Figure 8
Reduction of Set1 occupancy and RNA-binding activity in Set1YF/AA correlates with reduced H3K4me3 levels. (a) ChIP-seq and CRAC signals of PTH-Set1 (Set1) and PTH-Set1YF/AA (Set1 YF/AA) at representative genes. ChIP-seq bedGraphs were generated by normalization to 10 million mapped reads for each sample. CRAC signals were normalized as described above. (b) H3K4me3, me2, me1 occupancies at the indicated genes in W303 set1∆::TRP1 pRS415-nHTP-SET1 and SET1YF/AA strains. Levels of H3K4me3, me2 and me1 are normalized to total H3. Positions of the primers used for the ChIP-qPCR for each representative gene are indicated. Errors bars represent the s.d. from three independent experiments. (c) Recombinant chromatin template containing fully ubiquitylated H2B (H2Bub) was subjected to in vitro HMT assays with purified Set1C and Set1C YF/AA in the absence and presence of purified GAL10 RNA [0.1 (lanes 2 and 7), 0.2 (lanes 3 and 8), 0.5 (lanes 4 and 9) and 2 (lanes 5 and 10) molar ratio to Set1C. H3 methylated status was monitored by western blots with indicated antibodies.
