Fig. 6.

FBXO7 accelerates formation of the TAK1–NRAGE complex and NRAGE poly-ubiquitination upon BMP4 treatment. a HEK293 cells were mock-transfected or transfected for 24 h with FBXO7-siRNAs or control siRNAs, and treated with 10 ng/ml BMP4 for 24 h. Cell lysates were immunoprecipitated with the anti-TAK1 antibody and immunoblotted with the anti-NRAGE antibody. Control immunoprecipitations were performed with rabbit IgG. To determine equal loading, cell lysates were analyzed by immunoblotting with anti-tubulin. b HEK293 cells were transfected for 24 h with HA-tagged ubiquitin and Myc-tagged NRAGE alone or in combination with Flag-tagged FBXO7, and treated with 10 ng/ml BMP4 for additional 24 h. Cell lysates were immunoprecipitated with the anti-Myc antibody and immunoblotted with the anti-HA antibody. Ubiquitin, NRAGE, and FBXO7 expression in cell extracts was determined by immunoblotting with the appropriate antibodies