Receptor activator of NF-κB
ligand (RANKL)-RANK receptor signaling to induce
NFATc1 transcription factor is critical for osteoclast differentiation and
bone resorption. RANK adaptor
proteins,
tumor necrosis factor receptor-associated factors (TRAFs) play an essential role in RANKL signaling. Evidence indicates that NIP45 (NFAT interacting
protein) binds with TRAFs and NFATc2. We therefore hypothesized that NIP45 regulates RANKL induced osteoclast differentiation. In this study, we demonstrate that RANKL treatment down regulates NIP45 expression in mouse bone marrow derived pre-osteoclast cells. Lentiviral (pGIPZ) mediated
shRNA knock-down of NIP45 expression in RANKL stimulated pre-osteoclast cells resulted in increased levels of NFATc1, NFATc2, and
TRAF6 but not
TRAF2 expression compared to control
shRNA transduced cells. Also, NIP45 suppression elevated p-IκB-α levels and NF-κB-
luciferase reporter activity. Confocal microscopy demonstrated NIP45 colocalized with
TRAF6 in the cytosol of osteoclast progenitor cells. In contrast, RANKL stimulation induced NIP45 nuclear translocation and colocalization with NFATc2 in these cells. Coimmuneprecipitation assay demonstrated NIP45 binding with NFATc2 but not NFATc1. We further show that
shRNA knock-down of NIP45 expression in pre-osteoclast cells significantly increased RANKL induced osteoclast differentiation and
bone resorption activity. Taken together, our results indicate that RANKL signaling down regulates NIP45 expression and that NIP45 is a negative regulator of osteoclast differentiation.