Figure 7.
pS273R interacted with IKKε and disturbed the interaction between IKKε and STING. (a–c) HEK293T cells in 6-well plates (8 × 105 cells/well) were co-transfected with 1 μg each plasmid as indicated for 48 h and then the cells were harvested and subjected for Co-IP using the indicated antibodies and subsequent Western blot analysis. (d) Increasing amounts of pS273R plasmid (1 μg, 2 μg) were co-transfected with 1 μg STING-mCHERRY and 1 μg IKKε-MYC into 293T cells for 48 h and then cells were harvested and immunoprecipitated with anti-mCHERRY antibody and subjected to Western blot analysis. (E) PAM cells in 24-well plates (3 × 105 cells/well) were co-transfected with STING-GFP (0.5 μg) and IKKε-MYC (0.5 μg), with STING-GFP (0.5 μg) and pS273R-HA (0.5 μg), with IKKε-MYC (0.5 μg) and pS273R-HA (0.5 μg) for 24 h, and then cells were fixed and examined for cellular co-localization by con-focal microscopy. the co-localizations in multiple vision fields were analyzed using Image J, and the Pearson coefficient values from 10 positive cells were graphed and shown on the right. the value of Pearson coefficient reflects the level of co-localization, with 1 representing 100% co-localization.