FIGURE 5.

WNK1 and Akt1 are involved in SGK1 activation by H₂O₂ and IGF1. HeLa cells were cotransfected with plasmid encoding FLAG-ΔSGK1 and WNK1 (Wi), Akt1 (Ai), or scrambled (Ci) small interfering RNA (siRNA) oligonucleotides. Cells were untreated (–) or treated (+) with 2 mm H₂O₂ for 25 min or with 50 ng/ml IGF1 for 30 min before harvest. Lysates were probed with anti-FLAG, anti-WNK1, anti-Akt1, anti-phospho-ERK1/2, and either anti-OSR1 (A) or anti-ERK1/2 (B) antibodies. Immunoprecipitated ΔSGK1 was assayed with GST-Nedd4-2 as substrate. A, WNK1 knockdown; B, Akt1 knockdown (n = 3 in A and B). Ctrl, control.