FIG. 5.

TFIID is intact in WT and Ts48 cells. (A) Measurements of Taf1p and Taf4p co-occupancy on the RPS5 gene promoter analyzed by sequential ChIP (ChIP-ChIP). Sheared formaldehyde cross-linked WT and Ts48 chromatin preparations were subjected to a first IP with anti-HA (Taf1p) antibody. Immunoprecipitated Taf1p-DNA complexes were eluted with HA peptide, and a second IP was performed on this eluted material with anti-Taf4p antibodies. Aliquots of both immunoprecipitates were analyzed by real-time quantitative PCR, and results were normalized and plotted as RPS5 gene promoter enrichment over DNA PolI (POL1) ORF. (B) Gel filtration of TFIID extracted from WT and Ts48 cells. Equal aliquots of alternate fractions from gel filtration chromatography of WT and Ts48 WCE were trichloroacetic acid precipitated, fractionated by SDS-PAGE, and blotted, and different TFIID-specific Tafps were detected with specific antibodies (left). The WCE input (I) and column fractions analyzed are indicated (bottom); the elution positions of various size markers run in parallel are shown (top).