Figure 4
From: SGO1 is involved in the DNA damage response in MYCN-amplified neuroblastoma cells
SGO1 knockdown and MYCN overexpression induce DNA damage that is repaired primarily by NHEJ.
(a) Immunofluorescence of γ-H2AX and DAPI staining of IMR32 (MYCN-amplified) and SK-N-AS (MYCN-single copy) cells infected with non-targeting or SGO1-specific shRNA. Images were acquired 3 days after infection. (b) Percentage of γ-H2AX-positive cells in the cell lines shown in (a), as well as in NB39 (MYCN-amplified) and SH-EP (MYCN-single copy) neuroblastoma cells. Data represent means ± SE of three independent repeats. (c) DSB formation in SGO1-knockdown MYCN-overexpressing U2OS cells expressing EYFP-53BP1 and ECFP-Geminin 48 hrs after virus infection. Nocodazole (0.9 μg/ml) was added for 16 hrs to arrest cells in G2/M phase and cells were counterstained with DAPI. (d) Percentage of 53BP1-positive cells relative to the total number of Geminin-positive cells in (c). Data are expressed as the mean ± SE of four independent experiments. (e) NHEJ efficiency in SGO1-knockdown MYCN-overexpressing H1299dA3-1 #1 cells and HR efficiency in SGO1-knockdown MYCN-overexpressing DR-U2OS cells. NU7026 (a DNA-PK inhibitor) was used as a negative control for the NHEJ assay. shBRCA1 and shBRCA2 were used as a negative control for the HR assay.
