Figure 1.
Analysis of Bye1, TFIIS and Pol II chromatin occupancies in yeast: (A, B) Genome-wide scatter-plots of Bye1 and Pol II distribution assessed by ChIP-chip in the Bye1-Myc HA-TFIIS strain.
Comparison of fold enrichment (A) between Pol II and Bye1 and (B) between TFIIS and Bye1. Red dots correspond to probes complementary to class III genes (listed in Table S7); (C) Quantification of Bye1 enrichment in coding regions of class II and class III genes, promoters of some class II genes and Y’-containing telomeric ends by ChIP. 3′ORF and mORF stand for the 3′end and middle coding regions.
Figure 2.
Bye1 association with class II genes requires active transcription: (A) Quantification of Pol II enrichment in rpb1-1 mutant and Bye1-Myc enrichment in RPB1 WT and rpb1-1 strains at 30°C and 37°C by ChIP; (B) Quantification of Bye1 enrichment at the heat-induced HSP104 locus at 28°C and 37°C by ChIP.
An intergenic genomic locus is used for a relative occupancy calculation; 3′ORF and mORF stand for the 3′end and middle coding regions.
Figure 3.
Bye1 is associated with the Pol II Rpb1 subunit in vivo: (A) Co-immunoprecipitation of Bye1-Myc, HA-TFIIS and Rpb1 from cross-linked cellular extracts; (B) Bye1 and TFIIS interact with the jaw domain (aa 1154–1252) of Rpb1 by Y2H.
All three domains, TFIIS-like, PHD and SPOC, are essential for Rpb1-Bye1 interaction: overlay of the Y190 strain transformed with either empty GAD/GBD vectors or GAD fused to the Rpb1 jaw domain and GBD fused to Bye1 or TFIIS. Left panel: control for the autoactivated β-galactosidase activity. Right panel: interactions are observed for Rpb1 and TFIIS and for Rpb1 and Bye1 the whole and the PHD and SPOC deleted proteins.
Figure 4.
Bye1 association with promoters of class II genes requires H3K4me3: (A, B) Quantification of Bye1-Myc enrichment in WT and strains deleted for SET1 or SPP1 genes relative to the intergenic region by ChIP; (C) Quantification of Rpb1 enrichment in WT, set1Δ and spp1Δ strains relative to the intergenic region by ChIP.
3′ORF and mORF stand for the 3′end and middle coding regions.
Figure 5.
Bye1 and TFIIS exhibit a synthetic growth defect in response to high concentrations of NaCl and caffeine.
Figure 6.
Bye1 positively regulates the GAL10 gene transcription: (A) GAL10 steady-state mRNA levels measured by RT-qPCR in WT and bye1Δ strains grown in a glucose- or galactose-supplemented medium; (B) Quantification of Rpb1 occupancy of GAL10 in glucose and after 3 hours of induction in galactose media in WT and bye1Δ strains by ChIP; (C) Quantification of Bye1-Myc occupancy of promoter and coding regions of GAL10 relative to RPO21-ORF under repressed (in glucose) and activated (in galactose) conditions by ChIP.