Figure 4. N-terminal and C-terminal SP/TP sites of Rad9 act redundantly to control DNA damage checkpoint activation. Gel mobility shift assays of Rad9–3HA and Rad53 were performed using G₂/M-arrested cells and asynchronous cells as indicated in (A and C), respectively. (B) Plate sensitivity assay of WT, mrc1Δ, rad9Δ and various double mutants of mrc1Δ rad9. (D) Percentages of bi-nucleated cells after release from phleomycin treatment in the G₂/M phase, comparing WT, rad9Δ, rad9–8AP, rad9-K1088E and rad9–8AP,K1088E mutants. (E) Tetrad dissection analysis of the genetic interactions between various rad9 mutants, mrc1Δ and sml1Δ. Synthetic lethality between mrc1Δ and either rad9Δ or rad9–8AP,K1088E is rescued by sml1Δ. No synthetic lethality was detected between mrc1Δ and rad9–8AP or rad9-K1088E. Representative tetrads are shown here for simplicity. Unmarked spores contain RAD9 MRC1 SML1.