FIG 7.
Hyperammonemia regulates β-catenin via IKKβ. (A) [3H]phenylalanine incorporation in untreated myotubes and those treated for 24 h with AmAc and depleted of IKKβ or IKKα showing a partial reversal of AmAc-induced protein synthesis with silencing of IKKβ. (B) Immunoprecipitate of β-catenin probed for IKKβ and immunoprecipitate of IKKβ probed for β-catenin in untreated (UnT) murine C2C12 myotubes or those treated with 10 mM AmAc for 24 h. (C) Immunoprecipitate of IKKβ probed for β-catenin and immunoprecipitate of β-catenin probed for IKKβ in untreated HEK cells or those treated with 10 mM AmAc for 24 h. (D) Representative immunoblotting and densitometry of total β-catenin levels in C2C12 myotubes stably transfected with β-catenin mutants that are resistant to GSK3β phosphorylation (S33Y, S45Y, and N-terminal deletion [ΔN-ter] constructs). The β-catenin N-terminal deletion construct has a lower molecular weight. (E) In vitro kinase assay with purified β-catenin and either IKKβ or GSK3β (positive control) and negative controls. The blots were probed with phospho-β-catenin (Ser33/37/41) antibody. The purity of reagents was tested by immunoblotting. All cellular experiments and in vitro kinase assays were performed in at least 3 biological replicates, all blots were normalized to β-actin, and data are expressed as means ± SD. *, P < 0.05; **, P < 0.01; ***, P < 0.001; NS, not significant (versus mock-transfected untreated control myotubes).