Activation of GSK-3 kinase leads to a phosphorylation induced acidic shift of hNaa10p. Cells were in vivo stimulated with Wortmannin 1 μM for 8 hours, cells which remained untreated served as control. Furthermore, Wortmannin treated cells were divided in to two aliquots, one was treated with 20 U/ml CIAP enzyme, while the other remained untreated and served as negative control for this experiment. 50 μg of resuspended proteins were analyzed by 2D-SDS PAGE using 13 cm IPG pH 4–7 for IEF, and hNaa10p was detected by immunoblotting using anti-hNaa10p. Molecular size markers are indicated to the left. (A) control, unstimulated cells, and (B) cells were stimulated with 1 μM/8 hours Wortmannin. (C) Cells were stimulated with 1 μM/8 hours Wortmannin in vivo (control for D), and (D) cells were stimulated with 1 μM/8 hours Wortmannin in vivo, and the purified proteins from these cells were treated with CIAP in vitro.