Fig. 3: The NZF domain of HOIL-1L contributes to LUBAC-mediated cell death protection in concert with the NZF domain of SHARPIN. | Cell Death & Disease

Fig. 3: The NZF domain of HOIL-1L contributes to LUBAC-mediated cell death protection in concert with the NZF domain of SHARPIN.

From: Synergistic involvement of the NZF domains of the LUBAC accessory subunits HOIL-1L and SHARPIN in the regulation of LUBAC function

Fig. 3

A, F HOIL-1L/SHARPIN DKO MEFs stably reconstituted with the indicated proteins were stimulated with TNF-α (2.5 ng/mL) and CHX (20 μg/mL) for the indicated times and analyzed by immunoblotting with the indicated antibodies. Data are representative of three independent experiments. B HOIL-1L/SHARPIN DKO MEFs stably reconstituted with the indicated proteins were stimulated with TNF-α (2.5 ng/mL) and CHX (20 μg/mL) for the indicated times after pretreatment with Z-VAD-FMK (10 μM) for 1 h, and analyzed by immunoblotting with the indicated antibodies. Data are representative of three independent experiments. C, D, G, H HOIL-1L/SHARPIN DKO MEFs stably reconstituted with the indicated proteins were stimulated with TNF-α (2.5 ng/mL) and CHX (20 μg/mL), and cell viability was continuously measured using the xCELLigence system. A representative image (C, G) and statistical analyses of the normalized cell index at 6 h (D, H). Data are shown as the mean ± s.d. of five independent experiments. P values were calculated by a one-way ANOVA with Tukey’s post hoc test. E HOIL-1L/SHARPIN DKO MEFs stably reconstituted with the indicated proteins were stimulated with TNF-α (2.5 ng/mL) and CHX (20 μg/mL) for the indicated times after pretreatment with Z-VAD-FMK (10 μM) for 1 h. Lysates were immunoprecipitated with anti-FADD antibody and immunoblotted as indicated. The experiments were repeated three times, independently, with similar results. Uncropped western blots are available for this figure in the Supplementary Material.

Back to article page