Skip to main content
. 2015 May 27;11(7):975–994. doi: 10.1080/15548627.2015.1049800

Figure 2.

Figure 2.

RIPK1 protects melanoma cells from killing by TM or TG. (A) Whole cell lysates from Mel-RM and MM200 cells transduced with the control or RIPK1 shRNA treated with tunicamycin (TM) (3 μM) or thapsigargin (TG) (1 μM) for 16 h were subjected to western blot analysis of RIPK1, HSPA5, and GAPDH (as a loading control). The data shown are representative of 3 individual experiments. The numbers represent fold changes of HSPA5. (B) Mel-RM and MM200 cells transduced with the control or RIPK1 shRNA were treated with TM (3 μM) or TG (1 μM) for 48 h. Cell viability was measured by CellTiter-Glo assays (n = 3, mean ±SEM, *P < 0.05, Student t test). (C) Mel-RM and MM200 cells transduced with the control or RIPK1 shRNA were seeded onto 6-well plates at 2000 cells per well in medium containing TM (1 μM) or TG (0.3 μM). Twelve d later, cells were fixed and stained with crystal violet. The data shown are representative of 3 individual experiments. (D) Mel-RM and MM200 cells stably transduced with the control or RIPK1 shRNA were transiently transfected with a shRNA-resistant mutant form of RIPK1 (pCMV-MYC-mut-RIPK1). Twenty-four h later, whole cell lysates were subjected to western blot analysis of RIPK1 and GAPDH (as a loading control). The data shown are representative of 3 individual experiments. (E) Mel-RM and MM200 cells stably transduced with the control or RIPK1 shRNA were transiently transfected with a shRNA-resistant mutant form of RIPK1 (pCMV-MYC-mut-RIPK1). Twenty-four h later, cells were treated with TM (3 μM) or TG (1 μM) for another 48 h. Cell viability was measured by CellTiter-Glo assays (n = 3, mean ±SEM, *P < 0.05; Student t test). (F) Whole cell lysates from HEMn-MP melanocytes transfected with the pCMV-MYC or pCMV-MYC-RIPK1 were subjected to western blot analysis of RIPK1 and GAPDH (as a loading control). The data shown are representative of 3 individual experiments. (G) HEMn-MP melanocytes transfected with pCMV-MYC or pCMV-MYC-RIPK1 were treated with TM (3 μM) or TG (1 μM) for 48 h. Cell viability was measured by CellTiter-Glo assays (n = 3, mean ±SEM, *P < 0.05, Student t test).