Figure 2. Cells with hybrid-induced DNA damage are slow to repair.
(A) Assessment of cell-cycle delay in RNase H mutants. Asynchronously dividing cells were scored on the basis of their bud size and nuclear morphology. The percentage of cells with large buds and an undivided nucleus (single DAPI mass) are shown. Bars represent mean +/- standard deviation (n = 3, 100 cells scored per replicate) (B) Cell cycle profile of Rad52-GFP foci in dividing cells. rnh1∆ rnh201∆ cells were arrested in S-phase using hydroxyurea, washed, and released into alpha factor. Samples were taken at 30 min intervals and 300 cells per time point were scored for Rad52-GFP foci. If a cell had a Rad52-GFP focus, it was further scored for cell cycle phase. Cells with undivided nuclei (single DAPI mass) are labeled ‘pre-anaphase,’ while those that had undergone nuclear division (two DAPI masses or G1 arrested) are labeled ‘post-anaphase.’ (C) Cell cycle stage of dividing rnh1∆ rnh201∆ cells. Cells from (B) were subjected to flow cytometry (Figure 2—figure supplement 1) and quantified. The percentage of cells with 1C DNA content is shown.
Figure 2—figure supplement 1. Flow cytometry of rnh1∆ rnh201∆ cells released from hydroxyurea into alpha factor.
