Fig. 4
From: LSD1 is required for euchromatic origin firing and replication timing
LSD1 deficiency compromises replication initiation. a iPOND assays for LSD1 at replication forks or mature chromatin in HeLa and HEK293T cells. Pulldowns of control, nascent DNA (pulse), or mature chromatin (chase) were analyzed by western blotting with the indicated antibodies. b U2OS cells were pulse-labeled with EdU or followed by thymidine chase for in situ PLA assays. The efficacy of the antibodies was determined for each assay. PLA, red; biotin, green; DAPI, blue. Scale bar, 10 μm. Quantification of the PLA signals is shown on the right (n = 15 cells). c LSD1-deficient U2OS cells were pulse-labeled successively with IdU and CldU for DNA fiber assays. Distribution of replication forks is presented as beeswarm-box plots (n ≥ 200). The median fork rate for each experiment is shown. d IOD was measured on spread DNA fibers and presented as beeswarm-box plots (n ≥ 100). e Five classes of replication structures were defined (n ≥ 200). Left, representative classes of replication structures. Right, the percentage of the different classes is scored. f Synchronized U2OS cells were analyzed at different time points of release for EdU incorporation by flow cytometry. P-values are indicated
