Genomewide Comprehensive Analysis Reveals Critical Cooperation Between Smad and c-Fos in RANKL-Induced Osteoclastogenesis

We have previously reported that transforming growth factor (TGF-) plays an essential role in receptor activator of nuclear factor-B ligand (RANKL)-induced osteoclastogenesis. However, the detailed underlying molecular mechanisms still remain unclear. Formaldehyde-assisted isolation of regulatory elements (FAIRE) and chromatin immunoprecipitation (ChIP) followed by sequencing (FAIRE-seq and ChIP-seq) analyses indicated the cooperation of Smad2/3 with c-Fos during osteoclastogenesis. Biochemical analysis and immunocytochemical analysis revealed that physical interaction between Smad2/3 and c-Fos is required for their nuclear translocation. The gene expression of nuclear factor of activated T-cells, cytoplasmic 1 (Nfatc1), a key regulator of osteoclastogenesis, was regulated by RANKL and TGF-, and c-Fos binding to open chromatin sites was suppressed by inhibition of TGF- signaling by SB431542. Conversely, Smad2/3 binding to Nfatc1 was impaired by c-Fos deficiency. These results suggest that TGF- regulates RANKL-induced osteoclastogenesis through reciprocal cooperation between Smad2/3 and c-Fos. (c) 2014 American Society for Bone and Mineral Research.