PI3-kinase activity was needed for its translocation. Moreover, wortmannin inhibited osteoclast attachment to both bone and plastic and caused drastic changes in osteoclast morphology resulting in rounding of the cells, disappearance of F-actin structures or podosomes, and appearance of punctate or vesicular structures inside the cells. Osteoblastic MB1.8 cells and IC-21 macrophages did not show additional translocation of PI3-kinase or rho A upon attachment to bone or changes in attachment or morphology in response to wortmannin. Finally, PI3-kinase coimmunoprecipitated with alpha v beta 3 integrin from osteoclasts.
authors.affiliation: Department of Bone Biology and Osteoporosis Research, Merck Research Laboratory, West Point, Pennsylvania 19486, USA.
authors.forename: P T
authors.initials: PT
authors.lastname: Lakkakorpi
authors.forename: G
authors.initials: G
authors.lastname: Wesolowski
authors.forename: Z
authors.initials: Z
authors.lastname: Zimolo
authors.forename: G A
authors.initials: GA
authors.lastname: Rodan
authors.forename: S B
authors.initials: SB
authors.lastname: Rodan
mesh_terms: D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology; D001842:Bone and Bones / Q000166:cytology; D002448:Cell Adhesion; D002478:Cells, Cultured; D003599:Cytoskeleton / Q000201:enzymology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism; D051379:Mice; D010010:Osteoclasts / Q000201:enzymology* / Q000648:ultrastructure; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D019030:Receptors, Vitronectin / Q000378:metabolism; D000077191:Wortmannin; D020742:rhoA GTP-Binding Protein
chemical_list: D000730:Androstadienes; D004791:Enzyme Inhibitors; D019204:GTP-Binding Proteins; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D019030:Receptors, Vitronectin; D000077191:Wortmannin; D020742:rhoA GTP-Binding Protein