α h2b total

To prepare yeast cell extracts, yeast cultures were grown to an OD600 of 0.5-0.8 in YPD medium. Total proteins were extracted using the trichloroacetic acid (TCA) method. All tandem affinity purifications (TAPs) were performed as previously described [37] . Briefly, TAP-fusion proteins and their associated proteins were recovered from cell extracts by affinity selection on IgG Sepharose beads. After bead washing, the Tobacco Etch Virus (TEV) protease was added to release the bound material. The eluate was incubated with calmodulin-coated beads in the presence of calcium. After washing, the bound material was released with EGTA. This enriched fraction was called the calmodulin eluate. To analyze the TAP-purified protein complexes, TCAprecipitation, LysC/trypsin digestion and multidimensional protein identification technology (MudPIT) mass spectrometry analyses were performed as described previously [38] . Following electrophoresis and western blotting, membranes were probed with specific antibodies: α-PGK1 was used as a loading control (Invitrogen), α-HA (Roche), α-TAP (Thermo Fisher), α-H2B total (Active Motif), α-H2Bub1 (Cell Signaling), α-Spt8 (Santa Cruz) and α-GFP (Roche). Quantification of the western blot bands was performed by densitometry using ImageJ software and subsequent normalization using the ratio between the protein to study and loading control protein.

To prepare yeast cell extracts, yeast cultures were grown to an OD 600 of 0.5-0.8 in YPD medium. Total proteins were extracted using the trichloroacetic acid (TCA) method. All tandem affinity purifications (TAPs) were performed as previously described [45] . Briefly, TAP-fusion proteins and their associated proteins were recovered from cell extracts by affinity selection on IgG Sepharose beads. After bead washing, the Tobacco Etch Virus (TEV) protease was added to release the bound material. The eluate was incubated with calmodulin-coated beads in the presence of calcium. After washing, the bound material was released with EGTA. This enriched fraction was called the calmodulin eluate. To analyze the TAP-purified protein complexes, TCA-precipitation, LysC/trypsin digestion and multidimensional protein identification technology (MudPIT) mass spectrometry analyses were performed as described previously [46] . Following electrophoresis and western blotting, membranes were probed with specific antibodies: α-PGK1 was used as a loading control (Invitrogen), α-HA (Roche), α-TAP (Thermo Fisher), α-H2B total (Active Motif), α-H2Bub1 (Cell Signaling), α-Spt8 (Santa Cruz) and α-GFP (Roche). Quantification of the western blot bands was performed by densitometry using ImageJ software and subsequent normalization using the ratio between the protein to study and loading control protein.