Figure 2.

Immunoblot Analysis of Granulosa Cells Infected with Either the PKA-CQR or PKA-Cα Mutant Lentiviral Vectors

Granulosa cells were infected with either the PKA-CQR or PKA-Cα mutant lentiviral vectors (5 MOI) as described in Fig. 1. The next morning, unattached cells and free lentivirus were removed and replaced with M199. After 24 h, medium was removed and replaced with M199 containing 30 ng/ml testosterone with or without FSH (100 ng/ml). Cells infected with lentiviral vectors were cultured in M199 plus 30 mg/ml testosterone. Forty-eight hours later, cells were collected and processed for Western immunoblotting with anti-PKA-C followed by anti-GAPDH to verify equivalent loading of samples as described in Materials and Methods. Results are representative of three separate groups of granulosa cells.