Abl family kinases bind and phosphorylate HS1. (A) Jurkat T cells were treated with or without pervanadate for 10 minutes, and lysates were subjected to affinity precipitation with the SH2 domains of indicated proteins. The precipitates were analyzed by immunoblotting with anti-HS1. (B) Ba/F3 pTET-ON p210 Bcr-Abl–inducible cells were treated with 1 μg/mL doxycycline for the indicated times. Bcr-Abl expression was assessed by immunoblotting of whole-cell lysates (WCL). HS1 was immunoprecipitated from cell lysates and phosphorylation was assessed by blotting for phosphotyrosine (Anti-P-Tyr). (C) A total of 10 μg of recombinant HS1 in 30 μL phosphorylation buffer was incubated with or without 0.5 μg of recombinant Arg or Abl kinases at 30°C for 10 minutes and immunoblotted with antiphosphotyrosine and anti-HS1.