Analysis of the association of CrkL with DOCK2 truncated mutants.

(A) Schematic structure of wild-type DOCK2 and the mutants designated dN (aa 939 to 1830), dCS (aa 1 to 515), INT1 (aa 386 to 593), INT2 (aa 586 to 883), dBE (aa 926 to 1478), and CT (aa 1640 to 1830). ND indicates not determined; P, proline-rich sequence; B, basic domain; and S, SH3 domain. (B) Association of CrkL and DOCK2 mutants were analyzed by immunoprecipitation. The 293T cells were transfected with pCAGGS-CrkL and the DOCK2 expression vector, and cell lysates were immunoprecipitated with anti-CrkL Ab and were analyzed by immunoblotting with anti-Flag monoclonal Ab (Ab; lanes 6-10). Protein expressions of DOCK2 mutants and CrkL were examined by anti-Flag mAb (top panel, lanes 1-5) and anti-CrkL Ab (bottom panel). Transfected plasmids were as follows: lanes 1 and 6, pCAGGS; lanes 2 and 7, pCXN2-Flag-DOCK2-dCS; lanes 3 and 8, pCXN2-Flag-DOCK2-dN; lanes 4 and 9, pCXN2-Flag-DOCK2-dBE; and lanes 5 and 10, pCXN2-Flag-DOCK2-CT.