TCR-induced tyrosine phosphorylation of SLP-76 and LAT, and the association of LAT with SLP76 and Grb2 are not inhibited by RosA treatment.

Jurkat cells were preincubated with 30 μM RosA for 4 hours and stimulated for the indicated periods of time (A) or 2 minutes (B). Cells were then lysed, precleared with rabbit serum agarose, and immunoprecipitated with rabbit polyclonal anti–SLP-76 (5 μg/mL) (A) or rabbit polyclonal anti-LAT antibodies (5 μg/mL) (B), respectively. The immunoprecipitates were resolved by SDS-PAGE and immunoblotted with antiphosphotyrosine mAb (0.5 μg/mL) (top panels) or specific antibodies (0.5 μg/mL) (middle and bottom panels) as indicated. Representative data from more than 3 independent experiments are shown.