Activities of EE483, EE372, and EENb2 chimeras in mediating induced SKT6 cell hemoglobinization and globin expression. (A) In SKT6 cells stably expressing the above chimeric receptor forms (see Fig 1), hemoglobinization as induced by EGF first was analyzed. SKT6-EE483, -EE372, and -EENb2 cell lines (3 sublines for each designated by open, closed, and shaded histograms, respectively) were exposed to either EGF (15 ng/mL) or Epo (10 U/mL) and, at 72 hours of cytokine exposure, hemoglobin-positive cells were stained with 2, 7 diaminofluorene (DAF) and scored (>200 cells per sample). Graphed are mean frequencies of DAF-positive cells ± standard deviations (n = 3). (B) EGF-induced globin expression in SKT6-EE483, -EE372, and -EENb2 cell lines. Cells were exposed to EGF (15 ng/mL) or Epo (10 U/mL) for the indicated intervals (0, 48, and 72 hours), and levels of globin expression were assayed by direct Western blotting of cell lysates using purified antibodies to murine hemoglobin.