Figure 7.

Modulation of MAP2D at Thr256/Thr259 through LH-Mediated Regulation of Both GSK3β and PP2A

A, PO granulosa cells were left untreated or pretreated with 50 μm myristoylated-PKI (Myr-PKI) for 60 min and then left untreated (−) or treated with 10 μm forskolin (Fsk), 500 μm of 8-CPT-cAMP, or 1 IU/ml hCG for 10 min, as indicated. Western blot results are representative of two separate experiments. Lines between lanes indicate cropped images. B, PO granulosa cells were pretreated with 12.5 μm of the PI-3 kinase inhibitor LY294002 in dimethylsulfoxide or an equivalent concentration of dimethylsulfoxide for 60 min and then left untreated (−) or treated with 1 IU/ml hCG for 10 min, as indicated. Western blot results are representative of three separate experiments. C, PO granulosa cells were first left untreated (−) or pretreated with the indicated concentrations of the GSK3 inhibitor lithium chloride (LiCl) for 50 min; cells were then left untreated (−) or pretreated with the PP2A inhibitor, okadaic acid (OA), for an additional 35 min; cells were then left untreated (−) or treated with 1 IU/ml hCG for 10 min. Similar results were obtained using okadaic acid combined with the GSK3 inhibitor AR-A014418 (not shown). DMSO, Dimethylsulfoxide; ph-, phosphorylated.