Fig. 2

TSPAN18 maintains STIM1 stability via inhibiting ubiquitination in PCa. a Western blot (WB) analysis of STIM1 expression in DU145 and PC-3 cells transiently transfected with indicated siRNA. b WB analysis of STIM1 expression in TSPAN18-overexpressing DU145 and PC-3 cells. c-d The protein level of STIM1 in DU145 and PC-3 cells transfected with scramble or si-TSPAN18 were monitored by WB at indicated times after cycloheximide (CHX, 20μg/mL) (c) and were quantified by ImageJ software (d). e-f the protein level of STIM1 in TSPAN18 overexpressing DU145 and PC-3 cells by WB at indicated times after cycloheximide (e) and were quantified by ImageJ software (f). g WB analysis of lysates from DU145 or PC-3 cells transfected with scramble or si-TSPAN18 followed by treatment with DMSO, MG132 (10 μg/ml) or Baf for 4 h. Baf: Bafilomycin. h-i Coimmunoprecipitation (Co-IP) analysis of ubiquitination of STIM1 in TSPAN18-knockdown (h) or TSPAN18-overexpressing (i) DU145 and PC-3 cells co-transfected with Myc-STIM1 plasmid and HA-Ub plasmid. The values are expressed as the mean ± s.d. of three independent experiments. *p<0.05, **p<0.01, ***p < 0.001, Student’s t test