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Figure 3 | The EMBO Journal

Figure 3

From: Regulation of RNA polymerase II processivity by Spt5 is restricted to a narrow window during elongation

Figure 3The alternative text for this image may have been generated using AI.

Spt5 depletion causes a redistribution of actively engaged Pol II

A. Spt5dep/WT ratio heatmaps with static binning spanning the entire length of all expressed minus‐strand genes (n = 3,473) up to 100 kb ordered by increasing gene length. Read densities are in reads per million (RPM). The arrows within the plots indicate the point at which the WT and Spt5dep tracks intersect in the TSS‐proximal gene bodies (˜15–20 kb).

B. GROseq and Pol II occupancy at a long gene (Camk2d) and a short gene (Mafk) in WT and Spt5dep cells visualized in the UCSC genome browser. WT and Spt5dep tracks are overlaid in blue and red representing WT and Spt5dep tracks, respectively, and black representing the overlap between them. The insets provide magnified views of the gene body region.

C. Upper panel shows a composite metagene plot of Spt5dep/WT GROseq ratio of all expressed genes < 100 kb (n = 3,473). The dotted vertical lines demarcate the TSS +13 kb to +23 kb region magnified in the lower panel. In the lower panel, a zoom‐in view of the Spt5dep/WT GROseq ratio is shown for the TSS +13 kb to +23 kb region of all expressed genes > 30 kb (n = 2,094). The red line at TSS +17 kb highlights the transition region where the average Spt5dep GROseq signal decreases below the WT GROseq signal.

D. Analysis of the Spt5dep/WT GROseq ratios at three different locations within genes for all expressed genes > 50 kb (n = 1,037). As shown in the upper panel, each region is 8 kb long and is defined as follows: front region from TSS +2 to +10 kb (A), middle region from TSS +40 to +48 kb (B) and the back region from TTS −10 to −2 kb (C). The upper panel depicts the three regions schematically and the box plot in the lower panel shows the distribution of ratios for each region. Asterisks indicate P‐value < 2.2 × 10−16. ns, not significant calculated using the unpaired t‐test.

E. Identical analysis as in (D) above using genes > 50 kb but divided into three groups: all genes (n = 1,037), genes which have a 5′ UTR intron (n = 535) and genes without a 5′ UTR intron (n = 397). In all three gene body segments (A–C), no statistical significance was reached between the three gene categories.

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