FIGURE 1.

Rpb4/7 is required for Ccr4-Not to rescue arrested RNAPII. A, purification of intact (WT) and Rpb4/7-less RNA polymerase (Rpb4/7Δ). Gels were stained with Coomassie Blue. Rpb8 migrates differently in the last lane because of the calmodulin binding peptide (CBP) tag. M, molecular size markers. B, schematic diagram of elongation complex formation and transcription run-off assays. C, representative transcription run-off gels using intact (12-subunit) and Rpb4/7Δ RNAPII. Reactions were stopped at 1, 2, 4, and 8 min after the addition of nucleotides. Where indicated, 1.5 μg of Ccr4-Not was added to the reactions. D, averages of the quantification of three separate experiments. Error bars represent S.D. Intact RNAPII (black lines) or Rpb4/7Δ RNAPII (gray lines) reactions were incubated with Ccr4-Not (C/N), (solid lines), or an equivalent amount of bovine serum albumin carrier protein (dashed lines). The percentage of run off of ECs was plotted over time.