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. 2021 Apr 15;40(11):e99692. doi: 10.15252/embj.201899692

Figure 1. Targeting USP7 triggers mitotic signaling throughout the cell cycle.

Figure 1

  • A
    HTM of EdU levels per individual nucleus in response to USP7 inhibition in HCT‐116 cells treated with 50 μM P22077 for 1–2 h or 25 μM BAY 11‐7082 for 1–2 h, or with DMSO as a control.
  • B
    Western Blot (WB) showing the levels of USP7, MPM‐2, histone H3S10P, and histone H3 in whole nuclear extracts of HCT‐116 cells treated with 50 μM P22077 for 2–4 h (USP7i), 10 μM RO3306 for 8 h (CDK1i), or DMSO as a control. Ponceau staining is shown as a loading control.
  • C
    Flow cytometry profile illustrating the levels of histone H3S10P (left) and MPM‐2 (right) in HCT‐116 cells in control conditions (control, black) and after treatment with 50 μM P22077 for 4 h (USP7i, red). DNA content was measured with propidium iodide (PI).
  • D, E
    HTM‐mediated quantification of MPM‐2 (D) and H3S10P (E) levels per individual nucleus in U2OS cells treated with DMSO as a control or 50 μM P22077 (USP7i) for 4 h. Nuclei were stained with DAPI (blue). Scale bar, 30 μm. The quantification is shown to the right of representative images for each analysis (***P < 0.05; t‐test). Gray lines indicate mean values.
  • F
    Flow cytometry profile representing the levels of histone H3S10P and MPM‐2 in HCT‐116 cells transfected with two different siRNA against USP7 (red; si#1 and si#2) or with a non‐specific siRNA (control, black), for 96 h. DNA content was measured with propidium iodide (PI).
  • G
    WB illustrating the depletion of USP7 in the HCT‐116 cells used in (F). Experiments were repeated three times, and one representative result is shown.