Figure 3

ER stress transcriptionally increases PrP levels in MCF-7 cells. One representative Western blot from at least three independent experiments is shown in A-B. (A) Western blots of PrP with the 3F4 antibody, BiP, and β-actin in protein extracts from MCF-7 cells treated 18 hrs with increasing concentrations of Brefeldin A, Tunicamycin, or Thapsigargin. (B) Western blot of PrP (3F4) and β-actin in protein extracts from MCF-7 cells incubated for 18 hrs in the absence or presence of serum or transfected with pCep4β-PrP (pC-PrP). (C) PrP, XBP1, sXBP1 and β-actin RT-PCR cDNA amplicons from cells treated for 6 hrs. The ratio of PrP over β-actin was calculated from three independent experiments. (D) Western blot of various ER stress-regulated proteins in MCF-7 cells treated 18 hrs with BFA, TM or Thps. (E) Western blot of PrP with the 3F4 PrP, CHOP and β-actin antibodies in protein extracts from MCF-7 cells treated 18 hrs with DMSO (Ctl) or ER stressors in the presence or in absence of cycloheximide (CHX) or actinomycin D (Act D). The immunoreactive band at 37 kDa was not consistently detected with the anti-PrP 3F4 antibody suggesting a non-specific band (ns).