FIG 7.

Interaction of KDM2B with the SCF complex. (A) Composition of the SCF E3 ubiquitin ligase complex: the substrate receptor F-box protein, the adaptor protein SKP1, the scaffold protein Cullin-1 (CUL1), and the RING finger E3 ubiquitin ligase ROC1. (B) FLAG IP using 3×FLAG-KDM2B-transfected HEK293T cells. KDM2B interacts with endogenous SKP1. (C) HEK293T cells were cotransfected with 3×FLAG-KDM2B along with different combinations of epitope-tagged CUL1, SKP1, and ROC1. HA-ROC1 IP was analyzed by immunoblotting. (D) Schematic representation of the full-length KDM2B (WT) and its mutants. (E) FLAG IP using HEK293T cells transfected with WT, the F-box mutant, or the LRR domain mutant of 3×FLAG-KDM2B. The IPs were analyzed by anti-FLAG antibody and for the presence of endogenous SKP1. (F) HEK293T cells were cotransfected with myc-FLAG-SKP1 and WT or the indicated mutants of 3×FLAG-KDM2B, followed by myc IP at 48 h posttransfection. KDM2B and SKP1 were detected by FLAG and myc antibodies, respectively.