FIG. 8.

Role of actin binding activity of Fli-I gelsolin-like domains in coactivator function. (A) Binding of Fli-I Gel.A domain (wild type and mutants) to actin-like molecule BAF53 and to ER. Vectors encoding the human Fli-I gelsolin-like Gel.A fragment (amino acids 495 to 827) and two point mutants (E585K and G602S) (2.5 μg) were transfected into Cos-7 cells along with vectors encoding the actin-like protein BAF53 (2.5 μg, upper panel) or ER (2.5 μg, lower panel). Cell lysates were prepared and immunoprecipitated (IP) with antibodies against HA or ER. The precipitated fractions were analyzed by immunoblotting (W) with anti-Flag antibody. (B) Coactivator activity of wild-type and mutant Fli-I Gel.A fragments. pSG5.Flag vectors encoding wild-type or mutant Fli-I Gel.A fragments (0.5 μg) were transfected into CV1 cells with 0.1 μg of ER expression vector and 0.25 μg of MMTV(ERE)-LUC reporter plasmid; E2 was added after transfection, and luciferase activity was determined 40 h after transfection. The data shown are representative of seven independent experiments.