Hydrogen peroxide-dependent oxidation of ERK2 within its D-recruitment site alters its substrate selection

Extracellular signal-regulated kinases 1 and 2 (ERK1/2) are dysregulated in many pervasive diseases. Recently, we discovered that ERK1/2 is oxidized by signal-generated hydrogen peroxide in various cell types. Since the putative sites of oxidation lie within or near ERK1/2’s ligand-binding surfaces, we investigated how oxidation of ERK2 regulates interactions with the model substrates Sub-D and Sub-F. These studies revealed that ERK2 undergoes sulfenylation at C159 on its D-recruitment site surface and that this modification modulates ERK2 activity differentially between substrates. Integrated biochemical, computational, and mutational analyses suggest a plausible mechanism for peroxide-dependent changes in ERK2-substrate interactions. Interestingly, oxidation decreased ERK2’s affinity for some D-site ligands while increasing its affinity for others. Finally, oxidation by signal-generated peroxide enhanced ERK1/2’s ability to phosphorylate ribosomal S6 kinase A1 (RSK1) in HeLa cells. Together, these studies lay the foundation for examining crosstalk between redox- and phosphorylation-dependent signaling at the level of kinase-substrate selection. [Display omitted] •ERK2 is sulfenylated by H2O2 on Cys159 within its DRS ligand binding domain•Oxidation of ERK2 differentially alters its activity toward model ERK2 substrates•DRS ligands have alternate preferences for oxidized versus reduced ERK2•Signal-generated H2O2 promotes ERK2-mediated phosphorylation of RSK1 Biological sciences; Biochemistry; Molecular biology; Cell biology