Domains involved in Tec/Dok association.

(A) The 293 cells were cotransfected with HA-Dok-1 and different Tec mutants. Tec immunoprecipitates were analyzed by Western blotting with anti-HA (top panel), anti-Tec (middle panel), and antiphosphotyrosine (lower panel). The TH and SH2 domains of Tec are crucial for the direct interaction with HA-Dok-1. (B) Schematic representation of the Tec mutants used and their Dok-1 interaction and phosphorylation properties. (C) Schematic representation of full-length HA-Dok-1, HA-DokΔY5, and HA-DokΔY1. Tyrosine residues (Y), the PH and PTB domains and total number of amino acids are indicated. (D) The 293 cells were cotransfected with Tec and different HA-Dok-1 deletion constructs. Lysates were incubated with anti-HA for expression control and with anti-Tec. Precipitates were analyzed with anti-HA. Although the HA-Dok deletion constructs are equally expressed (left panel), only full-length HA-Dok and a reduced amount of HA-DokΔY5 are detected in Tec precipitates (right panel), indicating that 2 domains in Dok-1 are involved in the binding to Tec.