Fig. 4. Mitotic arrest deficient 2 like 2 (Mad2L2, also known as Mad2B) forms a complex with Rev3 and Rev1 that binds to cell division cycle (Cdc)-27 following cisplatin-induced DNA damage.
HEK293 cells were co-transfected with HA-Mad2B and either FLAG-tagged hRev 1 (a.a. 907–1251) (A) or FLAG-tagged hRev 3 (a.a. 1665–2070) (B). After transfection for 24 h, the cells were treated with cisplatin (50 μM) for 18 h. Cells were then lysed and HA-Mad2B immunoprecipitated and immunoblotted with anti-HA, anti-FLAG, and anti-γ-tubulin antibodies. Control immunoprecipitates were incubated with the lysis buffer alone. (C, D) HEK293 cells were co-transfected with either HA-Mad2B and FLAG-tagged hRev1 (a.a. 907–1251) or HA-Mad2B and hRev3 (a.a. 1665–2070). Cells were then lysed and Cdc27 was immunoprecipitated from the cell lysates. Immunoprecipitates were Western blotted with anti-FLAG (upper), anti-HA (middle), and anti-Cdc27 (lower) antibodies. For control immunoprecipitates, the Cdc27 antibody was incubated with the lysis buffer alone. Molecular weight markers (kDa) are indicated on the left of each Western blot (WB). Asterisk indicates the non-specific band. IP, immunoprecipitation; HA, hemagglutinin.
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