Peroxiredoxin interaction with the cytoskeletal-regulatory protein CRMP2: Investigation of a putative redox relay

Abstract

Hydrogen peroxide (H2O2) acts as a signaling molecule in cells by oxidising cysteine residues in regulatory proteins such as phosphatases, kinases and transcription factors. It is unclear exactly how many of these proteins are specifically targeted by H2O2 because they appear too unreactive to be directly oxidised. One proposal is that peroxiredoxins (Prxs) initially react with H2O2 and then oxidise adjacent proteins via a thiol relay mechanism. The aim of this study was to identify constitutive interaction partners of Prx2 in Jurkat T-lymphoma cells, in which thiol protein oxidation occurs at low micromolar concentrations of H2O2. Immunoprecipitation and proximity ligation assays identified a physical interaction between collapsin response mediator protein 2 (CRMP2) and cytoplasmic Prx2. CRMP2 regulates microtubule structure during lymphocyte migration and neuronal development. Exposure of Jurkat cells to low micromolar levels of H2O2 caused rapid and reversible oxidation of CRMP2, in parallel with Prx2 oxidation, despite purified recombinant CRMP2 protein reacting slowly with H2O2 (k~1 M−1s−1). Lowering Prx expression should inhibit oxidation of proteins oxidised by a relay mechanism, however knockout of Prx2 had no effect on CRMP2 oxidation. CRMP2 also interacted with Prx1, suggesting redundancy in single knockout cells. Prx 1 and 2 double knockout Jurkat cells were not viable. An interaction between Prx2 and CRMP2 was also detected in other human and rodent cells, including primary neurons. However, low concentrations of H2O2 did not cause CRMP2 oxidation in these cells. This indicates a cell-type specific mechanism for promoting CRMP2 oxidation in Jurkat cells, with insufficient evidence to attribute oxidation to a Prx-dependent redox relay.