Fig. 4

Sgf73p participates in transitioning Yra1p onto nascent transcripts. a Average plots of ChIP-seq signals representing the binding of Yra1p in wildtype, rpt2-1 and sgf73Δ cells from 1.0 kb upstream of the TSS to 0.5 kb downstream of the TES, regarding the Sgf73-peak gene set (n = 580). b Average plots of ChIP-seq signals representing the binding of Yra1p and Sgf73p in the rpt2-1 mutant. c Schematic diagram of in vitro competition assay between Sgf73p and Pcf11p with respect to Yra1p binding. The Yra1p-Pcf11p complex was immobilized on GST sepharose beads. After a brief wash, Sgf73p was added and the amount of released Pcf11p in supernatants was assessed by Western blotting. d (Left panel) Western blotting of an in vitro competition assay performed between Sgf73p and Pcf11p (α-HIS) with respect to the binding of Yra1p (α-GST). (Right panel) Band intensity of released Pcf11p in the supernatant following the addition of Sgf73p. e R-IP-qPCR analysis of Yra1p (α-FLAG) and control (α-IgG) against the PMA1 transcript in wildtype, rpt2-1, and sgf73Δ cells. f R-IP-qPCR analysis of Sgf73p (α-FLAG) and control (α-IgG) against the PMA1 transcript in wildtype cells. g (Left panel) In vitro RNA-pulldown analysis of Sgf73p (α-MBP) against the GAL1 transcript. (Right panel) Western blotting against bead-bound Sgf73p (α-MBP). Standard deviations of all quantification data were obtained from three independent experiments and are shown by error bars; P-values were evaluated by Student’s t-test (*P ≤ 0.05, **P ≤ 0.01, and ***P ≤ 0.001). kb, kilobase. TSS, transcription start site. TES, transcription end site