Fig. 3: srr1 but not skb1 mutations increase DNA damage sensitivity.

a A serial dilution assay to determine the sensitivity to DNA damaging agents. Log-phase cultures of wild-type, srr1-W157R, srr1Δ, rad51Δ, srr1-W157R rad51Δ, and srr1Δ rad51Δ (TNF3885, 8280, 5847, 5845, 8573, and 5849) prepared in YE3S medium were spotted onto YE3S plates supplemented with the indicated concentrations of MMS, HU, or CPT (top panels). Log-phase cultures of wild-type, skb1Δ, rad51Δ, and skb1Δ rad51Δ (TNF35, 8321, 8107, and 8320) prepared in YE medium were spotted onto YE plates (bottom panels). b Wild-type, srr1Δ, skb1Δ, and chk1Δ cells (TNF35, 5943, 8321, and 3559) in the log phase in EMM were treated with 0.01% MMS. The percentage of cells containing a septum is indicated. > 300 cells are counted at each point. Pearson’s Chi-square test of the septation index between wild-type and other strains at t = 8 h showed that srr1Δ or skb1Δ did not significantly change the septation index (p > 0.05) but chk1Δ increased it. c Chk1 phosphorylation in response to MMS treatment. Before and after 4 h treatment with 0.01% MMS, extracts were prepared from chk1⁺ (TNF7555) and chk1-HA⁺ cells of wild-type, srr1Δ, and skb1Δ (TNF8441, 8799, and 8802) and separated by 8% SDS-PAGE. Chk1-HA was detected by Western blotting using anti-HA antibodies (16B12). Whole proteins were stained using Coomassie brilliant blue. Size markers (Takara, 3454 A, CLEARLY stained protein ladder) are shown on the left. wt, wild-type. Uncropped images are shown in Supplementary Fig 6. d Wild-type and srr1Δ (TNF5369 and 5774) cells were plated onto adenine-limited YE plates, on which ade6^– cells form red colonies. e Chromosome loss rates of wild-type, srr1Δ, srr1-W157R, skb1Δ, rad51Δ, srr1-W157R rad51Δ, and skb1Δ rad51Δ strains (TNF5369, 5774, 8308, 5772, 5411, 8344, and 5788). The two-tailed Mann-Whitney test. **p < 0.01; ***p < 0.001. Numerical data underlying b, e are provided in Tables C and D, respectively, in Supplementary Data 1.