Convergence of αvβ3Integrin–And Macrophage Colony Stimulating Factor–Mediated Signals on Phospholipase Cγ in Prefusion Osteoclasts

Abstract

The macrophage colony stimulating factor (M-CSF) and αvβ3 integrins play critical roles in osteoclast function. This study examines M-CSF– and adhesion-induced signaling in prefusion osteoclasts (pOCs) derived from Src-deficient and wild-type mice. Src-deficient cells attach to but do not spread on vitronectin (Vn)-coated surfaces and, contrary to wild-type cells, their adhesion does not lead to tyrosine phosphorylation of molecules activated by adhesion, including PYK2, p130Cas, paxillin, and PLC-γ. However, in response to M-CSF, Src−/− pOCs spread and migrate on Vn in an αvβ3-dependent manner. Involvement of PLC-γ activation is suggested by using a PLC inhibitor, U73122, which blocks both adhesion- and M-CSF–mediated cell spreading. Furthermore, in Src−/− pOCs M-CSF, together with filamentous actin, causes recruitment of β3 integrin and PLC-γ to adhesion contacts and induces stable association of β3 integrin with PLC-γ, phosphatidylinositol 3-kinase, and PYK2. Moreover, direct interaction of PYK2 and PLC-γ can be induced by either adhesion or M-CSF, suggesting that this interaction may enable the formation of integrin-associated complexes. Furthermore, this study suggests that in pOCs PLC-γ is a common downstream mediator for adhesion and growth factor signals. M-CSF–initiated signaling modulates the αvβ3 integrin-mediated cytoskeletal reorganization in prefusion osteoclasts in the absence of c-Src, possibly via PLC-γ.

The mechanism involved in the regulation of phospholipase Cγ1 activity in cell migration

“…This raises the possibility that the ability of FAK to activate PI 3-K may lead to integrin stimulation of PLCg1 activity. A similar mechanism has been recently proposed for the FAKrelated protein tyrosine kinase 2 (PYK2) (Nakamura et al, 2001). It has been demonstrated that PLCg1 is a common mediator for adhesion and growth factor signals in prefusion osteoclasts and that adhesion or macrophage stimulating factors induce direct interaction of PLCg1 with PYK2 (Nakamura et al, 2001).…”

“…A similar mechanism has been recently proposed for the FAKrelated protein tyrosine kinase 2 (PYK2) (Nakamura et al, 2001). It has been demonstrated that PLCg1 is a common mediator for adhesion and growth factor signals in prefusion osteoclasts and that adhesion or macrophage stimulating factors induce direct interaction of PLCg1 with PYK2 (Nakamura et al, 2001). Interestingly, these effects were blocked by PI 3-K inhibitors, indicating that PLCg1 is a downstream effector of PI 3-K and that this pathway can mediate cell spreading and migration in response to growth factor and cytokines.…”

M-CSF, TNFα and RANK ligand promote osteoclast survival by signaling through mTOR/S6 kinase

“…A working model has been proposed, where the absence of either signal would prohibit activation of downstream targets. 52 In the Ocl, M-CSF was shown to signal through the PI3K pathway to activate the prosurvival kinase, Akt (see Nakamura et al 19 and the present study). Signaling in Ocl to Akt by RANKL and TNFa, via p60 c-src has also been reported.…”

“…19 Akt activity is necessary and in some cases sufficient for cell survival and has been shown to target the apoptotic machinery by phosphorylating downstream molecules like BAD, caspase-9, glycogen-synthase kinase and forkhead family members. 20,21 RANKL and TNFa act primarily via NF-kB activation leading to the transcription and de novo synthesis of antiapoptotic proteins.…”

Phosphatidylinositol 3,4,5-Trisphosphate Directs Association of Src Homology 2-containing Signaling Proteins with Gelsolin

“…Recently, it has been shown in c-SrcϪ/Ϫ osteoclasts that macrophage colony-stimulating factor causes recruitment of ␤ 3 integrin and phospholipase C␥ and induces stable association of ␤ 3 integrin with phospholipase C␥, PI3K, and PYK2 (108).…”