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. 2021 Apr 16;296:100674. doi: 10.1016/j.jbc.2021.100674

Figure 4.

Figure 4

PLD2 dependency of SK1 translocation. WT HA-PLD2–inducible or K758R-inactive HA-PLD2–inducible Chinese-hamster ovary cells were transiently transfected with WT GFP-mSK1 construct and then treated with 2 μg/ml doxycycline for 24 h. A, photomicrographs of cells of 40× oil magnification overexpressing WT GFP-mSK1 detected with GFP. Representative results of three independent experiments. Also shown is a Western blot of cell lysates with the anti-HA antibody, detecting induced PLD2, and ERK-2 (loading control). B, the bar graphs represent the AUC of transfected WT GFP-mSK1 translocation (n = 5) and the percentage of cells containing translocated WT GFP-mSK1 (n = 3). ++++p < 0.001 doxycycline versus control and ∗∗∗∗p < 0.001 for WT PLD2 versus K758R-inactive PLD2 in doxycycline-treated cells (two-way ANOVA with Tukey's post hoc test). AUC, area under the curve; mSK1, mouse SK1; SK1, sphingosine kinase 1.