Figure 5

Metformin disrupts the MID1-α4/PP2A complex and releases associated AR mRNA. DuCaP or VCaP prostate cancer cells were treated with 5 mM of metformin or vehicle control for 24 h. Afterwards the MID1-α4/PP2A complex was immunoprecipitated using an α4 specific antibody. Normal rabbit IgG was used as negative control. Complex-associated RNA was isolated and transcribed to cDNA. Using PCR and real-time PCR amplification of an AR cDNA fragment containing the CAG-repeat region (A) or a fragment of the AR hormone-binding region (B), respectively, were amplified. The agarose gel image (A) shows a representative PCR result of 3 independent experiments with DuCaP cells, the histogram (B) shows the relative real-time PCR AR fragment levels standardized to the input amount and normalized to the control for 3 independent experiments for DuCaP and VCaP cells. NeCo, negative control; Inpt, input. PCR primer and fragment information is provided in the supplementary data. Statistical significant differences are indicated as *, p < 0.05; **, p <0.01 and ***, p < 0.001.