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Figure 2 | Molecular Cancer

Figure 2

From: Nuclear export regulation of COP1 by 14-3-3σ in response to DNA damage

Figure 2

COP1 S387 phosphorylation is essential for 14-3-3σ -induced COP1 nuclear export. (A) Cytoplasmic COP1 localization is compromised after DNA damage in 14-3-3σ-null cells. HCT116 14-3-3σ+/+ and HCT116 14-3-3σ-/-cells were treated with 10 Gy IR. Cytoplasmic and nuclear extracts were prepared 2 hr after irradiation. N, nuclear fraction. C, cytoplasmic fraction. (B) Cytoplasmic COP1 localization is reduced in 14-3-3σ knockdown cells after IR. HCT116 cells were stably transfected with two specific 14-3-3σ shRNA (1 & 2). Luciferase shRNA was used as a control (C). Cytoplasmic and nuclear extracts were prepared 2 hours after cells were treated with 10 Gy IR. (C) COP1 S387 mutant is less sequestered by 14-3-3σ in the cytoplasm in response to DNA damage. HEK 293 cells were transfected with Flag-COP1 (WT), or Flag-COP1 (387A), followed by infection with Ad-β-gal or Ad-14-3-3σ. Lysates were immunoblotted with anti-Flag. Cytoplasmic and nuclear extracts were prepared 2 hr after cells were treated with 10 Gy IR. (D) 14-3-3σ-induced COP1 cytoplasmic translocation is S387 phosphorylation-dependent. U2OS cells were transfected with the indicated plasmids. GFP-COP1 localization of live cell images were captured using a confocal microscope. Arrows indicate the representative location of COP1 in the nucleus. The bar graph shows the percentage of COP1 localized in the nucleus from each group. Scale bar, 10 μm.

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