Inducible expression of TEL/PDGFβR leads to constitutive activation of signaling pathways. Cells were incubated for 24 hours in the presence (+) or in the absence (–) of 2 μg/mL Tet. (A) Samples were directly removed for immunoblotting. (B-C) Cells were either removed directly for immunoblotting (+ serum) or were washed 3 times with 1 × HBSS and serum starved for 1 hour prior to preparation of cell lysates (– serum). Twenty or 40 μg cell extract was immunoblotted using the following antibodies: panel A and lower part of panel B, a polyclonal antibody against PDGFβR that recognizes the TEL/PDGFβR fusion; panel B upper part, 4G10 to detect tyrosine-phosphorylated proteins; and panel C phosphospecific antibodies recognizing activated forms of STAT5, PKB, ERK, JNK, and p38. The same immunoblots were stripped and reprobed with anti-STAT5, anti-PKB, anti-ERK, anti-SHP-2, or anti-p38 antibodies as loading controls, respectively. For comparison of phospho-p38 in the presence of serum, see Figure 7.