Figure 5

Pol I, II, and III transcription in AC16 cells in response to TNFα. AC16 cell nuclei were treated with α-amanitin for 15 min. prior to GRO-seq analysis. The final read density for each gene was normalized to the total reads obtained in each condition. A) Effect of TNFα on the fraction of GRO-seq reads mapped to rDNA repeats (Pol I; green line), RefSeq genes (Pol II; red line), and tRNA genes (Pol III; blue line) over the time course of TNFα treatment. B) Heatmap showing the expression of 20 tRNA transcripts upregulated during a time course of TNFα treatment. C) Browser track representation of GRO-seq reads mapped to rDNA repeats (GenBank U13369.1) in 1 kb bins over a time course of TNFα treatment. D) Scheme for the GRO-seq experiments with α-amanitin showing the expected effects on Pol I, II, and III transcription. E) Relative change in GRO-seq reads at rDNA repeats in control and α-amanitin-treated AC16 nuclei. F and G) Metagene representations of the average number of GRO-seq reads distributed around tRNA (F) and RefSeq (G) gene TSSs in control and α-amanitin-treated AC16 nuclei. H) Fraction of different types of uniquely mapped transcripts transcribed by Pol II or other RNA polymerases, as revealed by α-amanitin treatment. I) The number of annotated, short, non-coding transcripts transcribed by Pol II or other RNA polymerases, as revealed by α-amanitin treatment. J) Metagene representations of the average GRO-seq read distributions ± 4 kb around the TSSs of 739 non-Pol II transcripts identified using α-amanitin. K) Histogram showing the length distribution of all 739 non-Pol II transcripts from (J). L) Pie chart showing the genomic distribution of the genes encoding all 739 non-Pol II transcripts from (J). M) Metagene representations of the average ChIP-seq read distributions ± 500 bp around the TSSs for RPC155 (Pol III subunit) in K652 cells (left), TFIIIC in K562 cells (middle), and CTCF in human cardiomyocytes (HCM) (right) relative to the TSSs (± 500 bp) of the identified non-Pol II transcripts.