Relevant parameters for ATR activation upon a subset of genotoxic treatments
| Parameter | Approach | Figure | NT | MMC (200 nM) | APH (100 nM) | CPT (25 nM) | HU (0.5 mM) |
| Fork reversal | EM analysis | 3, B and C | −/+ | ++ | ++ | ++ | ++ |
| Fork slowing | DNA fiber spreading | 1 A | − | ++ | ++ | + | +++ |
| Impaired DNA synthesis | EdU incorporation (FACS) | 4 D | − | −/+ | ++ | + | +++ |
| ssDNA at forks | EM analysis | 2, A and B; S2; and S3 | − | + | + | + | + |
| Total exposed ssDNA | Native BrdU staining | 4 B | − | − | − | ++ | ++ |
| ATR signaling at forks | iPOND γ-H2AX | 6 B | − | −/+ | ND | ++ | +++ |
| ATR signaling total | WB pCHK1 | 4 A | − | − | − | + | +++ |
| ATR signaling total | IF/FACS γ-H2AX | 4, B–D | − | − | − | ++ | ++ |
| Parameter | Approach | Figure | NT | MMC (200 nM) | APH (100 nM) | CPT (25 nM) | HU (0.5 mM) |
| Fork reversal | EM analysis | 3, B and C | −/+ | ++ | ++ | ++ | ++ |
| Fork slowing | DNA fiber spreading | 1 A | − | ++ | ++ | + | +++ |
| Impaired DNA synthesis | EdU incorporation (FACS) | 4 D | − | −/+ | ++ | + | +++ |
| ssDNA at forks | EM analysis | 2, A and B; S2; and S3 | − | + | + | + | + |
| Total exposed ssDNA | Native BrdU staining | 4 B | − | − | − | ++ | ++ |
| ATR signaling at forks | iPOND γ-H2AX | 6 B | − | −/+ | ND | ++ | +++ |
| ATR signaling total | WB pCHK1 | 4 A | − | − | − | + | +++ |
| ATR signaling total | IF/FACS γ-H2AX | 4, B–D | − | − | − | ++ | ++ |
Parameters were assessed by different investigation methods, as displayed in the indicated figures. −/+, +, ++, and +++ indicate increasingly clear phenotypes. IF, immunofluorescence; NT, not treated; WB, Western blot.