Fig. 4: Nrd1 CID fusion to Set1Δ200 partially restores CTD-binding and H3K4 methylation.
From: The Set1 N-terminal domain and Swd2 interact with RNA polymerase II CTD to recruit COMPASS
a Schematic diagram of the NSΔ200 fusion protein consisting of the Nrd1 CID (amino acids 1–153) and Set1Δ200. A FLAG-tag at the N-terminus is not shown. The positions of two CID mutants that disrupt CTD binding are also shown. b, c FLAG-tagged full-length Set1 (Set1), Set1Δ200 (SΔ200), or Nrd1 CID fusions (NSΔ200, D70RSΔ200, and I130RSΔ200) were transformed into set1Δ cells. Whole-cell lysates (b) and proteins immunoprecipitated using anti-FLAG beads (c) were analyzed by immunoblotting using indicated antibodies. Cells transformed with empty plasmid (set1Δ) served as a negative control. d, e ChIP-Seq heat maps of H3K4me3 (d) and H3K4me2 (e) from strains carrying full-length Set1 (Set1), Set1Δ200 (SΔ200), or Nrd1CID–Set1Δ200 fusion (NSΔ200). After normalizing to an internal spiked-in sample of S. pombe chromatin, SPMR values were mapped for individual RNApII transcriptional units and ordered according to H3K4me3 values in Set1 cells. H3K4me levels within 1500 bps from transcription start site (TSS) are shown. ChIP-seq reads from two technical repeats were averaged and plotted. f, g FLAG-tagged full-length Set1 (Set1), Set1Δ200 (SΔ200), and Nrd1CID–Set1Δ200 (NSΔ200) were transformed into set1Δ or set1Δswd2Δ cells. f Whole-cell lysates or g anti-FLAG immunoprecipitates were separated by SDS-PAGE and analyzed by immunoblotting using the indicated antibodies. Histone H3 was used as a loading control. Source data are provided as a Source data file.
