Figure 4

RET/PTC enhances tyrosine phosphorylation of PLD2, but it does not affect the enzyme activity of PLD. (A) HEK293 cells were transiently transfected with mock vector, flag-PLD2, RET/PTC1 expression plasmids, treated with pervanadate (50 μM) for 30 min, and subjected to immunoprecipitation with anti-flag antibody. Immunoprecipitate was analyzed by Western blotting with anti-phosphotyrosine specific antibody (4G10), and the efficiency of immunoprecipitation was measured with anti-PLD2 antibody. (B) NIH3T3 cells were transiently transfected with mock vector and RET/PTC1 expression plasmids, treated with pervanadate (50 μM) for 30 min. PLD activity was measured as described in the legend of Fig. 3A. Radioactivity of PtdBut was expressed in percent of radioactivity of total phospholipids. * P < 0.05, when compared with empty vector transfected group. Each bar shows mean ± SE of at least of three independent experiments.