Fig. 2.

Akt phosphorylates PDNF in T. cruzi–infected cells and in vitro. (A) Akt-dependent phosphorylation of PDNF in infected cells. Uninfected (Sc) or T. cruzi–infected (Sc-Inf) Schwann cells were treated with vehicle (0), with the PI3K inhibitor LY294002, or with the Akt inhibitor Akti VIII for 24 hours before harvesting. Cells were lysed and lysates were incubated with antibodies against pAkt (Ser⁴⁷³) or, after immunoprecipitation with an antibody against PDNF (TCN-2), with antibodies specific for Akt-phosphorylated substrates (αP-Akt sbstr). (B) Akt phosphorylates PDNF in vitro. Bacterially expressed PDNF (bPDNF) was used as a substrate for immunopurified Akt in kinase reactions with (+) or without (−) ATP. Phosphorylation of PDNF was analyzed with an antibody specific for Akt-phosphorylated substrates. Cs, Coomassie blue–stained bPDNF. Identical results were obtained in at least three experiments.