ABSTRACT
Maturation of MC3T3-E1 osteoblast cells in vitro can be divided into three major stages, namely, proliferation, differentiation, and mineralization. Our microarray analysis identified genes differentially expressed between proliferating and differentiated MC3T3-E1 osteoblastic cells. Immunohistochemical analyses of RNF11 protein encoded by one of the differentially expressed genes revealed it as highly expressed in osteoblasts of multiple skeletal elements during embryonic bone formation in mice. In contrast, cartilage, undifferentiated mesenchymal tissue and osteocytes did not express detectable amounts of the RNF11 protein. The RNF11 mRNA was found to be abundant during the proliferation stage of MC3T3-E1 osteoblast development with a short second peak of expression during the mineralization phase. This pattern of expression is similar to that of the Ets1 transcription factor during osteogenic differentiation of MC3T3-E1 cells, and we used immunohistochemistry to show that, in vivo, Ets1 protein is coexpressed with RNF11 in osteoblasts. The human and mouse RNF11 promoters each contain three Ets transcription factor binding sites (EBS) and we found that Ets1 binds specifically to one of them. The functionality of this site was tested in a transcription-transactivation assay, indicating that RNF11 expression in bone cells is regulated by the Ets1 factor.
