Fig. 4

miR-346 Induces Genome-Wide Double Strand DNA Breaks at Transcription Start Sites. A Schematic illustration of INDUCE-seq protocol, B Quantification of number of ds DNA breaks identified per ng of sequencing library, C Genome browser comparison of number of breaks per kb across chromosome 11 for C42, C42/NC, C42/miR-346 and C42/shNORAD cells treated ± Dox. Peak height corresponds to number of DNA breaks. Similar results were obtained for other chromosomes. D, E Integration of miR-346 and shNORAD dsDNA break coordinates with D) annotated transcription start sites (TSSs - hg19 refseq gene list) and E) PC ChIP-seq-identified CTCF, ZFX and histone H3K27ac sites. F) top 1000 genes expressed in C42 cells show greater enrichment of miR-346-induced dsDNA breaks at their TSSs (dark red) compared to total C42-expressed genes (yellow), G enrichment of miR-346 dsDNA breaks at TSSs of miR-346-downregulated (dark blue), but not –upregulated (dark green), genes. H Enrichment of miR-346 DSBs at (i) ChIP-seq-identified AR binding sites (LNCaP cells - pink) and (ii) canonical AREs (consensus ARE sequence from JASPAR [http://jaspar.genereg.net/]- blue). miR-346 DSB enrichment at TSSs from hg19 refseq gene list is shown for reference. J, K, L Integration of miR-346 dsDNA break coordinates with PC ChIP-seq-identified binding sites for (J) PC-implicated TFs/ pioneer factors cMyc (GSM1907204), CTCF (GSM2827203), ETV1 (GSM1145322), GATA2 (GSM941195), FOXA1 (GSM1863005), HOXB13 (GSM2480818), NKX3.1 (GSM989640) and POL2RA (GSM696843) (K) AR-related Nuclear Receptor family members PR (GSM50151), ESR1 (GSE43988) and NR3C1 (GR) (GSM759669) and (L) neuroendocrine PC-associated TFs nMyc (GSM2305251), EZH2 (GSM2305255) and RUNX2 (GSM838400). M Western blot analysis of phospho-Ser5-RNA PolII protein levels in C42 cells transfected with miR-346 or NC (20 nM) for indicated durations. GAPDH was used as a control for loading and a representative image is shown. See also Fig. S11