Figure 4.

Endogenous MDFIC expression and its interaction with GR. A, representative immunoblot from three independent experiments of COS-1 cells expressing FLAG-MDFIC or not (empty vector) using the anti-MDFIC antibody. B, representative immunoblot from two independent experiments of A549 cells using the anti-MDFIC antibody preincubated with PBS, the immunizing peptide, or a control peptide. C, A549 cells were transfected with NTC siRNA or MDFIC siRNA. The left panel shows RT-PCR analysis of MDFIC mRNA (Student's t test, mean ± S.E.; **, p < 0.01). The right panel shows representative immunoblot from three independent experiments with anti-MDFIC antibody and actin antibody. D, Endogenous GR and MDFIC associate in a complex. A549 cells were treated for 1 h with vehicle or 100 nm Dex and processed for co-immunoprecipitation (IP). Lysates were immunoprecipitated with the anti-MDFIC antibody, and recovered proteins were immunoblotted for GR and MDFIC. Nonspecific detection of the antibody light chain (Ab-LC) was observed just below the MDFIC protein. Shown are representative immunoblots from three independent experiments. con, vehicle. E, subcellular distribution of endogenous GR and MDFIC. A549 cells were treated for 1 h with vehicle or 100 nm Dex. Cytosolic and nuclear fractions were prepared, and immunoblots were performed with anti-GR, anti-MDFIC, anti-tubulin, and anti-Lamin-A antibodies. Shown are representative immunoblots from three independent experiments.