TRIM13 stabilizes CCNA1 through ubiquitination. (A-B) Coimmunoprecipitation (Co-IP) of indicated factors 48 hours after transfection in 293T cells. (C) Co-IP of indicated TRIM13 domain mutants 48 hours after transfection in 293T cells. (D) Denaturing IP for FLAG-CCNA1 in 293T cells expressing TRIM13-V5 and indicated HA-tagged R-K ubiquitin. (E) Cell-free ubiquitination assay with a library of E2s. Bottom panel is ponceau stain, E1/GST-CCNA1/TRIM13-E2s are indicated. (Top) Blot for ubiquitin. (F) Western blot analysis of indicated genotypes U937 cells after cycloheximide chase. (G) Statistical analysis of CCNA1 half-life replicates. (H-J) Western blot analysis of T13WT and T13RINGdel U937 cells treated with indicated inhibitors: MG132 (H), Bafilomycin (BAF) (I-J), and cyclohexamide (CHX) (J). (K) Statistical analysis of CCNA1 half-life replicates in the presence of BAF. Results shown are representative of 3 biological replicates (A-D,F,H-J), or are mean + SD from 3 biological replicates (G,K). (G,K) Normalized to T13WT CCNA1 Veh level.