Involvement of the orphan nuclear estrogen receptor-related receptor α in osteoclast adhesion and transmigration

The orphan nuclear receptor, estrogen receptor-related receptor alpha (ERR alpha) is expressed in osteoblasts and osteoclasts (OCs) and has been proposed to be a modulator of estrogen signaling. To determine the role of ERR alpha in OC biology, we knocked down ERR alpha activity by transient transfection of an siRNA directed against ERR alpha in the RAW264.7 monocyte-macrophage cell line that differentiates into OCs in the presence of receptor activator of nuclear factor kappa B-ligands and macrophage colony-stimulating factor. In parallel, stable RAW cell lines expressing a dominant-negative form of ERR alpha and green fluorescent protein (RAW-GFP-ERR alpha Delta AF2) were used. Expression of OC markers was assessed by real-time PCR, and adhesion and transmigration tests were performed. Actin cytoskeletal organization was visualized using confocal microscopy. We found that RAW264.7 cells expressing siRNA directed against ERR alpha and RAW-GFP-ERR alpha Delta AF2 OCs displayed abnormal spreading, and decreased osteopontin and beta 3 integrin subunit expression compared with the corresponding control cells. Decreased adhesion and the absence of podosome belts concomitant with abnormal localization of c-src were also observed in RAW-GFP-ERR alpha Delta AF2-derived OCs. In addition, RAW-GFP-ERR alpha Delta AF2-derived OCs failed to transmigrate through osteoblast cell layers. Our data show that the impairment of ERR alpha function does not alter OC precursor proliferation and differentiation but does alter the adhesion/spreading and migration capacities of mature OCs.