Fig. 1

Progranulin promotes AKT and MAPK activation in mesothelioma cells. A Progranulin (PGRN) protein levels were analyzed by immunoblot in cell lysates and media conditioned from MeT-5A, NCI-H2052, NCI-H2452, NCI-H28 and MSTO-211H cells. B Progranulin levels in media conditioned from MeT-5A, NCI-H2052, NCI-H2452, NCI-H28 and MSTO-211H cells was measured by ELISA assay. N = 3, ± SD, ** p < 0.01, *** p < 0.001. C Levels of total and phosphorylated EphA2 (S897 and S901) were analyzed by immunoblot in cell lysates derived from MeT-5A, NCI-H2052, NCI-H2452, NCI-H28 and MSTO-211H cells serum-starved for 24 h. D Levels of total and phosphorylated EphA2 (S897), AKT and ERK1/2 were assessed by immunoblot in NCI-H2052, NCI-H2452 and MSTO-211H cells serum-starved for 24 h and then treated with 50 nM progranulin for the indicated time. E MSTO-211H cells were transfected with siRNA targeting progranulin or non-targeting control siRNA. At 8 h post-transfection cells were transferred into serum-free medium for 40 h and then analyzed by immunoblot for total and phosphorylated levels of EphA2 (S897), AKT, ERK1/2 and progranulin levels. F Levels of total and phosphorylated EphA2, AKT, ERK1/2 and progranulin were assessed by immunoblot in parental MSTO-211H, MSTO-211H cells with progranulin knock-out by CRISPR/Cas9 (GRN KO) and GRN KO MSTO-211H cells with reconstituted progranulin expression (left panel) or parental and progranulin overexpressing NCI-H2052 cells (right panel), serum-starved for 24 h. Progranulin levels were also analyzed in conditioned media from cells starved for 48 h. G MSTO-211H and NCI-H2052 cells were serum-starved for 24 h, pre-incubated with either the MEK1/2 inhibitor U0126 (10 µM) or the AKT inhibitor AKTi VIII (5 µM) for 2 h and then treated with the same concentrations of MEK1/2 and AKT inhibitors alone or in combination with 50 nM progranulin for 15 min. Levels of total and phosphorylated EphA2, AKT and ERK1/2 were determined by immunoblot