Figure 4

Cross-tyrosine phosphorylation of PDGFRβ is not required for DRD4-mediated ERK1/2 activation. (A) Dose-response of DRD4- and PDGF-BB-mediated ERK1/2 phosphorylation. CHO/DRD4 (black square) or CHO/DRD4-PR (black triangle) cells were treated with different concentrations of dopamine or PDGF-BB for 5 min. (B) Effect of C-truncPDGFRβ on the phosphorylation of full-length PDGFRβ and ERK1/2 in CHO/DRD4-PR cells mediated by PDGF-BB and DRD4. CHO/DRD4-PR cells were transfected with C-truncPDGFRβ, or with lacZ as a control. IP: immunoprecipitation; IB: immunoblot. (C) DRD4-mediated phosphorylation of ERK1/2 in CHO/DRD4 was not blocked by C-truncPDGFRβ. (D) Requirement for PDGFRβ in the DRD4-mediated phosphorylation of ERK1/2 in C-truncPDGFRβ-transfected cells. Upper blot shows the phosphorylation of ERK1/2 in lacZ-transfected and C-truncPDGFRβ-transfected CHO/DRD4 cells that have been pre-treated with DMSO or 1 μM tyrphostin A9 prior to stimulation with dopamine. (B-D) The blots were stripped and re-probed with antibodies for PDGFRβ or ERK1/2 to verify that the total protein did not vary between lanes. In (C) and (D), the bar graphs show densitometric measurements of the phosphorylation signal from the respective experiments. The results are expressed as percentages relative to lacZ control and indicated as mean ± SEM. The number of experiments is indicated in parentheses.