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. 1999 Sep 14;96(19):10620–10625. doi: 10.1073/pnas.96.19.10620

Figure 3.

Figure 3

(A) PD98059 inhibits CA-induced activation of ERK p42/44, but not CA-induced STAT3 serine phosphorylation. T lymphoma cells were preincubated with or without a MEK inhibitor: PD98059 at 50 μM for 60 min, before incubation with CA (80 nM), OA (500 nM), or CyA (400 ng/ml) for 60 min, lysed and applied to SDS/PAGE, and immunoblotted with anti-phospho-p42/44 ERK, stripped and reblotted with anti-p42/44 ERK, anti-STAT3 pS727, or anti-STAT3. (B) Inhibition of staurosporine-sensitive kinases inhibits CA-induced serine phosphorylation of STAT3. In parallel experiments antigen-specific human CD4+ T cells were preincubated in either medium H7 at 200 μM, MEK inhibitor PD98059 at 50 μM (PD), or staurosporine (STA) at 100 nM for 60 min before incubation in medium, the PP1/PP2A-specific inhibitor CA at 80 nM or the PP2B-specific inhibitor CyA at 400 ng/ml for an additional 60 min. Cells were lysed and applied to SDS/PAGE as described in Materials and Methods, and immunoblotted with anti-STAT3 pS727 (Upper) and stripped and reblotted with anti-STAT3 (Lower).