Fig. 3
KDM4B promotes the membrane localization of AKT by promoting TRAF6-mediated ubiquitination of AKT. a The membrane fractions, cytoplasm fractions and whole cell extracts were collected in KDM4B-depressed LoVo cells to measure the phosphorylation of AKT at Thr308 and Ser 473. b KDM4B-depressed LoVo cells and control cells were fixed for immunofluorescence analysis by confocal microscopy. c In vivo ubiquitination assay in KDM4B-depressed 293 T cells transfected with HA-AKT, His-UB. Ubiquitinated AKT was detected in AKT immunoprecipitates. d HA-TRAF6 and Flag-KDM4B were transfected into 293 T cells as indicated. Flag-KDM4B proteins were immunoprecipitated by anti-HA antibody. Both the whole-cell lysates (WCE) and immunoprecipitated were analyzed by IB with anti-HA or anti-Flag antibody. e LoVo cells were stimulated with IGF-1 for the indicated times. The cell lysates were immunoprecipitated with anti-TRAF6 antibody or control IgG. Both the WCE and the immunoprecipitates were analyzed to detect the expression of KDM4B and TRAF6