FIG. 5.
Stat3 and Stat3 dominant negative mutants do not affect v-ras transformation or overall tyrosine phosphorylation by v-src. (A) NIH 3T3 cells (5 × 105 cells in a 35-mm-diameter dish) were transfected (as in the legend to Fig. 2A) with 500 ng of v-ras (pRSV Ha-rasLeu61) (11) in conjunction with 1 μg of RcCMV, Stat3, Stat3Y, Stat3DB, or Stat3S. Twenty hours after transfection, the cells were fed with DMEM plus 10% BCS. Twenty-four hours later, the cells were trypsinized and plated (5 × 105 cells) into soft agar. Seventeen days later, colonies were counted. Each bar represents the average of four independent transfections. (B) NIH 3T3 cells were transfected (as in the legend to Fig. 2A) with 500 ng of v-src and 1 μg of RcCMV-based Stat3 constructs. Whole-cell extracts were isolated, and 100 μg per lane was analyzed by Western blotting with an antiserum to phosphotyrosine (PY20).