Figure 4: Telomeric DSB sensing and preparation for microtubule-dependent repair.

(a) Anti-Myc-tagged protein ChIP followed by qPCR analysis reveals Lrs4-dependent increased interactions between the subtelomeric DSB and Cik1 (mean±s.d.; N=2 of triplicates) (*, t-test P≤0.01). (b) Anti-Myc immunoblotting indicates that DSB induction did not alter the level of Myc-tagged proteins used in ChIP analyses. (c) Co-immunoprecipitation using a TAP pull-down reveals no detectable interaction between TAP-tagged Lrs4 and Myc-tagged Nup84. (d) Genetic analysis of subtelomeric DSB repair. Survival (mean±s.d.; N=5) is compiled and presented as the ratio of colony-forming units on galactose (GAL) relative to glucose (GLU). P values (t-test) relative to wild type are presented immediately above mutants. Other P values are as illustrated on the graph. (e) Anti-Myc-tagged Nup84 and anti-Htz1 ChIP followed by qPCR analysis reveals Swr1-dependent chromatin remodelling (mean±s.d.; N=2 of triplicates) (*, t-test P≤0.01). (f) Anti-Myc immunoblotting indicates that DSB induction does not alter the level of Myc-tagged Nup84. (g) Anti-Mps3 ChIP followed by quantitative PCR analysis reveals that induced subtelomeric DSB–Mps3 interactions are increased in cells lacking Lrs4 or Cik1 (mean±s.d.; N=2 of triplicates) (*, t-test P≤0.01). (h) Anti-Mps3 immunoblotting indicates that DSB induction did not alter the level of the protein in our ChIP analysis. (i) Representative immunofluorescence images indicating that the loss of Lrs4 or Cik1 does not alter Mps3 protein localization within the nucleus. Composite images are shown with the scale bar, 2 μm. Untag., untagged.