(A) Serial dilutions (ten-fold) of the indicated mutants were spotted on YPAD alone or on YPAD containing the indicated drug (B) The DNA content of asynchronous cultures of wild-type (WT) and rtt109Δ strains were determined by flow cytometric analysis. (C) Electrophoretic mobility of Rad53p was analyzed in extracts from asynchronous cultures of wild-type (WT), rtt109Δ and asf1Δ cells, and WT and rtt109Δ cells treated with phleomycin for 1 h. (D) Cell cycle distribution of WT and rtt109Δ cells with Rad52-YFP foci as determined by bud size. The average and standard deviation of two independent experiments are shown (E) GCR frequency was measured for the indicated strains (7). The average and standard deviation of three fluctuation tests are shown. (F) Recombination frequencies were measured in the indicated strains with a direct repeat recombination assay (8); the average and standard deviation of three fluctuation tests are shown.

(A) Western blot analysis of whole cell extracts isolated from the indicated strains was performed with antibodies specific for histone H3 or histone H3 acetylated on K56 (K56 Ac). (B) Serial dilutions (ten-fold) of the indicated strains were plated on YPAD or YPAD containing HU. (C) Whole cell extracts from the indicated strains were probed with antibodies for histone H3 and histone H3 K56 Ac. (D) Serial dilutions (ten-fold) of the indicated strains were plated on YPAD at the indicated temperature. (E) A strain expressing a HA-tagged version of Rtt109p was arrested in G1 with α-factor and released. Samples were taken at the indicated times for FACS and western analysis performed, probing for the indicated proteins with Pgk1p and Histone H3 as loading controls and Clb2p as a G2-M marker.

(A) Serial dilutions (ten-fold) of the indicated strains were plated on YPAD or YPAD containing HU or MMS. (B) DNA isolated from the indicated strains was electrophoresed on an agarose gel containing chloroquine. Superhelical density of the 2μ plasmid was analyzed by southern blotting and hybridization with a radioactively-labelled probe (left panel). Topoisomers were quantified by densitometric tracing (right panel).

(A) (³H) acetyl-CoA was incubated with histone octamers, BSA, Rtt109p or Asf1p as indicated. The ³H counts were measured and the mean and standard deviation from six independent experiments are shown. (B) Half the reaction mixture from (A) was electrophoresed on a SDS-PAGE gel and subsequently stained with Coomassie brilliant blue, or transferred onto a nitrocellulose membrane and subjected to autoradiography (C). Asterisks mark a degradation product of Asf1p. (D) Alternatively, a nitrocellulose membrane was prepared as in (C) and probed with an antibody specific for H3 K56 Ac.