Okadaic acid induces phosphorylation of p65NF-κB on serine 536 and activates NF-κB transcriptional activity in human osteoblastic MG63 cells

Nuclear factor-kappa B (NF-κB) is an essential transcription factor in the control of expression of genes involved in cell growth, differentiation, inflammation, and neoplastic transformation. Previously, we reported that okadaic acid (OA), which is a specific inhibitor of serine/threonine protein phosphatases, induced apoptosis in cells of human osteosarcoma cell line MG63. However, to date, it is not clear whether the phosphorylation status of NF-κB could be affected by the treatment with OA. In this report, we demonstrate that treatment of MG63 cells with OA enhanced the phosphorylation level of NF-κB, as judged from the results of Western blot analysis and a λ protein phosphatase dephosphorylation assay. The phosphorylation level of NF-κB was enhanced in both time- and dose-dependent manners. In the eel Is treated with 100 nM OA for 3 h, consequential translocation of NF-κB from the cytosol to the nucleus occurred. Western blotting experiments with an anti-phospho-p65NF-κB antibody disclosed that the NF-κB was phosphorylated on serine 536. Furthermore, OA stimulated the transcriptional activity of NF-κB in MG63 cells, as judged from the results of a luciferase assay. Our findings indicate that OA elicit phosphorylation of NF-κB on serine 536 in MG63 cells, resulting in the translocation of phospho-NF-κB to the nucleus, thereby promoting transcriptional activity of genes.