Downregulation of the expression of MHC class II and CD74 in LCLs. CD19-positive B cells were seeded in a 12-well plate at a density of 1 × 106 cells per well and infected with EBV. RNA and proteins were harvested at the time points indicated. (A) The expression of HLA-DRA, HLA-DRB1, and CD74 transcripts was measured by RT-Q-PCR. The relative fold of the transcripts was normalized to uninfected B cells with the corresponding β-actin mRNA. This is a representative result from 6 independent experiments from anonymous donors. (B) Protein expression of MHC class II, CD74, EBNA1, LMP2A, and β-actin was detected by western blotting. Detection of β-actin served as an internal control. (C) BJAB cells were transfected with EBNA1 or BARF0 expression plasmids or infected with EBER1, EBER2, LMP1, LMP2A, or Zta lentiviruses. Expression of MHC class II, CD74, EBNA1, EBER1, EBER2, BARF0, LMP1, LMP2A, and Zta transcripts in the transfectants and lentiviruses-infected cells were analyzed by RT-PCR. β-actin was detected as an internal control. (D) Akata and BJAB cells were infected with LMP2A-expressing lentiviruses. Cell lysates were harvested and the expression of MHC class II, CD74, and LMP2A was detected by western blotting. β-actin was detected as an internal control. (E) BJAB cells were infected with various doses of LMP2A-expressing lentiviruses. At day 5 postinfection, cell lysates were harvested for the detection of MHC class II, CD74, and LMP2A by western blot analysis. β-actin was detected as an internal control. The experiment was performed 3 times, and 1 representative is shown.
