FIG. 2.

SH3 domain is critical for SKAP55-dependent β1- and β2-integrin activation. (A) Schematic representation of the individual FLAG (F)-tagged SKAP55 mutants used in this study. (B) Jurkat T cells were transiently transfected with either empty vector (vector), FSKAP55, FSKAP55(Y3F), FSKAP55ΔN, FSKAP55ΔPH, FSKAP55ΔSH3, or FSKAP55(W333K). The expression of the mutants was analyzed by anti-FLAG Western blotting. (C) The above transfectants were either left unstimulated (non) or were stimulated for 30 min with CD3 MAb OKT3 (TCR). Cells were analyzed for adhesion to fibronectin or to ICAM-1 as described in the legend to Fig. 1C. Data are the average means and SE from three independent experiments. (D) Jurkat T cells overexpressing the individual FLAG-tagged SKAP55 mutants were lysed, and FLAG-tagged proteins were immunoprecipitated using an anti-FLAG MAb. Precipitates were analyzed by Western blotting for the presence of SKAP55 and ADAP. IP, immunoprecipitation.