P stat5a

For protein expression of p-STAT5A and total-STAT5A, cells were lysed and whole cell lysates were prepared using radioimmunoprecipitation assay buffer (Sigma-Aldrich) supplemented with protease and phosphatase inhibitors. Total cell lysate (20 µg) was separated by SDS-polyacrylamide gel electrophoresis according to the manufacturer’s instructions (Life Technologies) and transferred to PVDF membranes. Western blot analysis was then performed using p-STAT5A (1:1000, Cell Signaling Technology, Danvers, MA) and t-STAT5A (1:1000, Cell Signaling Technology); secondary α-rabbit or α-mouse antibodies (1:2000, Jackson ImmunoResearch, West Grove, PA) were used and proteins were visualized using the SignalFire ECL Reagent (Cell Signaling Technology) on an Odyssey Fc Imaging System (LI-COR, Lincoln, NE). GAPDH (Santa Cruz Biotechnology, Inc., Dallas, TX) was used a loading control; immunoblots were performed a minimum of three times on samples collected from different experiments. Uncropped and unprocessed blots are provided in the Data Source file.

We used the following antibodies for immunoblotting: BCR-ABL1 (#2802), pBCR-ABL1 (#2861), BIM (#2819), cleaved CASPASE 3 (#9661), STAT5A (#9310), pSTAT5A (#9359), PARP (#9542), pERK (#4377), ERK (#9102), MCL-1 (#4572) (all from Cell Signaling Technology), Lyn (G-7, Santa Cruz, USA), pLYN (Epitomics, USA), BCL-2 (AbCam, UK) and β-actin (#AC-15, Sigma, USA). The antibody dilutions used were 1 in 1,000; except for β-actin (1 in 5,000). HRP-conjugated secondary antibodies were specific to rabbit (Sigma) or mouse IgG (Santa Cruz biotechnology). The protein bands on the membrane were visualized using the Western Lightning chemiluminescence reagent (pERKinElmer, USA).