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. 2014 Feb 17;33(8):862–877. doi: 10.1002/embj.201386064

Figure 1. Rad17 regulates HR repair in association with Rad51 recruitment to DSBs.

Figure 1

  1. Rad17 is required for HR repair in human cells. U2OS cells were transfected with the indicated siRNA for 2 days. HR assay was performed as previously described (Pierce et al, 1999). pGFP construct was transfected into cells to ensure similar transfection efficiency.
  2. Rad17 promotes Rad51 recruitment to DSBs. U2OS cells transfected with the indicated siRNA for 2 days were either left untreated or exposed to 5 Gy of IR. Immunostaining with anti-Rad17 and anti-Rad51 was performed. A cell containing 10 or more foci was considered as a foci-positive cell. The percentage of Rad51 foci-positive cells was plotted.
  3. Rad17 is required for γ-H2AX foci formation at DSB sites. U2OS cells transfected with the indicated siRNA for 2 days were either untreated or exposed to 5 Gy of IR. Immunofluorescence staining was performed with anti-Rad17 and anti-γ-H2AX. The intensity of γ-H2AX foci was analyzed using Image J software.
  4. Rad17 is required for γ-H2AX production. Inducible shRNA-mediated Rad17 knockdown cells (U2OS) treated with either Dox or DMSO for 2 days were either left untreated or exposed to 5 Gy of IR. Cell lysates were prepared at the indicated time points, and Western blots were performed as indicated. Dox, doxycycline; IR, ionizing radiation.

Data information: In (A–C): Error bars represent mean ± s.d. (n = 3).

Source data are available online for this figure.