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. 2024 Jan 16;31(2):203–216. doi: 10.1038/s41418-024-01256-y

Fig. 6. ABLIM1 interacts with IκBα and facilitates its ubiquitination.

Fig. 6

A Relative mRNA levels of NFKBIA and ABLIM1 in HCT116, RKO, and SW620 cells assayed by qRT-PCR. n = 3 for each group. Kruskal–Wallis test: *P < 0.05. B Immunoblotting of IκBα and ABLIM1 in HCT116 cells treated with CHX (10 μM) for indicated time. Quantification of the IκBα levels relative to GAPDH expression was displayed at the right. C Immunoblotting results of IκBα and ABLIM1 in ABLIM1 knockdown or overexpression HCT116 cells treated with PS341 (100 nM). D Co-immunoprecipitation assays revealed the interaction between ABLIM1 and IκBα in HCT116 and RKO cells. Cells were transfected with NFKBIA in combination with ABLIM1 or empty vector for 48 h. In control IP, IgG was used instead of ABLIM1 antibody. E Co-immunoprecipitation assayed the effect of ABLIM1 overexpression on IκBα ubiquitination in HCT116 and 293 T cells. Cells were transfected with NFKBIA, UBB, and ABLIM1 or empty vector for 48 h and treated with MG132 (20 μM) for 4 h before harvest. F In vivo ubiquitination assay using Ni-NTA pull-down confirmed that ABLIM1 promotes the poly-ubiquitination of IκBα.