Redox-regulated rotational coupling of receptor protein-tyrosine phosphatase α dimers

Abstract

Receptor protein-tyrosine phosphatase α (RPTPα) constitutively forms dimers in the membrane, and activity studies with forced dimer mutants of RPTPα revealed that rotational coupling of the dimer defines its activity. The hemagglutinin (HA) tag of wild type RPTPα and of constitutively dimeric, active RPTPα-F135C with a disulfide bond in the extracellular domain was not accessible for antibody, whereas the HA tag of constitutively dimeric, inactive RPTPα-P137C was. All three proteins were expressed on the plasma membrane to a similar extent, and the accessibility of their extracellular domains did not differ as determined by biotinylation studies. Dimerization was required for masking the HA tag, and we identified a region in the N terminus of RPTPα that was essential for the effect. Oxidative stress has been shown to induce a conformational change of the membrane distal PTP domain (RPTPα-D2). Here we report that H2O2 treatment of cells induced a change in rotational coupling in RPTPα dimers as detected using accessibility of an HA tag in the extracellular domain as a read-out. The catalytic site Cys723 in RPTPα-D2, which was required for the conformational change of RPTPα-D2 upon H2O2 treatment, was essential for the H2O2-induced increase in accessibility. These results show for the first time that a conformational change in the intracellular domain of RPTPα led to a change in conformation of the extracellular domains, indicating that RPTPs have the capacity for inside-out signaling.