Elevated DDR signalling is deleterious for growth of <i>yku70Δ</i> cells upon NAM- and MMS-induced replicative stress.

<p>(A) Mutations that abolish H4K16ac cause a reduction in H3K79me3 levels. Protein samples from asynchronous cells were immunoblotted with indicated antibodies. Bar graph represents the ratio of H3K79me3 onto H4 signals as quantified by densitometry. Error bars: standard error of the mean (B) Deletion of <i>DOT1</i> rescues growth of <i>yku70Δ</i> mutants in NAM (C) <i>sas2Δ</i> and <i>dot1Δ</i> mutations reduce Rad53 activation in <i>yku70Δ</i> cells. Cells were exposed to 20 mM NAM for 8 hours at 30°C and samples were taken for Rad53 <i>in situ</i> autophosphorylation assays (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1007356#sec009" target="_blank">materials and methods</a> for details). (D-E) <i>rad9Δ</i> (D) and <i>RAD53-HA</i> (E), which limit DDR signalling in response to replicative stress, rescue growth of <i>yku70Δ</i> mutants in NAM. Growth assay in 96-well plates (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1007356#sec009" target="_blank">materials and methods</a>). Error bars: standard deviation. (F) Deletion of <i>RAD9</i> rescues S phase progression defects of <i>yku70Δ</i> cells exposed to NAM. Asynchronous cells were incubated in YPD for 8 hours at 30°C in the presence of 20 mM NAM. Samples were taken at indicated time for flow cytometry-based DNA content analysis. (G-H) <i>yku70Δ</i> display synthetic sensitivity to MMS when combined with <i>slx4Δ</i> (G) and <i>pph3Δ</i> (H) mutants.