Fig. 7.

GR and FOXO1 associate directly with the MuRF1 promoter in C₂C₁₂ myotubes. A: C₂C₁₂ myotubes were treated with 1 μM DEX for 0 or 60 min (1 10-cm plate/time point), and cross-linked chromatin was immunoprecipitated with normal rabbit IgG, an anti-FOXO1 antibody, or one of two anti-GR antibodies (P20 or M20). After reversal of cross-links, immunoprecipitated MuRF1 promoter fragments were detected by PCR using primers flanking the predicted GRE and FOXO sites in the mouse MuRF1 promoter, followed by agarose gel electrophoresis. PCRs of a no-template control (NTC) and input DNAs are shown at left. B and C: C₂C₁₂ myotubes were treated with vehicle (open bars), 1 μM DEX, 20 ng/ml R3-IGF-I, or 1 μM DEX + 20 ng/ml R3-IGF-I for 15 min (black bars) or 30 min (gray bars) before harvest. Four plates for each treatment were processed in parallel as in A. Values (fold enrichment) are expressed as the mean MuRF1 promoter copies immunoprecipitated with either the anti-GR antibody (B) or anti-FOXO1 antibody (C) divided by that immunoprecipitated by a nonspecific antibody (anti-HA), as determined by quantitative PCR using the same primer pairs as in A. Error bars reflect SE.