Bone Morphogenetic Proteins Signal Via SMAD and Mitogen-activated Protein (MAP) Kinase Pathways at Distinct Times during Osteoclastogenesis

Aaron Broege,Lan Pham,Eric D Jensen,Ann Emery,Tsang-Hai Huang,Melissa Stemig,Hideyuki Beppu,Anna Petryk,Michael O'Connor,Kim Mansky,Raj Gopalakrishnan

doi:10.1074/jbc.m113.496950

Abstract

To investigate the role of bone morphogenetic protein (BMP) signaling in osteoclastogenesis in vivo, we eliminated BMPRII in osteoclasts by creating a BMPRII(fl/fl);lysM-Cre mouse strain. Conditional knock-out (cKO) mice are osteopetrotic when compared with WT controls due to a decrease in osteoclast activity. Bone marrow macrophages (BMMs) isolated from cKO mice are severely inhibited in their capacity to differentiate into mature osteoclasts in the presence of M-CSF and receptor activator of NF-κB (RANK) ligand. We also show that BMP noncanonical (MAPK) and canonical (SMAD) pathways are utilized at different stages of osteoclast differentiation. BMP2 induces p38 phosphorylation in pre-fusion osteoclasts and increases SMAD phosphorylation around osteoclast precursor fusion. Phosphorylation of MAPKs was decreased in differentiated BMMs from cKO animals. Treating BMMs with the SMAD inhibitor dorsomorphin confirms the requirement for the canonical pathway around the time of fusion. These results demonstrate the requirement for BMP signaling in osteoclasts for proper bone homeostasis and also explore the complex signaling mechanisms employed by BMP signaling during osteoclast differentiation.

Highlights

bone morphogenetic protein (BMP) affect osteoclastogenesis in vitro, but the effects of Bone morphogenetic proteins (BMPs) signaling on osteoclastogenesis in vivo are not well understood
BMPRII Is Required for Efficient Osteoclast Differentiation— Previously, our laboratory showed that BMP2 enhances RANKL-mediated osteoclast differentiation and that noggin, an inhibitor of BMP signaling, blocks osteoclast differentiation [11]
Real-time RT-PCR confirmed that BMPRII and cathepsin K (Ctsk, a marker for osteoclast differentiation) mRNA expression is decreased in the Ad-Cre-treated BMPRIIfl/fl osteoclasts (Fig. 1, D and E)

Summary

Introduction

BMPs affect osteoclastogenesis in vitro, but the effects of BMP signaling on osteoclastogenesis in vivo are not well understood. Conclusion: BMP signaling is required for proper bone remodeling in vivo. To investigate the role of bone morphogenetic protein (BMP) signaling in osteoclastogenesis in vivo, we eliminated BMPRII in osteoclasts by creating a BMPRIIfl/fl;lysM-Cre mouse strain. Conditional knock-out (cKO) mice are osteopetrotic when compared with WT controls due to a decrease in osteoclast activity. We show that BMP noncanonical (MAPK) and canonical (SMAD) pathways are utilized at different stages of osteoclast differentiation. Treating BMMs with the SMAD inhibitor dorsomorphin confirms the requirement for the canonical pathway around the time of fusion. These results demonstrate the requirement for BMP signaling in osteoclasts for proper bone homeostasis and explore the complex signaling mechanisms employed by BMP signaling during osteoclast differentiation

Methods

Results

Conclusion