Constitutive tyrosine phosphorylation of STAT3 in leukemia cells expressing Asp816 mutant c-Kit.
Total cellular lysates (20 μg) from indicated cells unstimulated or stimulated with SCF (40 ng/mL) for the times shown were analyzed by Western blotting. (A) The upper panels show the antibody against tyrosine- phosphorylated STAT3 reacting to 2 cellular-phosphorylated proteins. The top band was confirmed to be STAT3 by using the antibody against the internal domain of the protein, which recognizes both STAT3 and STAT3β, as shown in the lower panels using the stripped blots from the upper panels. The lower band observed as part of the doublet recognized by the antibody against tyrosine-phophorylated STAT3, could not be definitively identified as STAT3, 1, or 5. Size markers are shown in kilodaltons. (B) Blots in the upper panels were probed with the antibody against tyrosine- phosphorylated STAT5. Blots from the upper panels were stripped and reprobed with the antibody against STAT5 to illustrate the total STAT5 protein, as shown in the lower panels.
