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. 2018 Jul 31;37(17):e99154. doi: 10.15252/embj.201899154

Figure 6. Rtt105 is important for DNA replication and genome stability maintenance.

Figure 6

The procedure for BrdU IP‐seq is described in Fig 2A.
  1. Snapshots of BrdU IP‐seq peaks at ARS305 from cells released into YPD medium containing HU for 45 min.
  2. The average BrdU IP‐seq read density around fired ACS sites from cells released into YPD medium containing HU for 45 min.
  3. Snapshots of BrdU IP‐seq peaks at ARS305 from cells released into YPD medium at 16°C for 72 min.
  4. The average BrdU IP‐seq read density around ACS sites from cells released into 16°C for 72 min.
  5. Rad52‐GFP foci were counted in WT and rtt105Δ cells, and the percentage of cells with Rad52‐foci is reported. Error bars indicate one standard deviation from three independent experiments with at least 150 cells counted in each replicate. Statistical significance was evaluated based on Student's t‐tests (****P‐value < 0.0001).
  6. Rad53 phosphorylation was analyzed in WT and rtt105Δ cells by immunoblotting of protein extracts with anti‐Rad53 antibodies. Ponceau S (Pon S) staining was applied as a loading control.
  7. Deletion of RTT105 leads to an increased rate of gross chromosomal rearrangements (GCRs). Wild‐type (WT) and rtt105Δ mutant strains integrated with the yWSS439‐5oriΔ yeast artificial chromosomes (YACs). GCRs rates in the evaluation of telomere marker loss were assayed and calculated as previously described (Huang & Koshland, 2003). Data represent the mean (±SD) of three independent experiments. Statistical significance was evaluated based on Student's t‐tests (**0.001 ≤ P‐value < 0.01).
  8. Ten‐fold serial dilutions of WT or rtt105Δ cells in two different backgrounds were assayed on normal growth media (YPD) and on media containing the indicated DNA‐damaging agents, methyl‐methanesulfonate (MMS), camptothecin (CPT), bleomycin (Bleo), and hydroxyurea (HU).
Source data are available online for this figure.