Figure 7. Pef1 and PP4 oppose each other for regulating Rad21 binding to chromosomes.

(A) Western blot analysis of total protein extracts from cycling cells probed with anti-Rad21 antibodies. (B) Cell growth assays showing that the ts growth defect of psm3^NN pph3Δ is efficiently rescued by pef1 and psl1 deletion mutants (left) and rad21-T262A (right). (C) Rad21-ChIP after one complete cell cycle at 36°C. Data are expressed relative to wild-type. Bars indicate mean ± SD from four ratios. t-tests are shown in Figure 7—figure supplement 1 and Figure 7—source data 1. (D) Cell growth assays showing that pef1Δ, rad21-T262A and pph3Δ display opposite genetic interactions with mis4-367. (E) Rad21 and Mis4 ChIP relative to wild-type in cdc10-129-arrested cells. Bars indicate mean ± SD from four ratios (Figure 4—source data 1).

Figure 7—figure supplement 1. Pairwise comparisons of ChIP data from Figure 7C and quantification of Rad21 in Mis4 immunoprecipitates.

(A,B) Pairwise comparisons of ChIP data from Figure 7C. Data are shown in two separate graphs for clarity. Bars indicate mean ± SD, n = 4. ***p≤0.001, **p≤0.01, *p≤0.05, by two-tailed, unpaired t-test with 95% confidence interval (Figure 7—source data 1). (C) Mis4-GFP was immuno-purified from G1 (cdc10-129) protein extracts and co-purifying proteins were analyzed by western blotting. Band intensities were measured and the ratios Rad21/Mis4 were normalized to wt. Bar = mean+/-SD from two biological replicates.