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. 2021 Sep 24;41(10):e00171-21. doi: 10.1128/MCB.00171-21

FIG 5.

FIG 5

ICP4 regulates promoter proximal Pol II pausing at host cell genes activated by HSV-1 infection but not at genes repressed by infection. (A) At cellular genes activated by the WT virus there is a strong decrease in the pausing index that is mostly recovered in the absence of ICP4. Plotted is a cumulative distribution of the log2 pausing indices (gene body RPK/promoter proximal RPK) for the 463 genes activated by WT virus infection for the mock, WT, and n12 conditions. P values were determined by using a two-sided Kolmogorov-Smirnov test: mock versus WT, P < 2.2 × 10−16; mock versus n12, P < 6.9 × 10−9; and WT versus n12, P < 2.2 × 10−16. (B) The decrease in the pause index at host cell activated genes is primarily due to a large increase in Pol II occupancy in the gene bodies after infection. Shown are box plots comparing promoter proximal (positions +1 to +250) RPK and gene body (+250 to the 3′ end) RPK at the 463 WT-activated genes, across the three experimental conditions. P values were determined by an unpaired Mann-Whitney test (*, P < 0.050; **, P < 1.0 × 10−10; ***, P < 2.2 × 10−16). (C) ICP4 regulates promoter proximal pausing at genes dependent on ICP4 for activation. Plotted is a cumulative distribution of the log2 pausing indices (gene body RPK/promoter proximal RPK) for the 294 genes dependent on ICP4 for activation upon WT infection for the mock, WT, and n12 conditions. P values were determined by using a two-sided Kolmogorov-Smirnov test: mock versus WT, P < 2.2 × 10−16; mock versus n12, P < 0.0050; and WT versus n12, P < 2.2 × 10−16. (D) At cellular genes repressed by the WT virus, there is a moderate increase in the pausing index that occurs independent of the presence of ICP4. Plotted is a cumulative distribution of the log2 pausing indices (gene body RPK/promoter proximal RPK) for the 626 genes repressed by WT virus infection for the mock, WT, and n12 conditions. P values were determined by using a two-sided Kolmogorov-Smirnov test: mock versus WT, P < 9.3 × 10−10; mock versus n12, P < 2.8 × 10−9; and WT versus n12, P = 0.28. (E) The increase in the pause index at cellular repressed genes is dominated by a decrease in the gene body levels of Pol II after infection. Shown are box plots comparing promoter proximal (positions +1 to +250) RPK and gene body (positions +250 to the 3′ end) RPK across the three experimental conditions for the set of WT-repressed genes. P values were determined by an unpaired Mann-Whitney test (*, P < 0.05; ***, P < 2.2 × 10−16).