Enhancement of RANKL-induced MITF-E expression and osteoclastogenesis by TGF-β

Microphthalmia-associated transcription factor (MITF) is a transcription factor that is expressed in limited types of cells, including osteoclasts, but the expression and role of MITF during osteoclastogenesis have not been fully elucidated. The expression of the MITF-E isoform but not that of the MITF-A isoform was induced in response to differentiation stimulation towards osteoclasts by receptor activator of NF-κB ligand (RANKL) in both RAW264.7 cells and primary bone marrow cells. The RANKL-induced formation of tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells was inhibited in RAW264.7 cells expressing siRNA for MITF-E. Transforming growth factor-β (TGF-β) enhanced RANKL-induced MITF-E expression and -TRAP positive multinucleated cell formation. In particular, TGF-β potentiated the formation of larger osteoclasts. The expression levels of NFATc1, TRAP and CtsK, genes related to osteoclast development and activity, were concurrently enhanced by TGF-β in the presence of RANKL. Furthermore, the expression of dendritic cell-specific transmembrane protein (DC-STAMP), Itgav, Itga2, Itga5, Itgb1, Itgb3 and Itgb5, genes related to cell adhesion and fusion, were up-regulated by co-treatment with TGF-β. In particular, the regulatory expression of Itgav and Itgb5 in response to RANKL with or without TGF-β resembled that of MITF-E. Because MITF is involved in cell fusion in some cell systems, these results imply a role for MITF-E as an enhancer of osteoclastogenesis and that RANKL-induced levels of both MITF-E mRNA and of MITF-dependent gene expression are enhanced by treatment with TGF-β.