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. 2022 Sep 15;82(18):3382–3397.e7. doi: 10.1016/j.molcel.2022.07.011

Figure 3.

Figure 3

Genes within MiDAS sites are transcribed in early S-phase

(A) Top row: MiDAS-seq profiles (σ value) at representative genomic regions in BRCA2-deficient (−BRCA2) H1299 cells. Bottom row: EU-seq profiles (σ value) for nascent transcription at the same genomic sites, detected 100 min after release from single thymidine block (green, forward direction of transcription; red, reverse). Replication initiation profiles are shown in gray. RT, replication timing; Ge, genes; IGe, intergenic regions. Bin resolution, 10 kb.

(B) Average EU-seq signal for nascent transcription (yellow) and MiDAS-seq signal (black), centered across MiDAS peaks (n = 150) identified in BRCA2-deficient (−BRCA2) H1299 cells. Span of genomic region, 200 kb.

(C) BRCA2-proficient (+BRCA2) and -deficient (−BRCA2) H1299 cells were treated with 75 μM DRB for 4 h, pulse-labeled with EdU for 30 min. QIBC scatterplots of DNA content versus EdU mean intensity show RAD51 foci number in color scale. Boxes indicate cell populations used for quantification. Representative images of RAD51 foci (green) and DAPI staining (blue) for the 4 conditions are shown. DAPI, 4′,6-diamidino-2-phenylindole. Scale bars, 10 μm.

(D) Quantification of cells treated as in (C) and having ≥7 RAD51 foci per cell. Graph shows data from one experiment representative of n = 2 independent experiments.