PLCγ2 regulates osteoclastogenesis via its interaction with ITAM proteins and GAB2
Dailing Mao, … , Deborah V. Novack, Roberta Faccio
Dailing Mao, … , Deborah V. Novack, Roberta Faccio
Published November 1, 2006
Citation Information: J Clin Invest. 2006;116(11):2869-2879. https://doi.org/10.1172/JCI28775.
View: Text | PDF
Research Article Bone biology

PLCγ2 regulates osteoclastogenesis via its interaction with ITAM proteins and GAB2

Abstract

Excessive bone loss in arthritic diseases is mostly due to abnormal activation of the immune system leading to stimulation of osteoclasts. While phospholipase Cγ (PLCγ) isoforms are known modulators of T and B lymphocyte–mediated immune responses, we found that blockade of PLCγ enzymatic activity also blocks early osteoclast development and function. Importantly, targeted deletion of Plcg2 in mice led to an osteopetrotic phenotype. PLCγ2, independent of PLCγ1, was required for receptor activator of NF-κB ligand–induced (RANKL-induced) osteoclastogenesis by differentially regulating nuclear factor of activated T cells c1 (NFATc1), activator protein–1 (AP1), and NF-κB. Specifically, we show that NFATc1 upregulation is dependent on RANKL-mediated phosphorylation of PLCγ2 downstream of Dap12/Fc receptor γ (Dap12/FcRγ) receptors and is blocked by the PLCγ inhibitor U73122. In contrast, activation of JNK and NF-κB was not affected by U73122 or Dap12/FcRγ deletion. Interestingly, we found that in osteoclasts, PLCγ2 formed a complex with the regulatory adapter molecule GAB2, was required for GAB2 phosphorylation, and modulated GAB2 recruitment to RANK. Thus, PLCγ2 mediates RANKL-induced osteoclastogenesis and is a potential candidate for antiresorptive therapy.

Authors

Dailing Mao, Holly Epple, Brian Uthgenannt, Deborah V. Novack, Roberta Faccio

×

Figure 7

PLCγ2 forms a complex with GAB2 and modulates its activation.

Options: View larger image (or click on image) Download as PowerPoint
PLCγ2 in RANKL signaling. PLCγ2 is phosphorylated by RANKL in an SFK-dep...
(A) WT preOCs cultured with or without the PLC inhibitor U73122 (5 μM) for 3 days were stimulated with RANKL and subjected to Western blot analysis for phospho-JNK, phospho-IκBα, and NFATc1. β-Actin served as control. (B) WT and Plcg2–/– BMMs retrovirally transduced with empty vector (pMX), WT PLCγ2, or catalytically inactive PLCγ2 (PLCγ2 H/F) were cultured with RANKL (100 ng/ml) and M-CSF (10 ng/ml) for 7 days, and multinucleated OCs were detected by TRAP staining. Objective, ×10. (C) The same cells as shown in B were subjected to RANKL stimulation and Western blot analysis to detect NFATc1 expression. (D) Plcg2–/– BMMs retrovirally transduced with pMX, WT PLCγ2, or PLCγ2 H/F were treated with RANKL, and phosphorylation of IκBα and JNK was determined by Western blot analysis. PLCγ2 expression levels are shown. β-Actin served as loading control in C and D. (E) PLCγ2 and GAB2 were reciprocally immunoprecipitated in WT and Plcg2–/– BMMs treated with RANKL and subjected to Western blot analysis using anti-PLCγ2 and anti-GAB2 Abs, respectively. TCL, total cell lysate. (F) GAB2 was immunoprecipitated in WT and Plcg2–/– BMMs and subjected to Western blot analysis using anti-phosphotyrosine Ab (clone 4G10), anti-RANK, and anti-PLCγ2.