FIG. 6.

Phosphorylation of tyrosine 705 of Stat3 by IGF-I. The cell lines are indicated above the panels. (A) The times (in minutes) are times after placing the cells in serum-free medium (SFM) and stimulation with IGF-I. The lysates were immunoprecipitated with an antibody to Stat3, and the blots were developed with a phosphoantibody to tyrosine 705 of Stat3. The blots were then reprobed with an antibody to Stat3. (B) Western blots on lysates of the same cell lines. The blots were probed directly with an antibody to Y705 (upper row) and then reprobed with anti-Stat3 antibody (lower row). In the experiment on the right, G-CSF was used as control for the phosphorylation of Y705. (C) A mutation at Y950 of the IGF-IR abrogates the detection of Y705 phosphorylation of Stat3, regardless of the presence or absence of DNStat3 (left). The right panel shows that Y705 phosphorylation of Stat3 is not abrogated in the same cell lines stimulated with G-CSF.