Osteoclast Differentiation by RANKL Requires NF-κB-Mediated Downregulation of Cyclin-Dependent Kinase 6 (Cdk6)

Abstract

This study investigated the involvement of cell cycle factors in RANKL-induced osteoclast differentiation. Among the G1 cell cycle factors, Cdk6 was found to be a key molecule in determining the differentiation rate of osteoclasts as a downstream effector of the NF-B signaling.Introduction: A temporal arrest in the G1 phase of the cell cycle is a prerequisite for cell differentiation, making it possible that cell cycle factors regulate not only the proliferation but also the differentiation of cells. This study investigated cell cycle factors that critically influence differentiation of the murine monocytic RAW264.7 cells to osteoclasts induced by RANKL. Materials and Methods: Growth-arrested RAW cells were stimulated with serum in the presence or absence of soluble RANKL (100 ng/ml). Expressions of the G1 cell cycle factors cyclin D1, D2, D3, E, cyclin-dependent kinase (Cdk) 2, 4, 6, and Cdk inhibitors (p18 and p27) were determined by Western blot analysis. Involvement of NF-B and c-jun N-terminal kinase (JNK) pathways was examined by overexpressing dominant negative mutants of the IB kinase 2 (IKK DN ) gene and mitogen-activated protein kinase kinase 7 (MKK7 DN ) gene, respectively, using the adenovirus vectors. To determine the direct effect of Cdk6 on osteoclast differentiation, stable clones of RAW cells transfected with Cdk6 cDNA were established. Osteoclast differentiation was determined by TRACP staining, and cell cycle regulation was determined by BrdU uptake and flow cytometric analysis. Results and Conclusion: Among the cell cycle factors examined, the Cdk6 level was downregulated by RANKL synchronously with the appearance of multinucleated osteoclasts. Inhibition of the NF-B pathway by IKK

The Rho GTPase Wrch1 regulates osteoclast precursor adhesion and migration

“…Osteoclastic differentiation stimulated by RANKL requires osteoclast precursors to exit the cell cycle (Ogasawara et al, 2004). To test whether Wrch1 could be involved in this process, we analyzed the effect of Wrch1 overexpression on undifferentiated RAW264.7 cell growth.…”

“…They exit the cell cycle (Ogasawara et al, 2004) while a complex transcriptional program is activated.…”

Targeting CDK6 in cancer: State of the art and new insights

“…Among them is the ability of CDK6 to modulate differentiation in specific cell types including murine erythroid leukemia, primary mouse astrocytes, osteoblasts and osteoclasts, thymocytes, neurons, cardiomyocytes and hematopoietic cells. [22][23][24][25][26][27][28][29][30][31] In hematopoietic cells, for example, down-regulation of CDK6 allows terminal differentiation due to the release of runt-related transcription factor 1 (Runx1), a transcriptional factor required for the opening of chromatin of important hematopoietic regulator genes. 27 Location of CDK6 CDK6 has been shown to reside in the cytoplasm of many cell types.…”

Modulation of osteoclast differentiation and bone resorption by Rho GTPases

“…17,18 The main source of RANKL required for the formation of osteoclasts in bone remodeling comes from osteocytes. 18,19 RANK activation induces its trimerization and the recruitment of the adaptor molecule TRAF6.…”