FIG. 4.
The GR LS7 and dimer mutants function like the wt GR in U2OS cells. U2OS-LS7 and U2OS-dim clones were generated as described in Materials and Methods. (A) The GR LS7 and dimerization mutants induce cell cycle arrest in U2OS cells. U2OS-GR(+) (wt GR), U2OS-LS7 (LS7), and U2OS-dim (dim) clones were seeded on day 0 into six-well plates in duplicate and cultured in the absence or presence of 100 nM Dex as indicated. The total numbers of viable cells were determined on days 2, 4, and 6 by the trypan blue exclusion method. The graph represents cell counts on day 6. (B and C) The GR LS7 (B) and dim (C) mutants repress the expression of CAS, Bcl2, and CDK4. U2OS cells expressing the wt GR or the LS7 or dim mutant were cultured in the absence or presence of 100 nM Dex for 40 h and harvested, and the expression of CAS, Bcl2, CDK4, and ERK was assessed by immunoblotting as described in Materials and Methods. Similar results were obtained with at least three independent clones expressing each GR mutant.