Rad4 mainly functions in Chk1-mediated DNA damage checkpoint pathway.

Abstract

<p><b>A</b>, wild type cells or cells with the integrated mutations were treated with (+) or without (−) MMS at 30°C for 1 hour. Phosphorylation of Chk1 was monitored by the mobility shift assay as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0092936#s2" target="_blank">Materials and Methods</a>. A section of the Ponceau S stained membrane is shown for the loading (bottom). <b>B</b>, Rad3-dependent phosphorylation of Cds1-Thr¹¹ was examined by phosphor-specific antibody (top panel). Loading of Cds1 is shown in the lower panel. <b>C</b>, sensitivity of the cells to acute HU (left) or MMS (right) treatment. Cells were treated with the drugs in YE6S medium at 30°C. At each time point, an aliquot of the culture was removed and spread on YE6S plates for the cells to recover. Colonies were counted and viability was presented as percentages of the untreated cells. Each data point represents an average of three independent experiments for each mutant. <b>D</b>, synthetic effects of the double mutants containing Δ<i>cds1</i> or Δ<i>chk1</i> with the indicated <i>rad4</i> mutations were examined by spot assay.</p