Figure 4. MDC1 and BRIT1 were negative regulators of Aurora A and Plk1.
(A) U2OS cells were transfected with luciferase siRNA or MDC1 siRNA (left) or BRIT1 siRNA (right). At 48 h after transfection, cells were harvested and the lysates were subjected to Western blotting using antibodies against (left) MDC1, Plk1 and Aurora A or (right) BRIT1, Plk1 and Aurora A. Actin is for the loading control. (B) U2OS cells were transfected with the siRNA against luciferase, MDC1, or BRIT1. At 48 h after transfection, cells were untreated or treated with cycloheximide for 8 or 12 h. Cells were then harvested, and the lysates were subjected to Western blotting using antibodies against Plk1, Aurora A or actin. (C) U2OS cells were transfected with siRNA against luciferase or MDC1 along with cotransfection of empty expression vector or the vectors expressing wild-type pEGFP-C2-MDC1 or pEGFP-C2-ΔFHA-MDC1 mutant. At 48 h after the transfection, cells were harvested, and the lysates were subjected to Western blotting using antibodies against MDC1, Plk1, Aurora A. Actin is used for the loading control.