Fig. 4: Transcriptional readthrough impacts expression of neighboring genes.

a Schematic of a gene pair with opposite transcriptional directions. GOI, gene of interest; NNG, nearest neighbor gene. The last 1 kb region of GOI was used to calculate ChrRNA-seq Overlapping Signal Score (CROSS) as described. b Expression changes of GOI as a function of distance from NNG. All GOI-NNG pairs were divided into five equally sized bins based on their distance. The number of genes used in each plot is indicated. P value (K–S test) for bin 1 vs. bin 5 comparison is indicated for HepG2 cells treated with 10 μM JTE-607. c CROSS as a function of distance between GOI and NNG. P value (K–S test) for bin 1 vs. bin 5 comparison is indicated for both HepG2 and HeLa cell data. d CDF curves of CROSS for genes regulated in HepG2 cells treated with 10 μM JTE-607. P values (K–S test) for comparisons between upregulated genes (red line) or downregulated genes (blue line) and not regulated genes (gray line) are indicated. e Heatmap showing gene expression changes based on CROSS (x-axis) and GOI-NNG distance (y-axis). Each gene was assigned to one of the 25 bins based on CROSS and distance values. The median of gene expression changes (log₂Ratio in HepG2 cells treated with 10 μM JTE-607) of each bin is represented by color. f Schematic indicating that collision of Pol II on two opposite strands leads to gene downregulation. g Schematic showing GOI and its upstream NNG having the same transcriptional direction. Calculation of ChrRNA-seq Intervening Signal Score (CRISS) is described. h As in b except that GOI-NNG pair is based on g. i. As in e, except that the 25 bins are based on GOI expression level (y-axis) and NNG expression level (x-axis). j Schematic indicating that readthrough Pol II from an upstream gene leads to read-in of downstream gene, causing the latter being upregulated. k CDF curves showing CRISS vs. gene regulation for lowly expressed genes (left) and all genes (right). P values (K–S test) for comparisons of upregulated genes or downregulated genes with not regulated genes (gray lines) are indicated. Source data are provided as a Source Data file.