Figure 3. ROZA is specific for ZAP-70.

(A) WT J77cl20 or cells transiently expressing either ROZA or ROZA YF (5×10⁶ cells/point) were preincubated for 30 minutes with inhibitors (A. 10 µM PP2 or B 200 µM piceatannol) or medium alone, followed by a 2 minute activation with 5 mM freshly prepared PV. ROZA was immunoprecpitated with anti-GFP antibody. Upper panel: anti-phosphoTyr (4G10) western blot. Lower panel: anti-GFP western blot. (B) PV induces an increase in ZAP-70 activity in Jurkat T-cells expressing ROZA. No change in FRET ratio was observed in ZAP-70-deficient Jurkat T-cells (P116) cells expressing ROZA or in normal Jurkat T-cells transfected with the mutated probe (YF).