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. 2018 Jan 24;14(2):347–358. doi: 10.1080/15548627.2017.1407889

Figure 2.

Figure 2.

PolyUb(WT)-FC puncta were visualized after NCS and LLOMe treatment. (A) Visualization of PolyUb-FC puncta after NCS stimuli. HEK 293T cells were transfected with a plasmid encoding PolyUb(WT)-FC, PolyUb(K63)-FC and PolyUb(K0)-FC. After 20 h, cells were stained with anti-K63 polyubiquitin antibodies and analyzed by immunofluorescence microscopy. Images were acquired using a confocal laser microscope (FV10i, Olympus). Cells were treated with 5 ng/mL NCS for the last 180 min. Scale bars: 20 µm. We counted cells with more than 10 foci in the nuclear area. In all, 100 cells from each indicated strain were analyzed. Data are representative of 3 independent experiments. Error bars indicate standard deviations. (B) Dynamic visualization of PolyUb-FC puncta after LLOMe stimuli in living cells. HEK 293T cells were transfected with plasmids encoding PolyUb(K0)-FC and PolyUb(WT)-FC. After 17 h, cells were treated with 500 µg/mL LLOMe. Images were acquired using an FV10i confocal laser microscope every 3 min. Data are representative of 4 independent experiments. Scale bars: 10 µm. Data are representative of 2 independent experiments.