Transmembrane Protein 208: A Novel ER-Localized Protein That Regulates Autophagy and ER Stress (original) (raw)
Figure 6
TMEM208 regulates ER stress.
(A) U2OS cells were transfected with TMEM208 or pcDNA3.1 vector alone for 24 hours and cellular RNA were extracted for quantitative real-time PCR analysis of the genes indicated. (B) U2OS cells were transfected with shTMEM208 or shRNA vector for 72 hours, and relative mRNA levels of the indicated genes were analyzed by quantitative real-time PCR. The average value in the vector-transfected cells was normalized as 1. Data are the mean ± SD of results from three experiments (*P<0.05). (C and D) Western blot analysis of the protein levels of GRP78, ATG12-ATG5 conjugates and CHOP in U2OS cells treated as in (A) or treated as in (B). (E) U2OS cells were transfected with pcDNA3.1 vector or TMEM208 for 24 hours, then DTT (1 mM) and Baf. A1 were added to culture media for 4 hours as indicated. Cells were harvested and the levels of CHOP, P62 and TMEM208 were analyzed by western blot. (F) U2OS cells were transfected with shRNA vector or TMEM208 shRNA for 72 hours. DTT (1 mM) and Baf. A1 were added to culture media for the final 4 hours as indicated. Cells were harvested and the levels of CHOP and P62 were analyzed by western blot. Actin was detected as the protein loading control.