Fig. 7

TRIM67 inhibits NF-κB signaling by regulating the level of K48- and K63-linked ubiquitination of IκBα. A Primary cultured microglia were infected with recombinant adenovirus carrying Flag-TRIM67 and treated with OGD/R. Western blotting was used to assess the effect of TRIM67 on total IκBα protein levels. B In HEK293T cells transfected with Flag-TRIM67 and Myc-IκBα, the effects of TRIM67 overexpression on IκBα degradation were studied. C In HEK293T cells transfected with Flag-TRIM67, Myc-IκBα, and HA-ubiquitin, a representative co-IP study was performed to identify the ubiquitination of IκBα. D In HEK293T cells transfected with Flag-TRIM67, Myc-IκBα, and Flag-K48/K63-Ubiquitin (Ub), a representative co-IP analysis was performed to assess the degree of K48- and K63-linked polyubiquitination of IκBα. E A representative co-IP study was performed to examine the interaction of endogenous TRIM67 with IκBα in brain tissue. F A representative co-IP study was performed to examine the interaction of ectopically expressed TRIM67 with IκBα in HEK293T cells transfected with Flag-TRIM67 and Myc-IκBα plasmids. Data are representative of three independent experiments