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Fig. 2 | BMC Research Notes

Fig. 2

From: Fanconi anemia core complex-dependent HES1 mono-ubiquitination regulates its transcriptional activity

Fig. 2

HES1 mono-ubiquitination is not required for foci formation. a HES1K109E mutation does not impair MMC-induced HES1 foci formation. HeLa cells transfected with HA-tagged HES1 or HES1K109E coding vectors were processed for immunofluorescence 16 h following treatment with MMC (120 ng/ml). Cells were double-stained with anti-HA and anti-FANCD2 antibodies (mouse anti-HA at 1:500; rabbit anti-FANCD2, Novus Biologicals NB100-182 at 1:2000) followed with secondary antibodies (Goat anti-mouse Alexafluor-555, ThermoScientific A-32727 and goat anti-rabbit Alexafluor-488, ThermoScientific, A-11008). Cell nuclei were labelled with TO-PRO-3 Iodide stain (1:400, ThermoScientific, T3605). Cells were visualized at 100× magnification. b HES1 lysine 109 plays a role in MMC-induced activation of the HES1 promoter. COS-1 cells transfected with pHES1pro-Luc and HES1 or HES1K109E coding vectors (0.034 μg) were treated with 120 ng/ml MMC. Empty vectors were used as controls. Experiments were done twice in triplicates. 2-way ANOVA with Bonferroni correction test, *p < 0.01 and ***p < 0.0001, as compared to Empty; ###p < 0.0001, as compared to Untreated

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