FIGURE 6.

Mutation of Mad1 Thr⁶⁸⁰ abrogated SAC activity. A, mitotic indices of cells expressing Mad1-WT, Mad1-S22A, Mad1-S29A, Mad1-T680A, or Mad1-T680E after siRNA knockdown of cell endogenous Mad1 protein. Cells were pre-treated with Mad1-RNAi for 48 h and then transfected with Mad1-WT/mutants for another 24 h, followed by nocodazole (200 nm) treatment. 24 h later, cells were fixed and stained with anti-histone H3 phospho-Ser¹⁰ antibody (green) to visualize mitotic cells. Mitotic indices were scored (1000 cells were counted in each case) in panel B. DIC, differential interference contrast.