Efficient activation of STAT5 by EE-T-Y343, but not by EE-T-Y343F, in primary erythroid progenitor cells.

EE-T-Y343 and EE-T-Y343F mice were treated with TAP, and, at 72 hours after TAP withdrawal, erythroid splenocytes were prepared. Cells were cultured for 5 hours in the absence of cytokines and were exposed to EPO (±10 U/mL) or hEGF (±15 ng/mL) for 7.5 minutes. Nuclear extracts were prepared, and levels of STAT5 DNA-binding activity were assayed by electrophoretic mobility shift using a³²P-labeled element from the β-casein promoter. Extracts prepared from hEGF-stimulated EE-T-Y343 splenocytes were preincubated with 50-fold molar excess of self (S, β-casein) or irrelevant (I, NFκB) unlabeled oligonucleotide cassettes.