Figure 4

Mature and primary miR-146a expression in EBV-infected, LMP1- and EBNA2-transfected U2932 cells. (a) U2932 cells were transfected with expression vector for LMP1 and EBNA2 individually. Stable LMP1 and EBNA2 expressing clones were analyzed for expression of these proteins by S12 and PE2 monoclonal antibodies, using immunoblotting. EBV-infected U2932 cells were also included in this experiment. β-Actin expression was verified to check protein loading. (b) miR-146a expression in U2932 LMP1 and EBNA2 transfectants with corresponding vector control cell lines and in EBV-infected U2932 clones was verified by northern blotting. LNA oligonucleotide corresponding to sequences complementary to miR-146a was used as a probe. U2 expression was used to verify RNA loading and quality. (c) Two separate RNA preparations of U2932 parentals and EBV-infected clones EBVGFP cl-A (EBNA2 negative) and cl-B (EBNA2 positive) were tested for pri-miR-146a levels (left panel). Each experiment was performed in triplicates. In the right panel, U2932 LMP1- and EBNA2-transfected clones were tested for pri-miR-146a.