Me180 cells expressing Stat3-C. (A) Stable clones of
Me180 cells were isolated expressing either a control plasmid or a
Stat3-C expressing plasmid. The clones were maintained in G418
containing media. Nuclear extracts were isolated from these clones and
one example is shown here. Ten micrograms of protein from each cell
line (3T3, 3T3–3C, Me, Me3-C) was resolved on an SDS polyacrylamide
gel, transferred to nitrocellulose, and probed with anti-FLAG antisera.
Two micrograms of nuclear extract from the same cell lines (see above)
were incubated with radiolabeled m67 and resolved on a nondenaturing
EMSA gel. (B) Parental Me180 cells (Me) and Me180
expressing Stat3-C cells (Me 3-C) were either left untreated (serum
containing media) or treated with IFNγ (0.5 ng/ml) for 0.5, 1, 2,
or 6 h. Nuclear extracts were isolated from these, incubated with
radiolabeled m67, and resolved on an EMSA gel.