PLD synergistically enhances RET/PTC-mediated transcriptional activation of STAT3. HEK293 cells were transfected with various combinations of plasmids encoding an empty vector, STAT3, RET/PTC1, RET/PTC3, flag-tagged wild-type (wt)- or lipase inactive mutant (KRM)- PLD2 and m67 luciferase reporter plasmid. After 24 h transfection, the cells were lysed and the luciferase activities were measured as described in Methods, and the activity of each sample was normalized according to Renilla luciferase activity (upper panel). Results show mean ± SE of at least of three independent experiments. * P < 0.05, when compared with STAT3 alone-expressed group. # P < 0.05, when compared with STAT3 and RET/PTC1- or RET/PTC3-expressed group. To confirm the expression level of each plasmid in transfected cells, the total cell lysates were analyzed by SDS-PAGE and Western blotting with antibodies against flag and RET (lower panel).