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Osteoclasts are multinuclear cells which maintain bone homeostasis by resorbing bone. During bone resorption, osteoclasts attach to the bone matrix via a sealing zone formed by an actin ring. Rous sarcoma oncogene (Src) is essential for actin ring formation and bone resorption. Recently, we demonstrated that plectin, a cytolinker protein, is a Src-binding protein in osteoclasts. However, the function of plectin in osteoclasts remains unknown. In this study, we demonstrated that shRNA knockdown of plectin in RAW 264.7 cells resulted in tartrate resistant acid phosphatase positive multinuclear cells (TRAP (+) MNCs) with impaired actin ring formation and bone resorption activity. Moreover, we found that in plectin-silenced TRAP (+) MNCs, Src and protein tyrosine kinase 2 beta (Pyk2), two critical kinases in osteoclastic bone resorption, were inactivated and microtubule polarity was disturbed. These results suggest that plectin plays a critical role in osteoclast biology by acting as a scaffold to facilitate Src and Pyk2 activation during microtubule organization.

Endochondral ossification is important for skeletal development. Recent findings indicate that the p65 (RelA) subunit, a main subunit of the classical nuclear factor-κB (NF-κB) pathway, plays essential roles in chondrocyte differentiation. Although several groups have reported that the alternative NF-κB pathway also regulates bone homeostasis, the role of the alternative NF-κB pathway in chondrocyte development is still unclear. Here, we analyzed the in vivo function of the alternative pathway on endochondral ossification using p100-deficient (p100−/−) mice, which carry a homozygous deletion of the COOH-terminal ankyrin repeats of p100 but still express functional p52 protein. The alternative pathway was activated during the periarticular stage in wild-type mice. p100−/− mice exhibited dwarfism, and histological analysis of the growth plate revealed abnormal arrangement of chondrocyte columns and a narrowed hypertrophic zone. Consistent with these observations, the expression of hypertrophic chondrocyte markers, type X collagen (ColX) or matrix metalloproteinase 13, but not early chondrogenic markers, such as Col II or aggrecan, was suppressed in p100−/− mice. An in vivo BrdU tracing assay clearly demonstrated less proliferative activity in chondrocytes in p100−/− mice. These defects were partly rescued when the RelB gene was deleted in p100−/− mice. Taken together, the alternative NF-κB pathway may regulate chondrocyte proliferation and differentiation to maintain endochondral ossification.