Insufficient Phosphorylation Prevents FcγRIIB from Recruiting the SH2 Domain-containing Protein-tyrosine Phosphatase SHP-1

Abstract

FcγRIIB are IgG receptors that inhibit immunoreceptor tyrosine-based activation motif (ITAM)-dependent cell activation. Inhibition depends on an immunoreceptor tyrosine-based inhibition motif (ITIM) that is phosphorylated upon FcγRIIB coaggregation with ITAM-bearing receptors and recruits SH2 domain-containing phosphatases. Agarose bead-coated phosphorylated ITIM peptides (pITIMs) bind in vitro the single-SH2 inositol 5-phosphatases (SHIP1 and SHIP2) and the two-SH2 protein tyrosine phosphatases (SHP-1 and SHP-2). Phosphorylated FcγRIIB, however, recruit selectively SHIP1/2 in vivo. We aimed here at explaining this discordance. We found that beads coated with low amounts of pITIM bound in vitro SHIP1, but not SHP-1, i.e. behaved as phosphorylated FcγRIIB in vivo. The reason is that SHP-1 requires its two SH2 domains to bind on adjacent pITIMs. Consequently, the binding of SHP-1, but not of SHIP1, increased with pITIM density on beads. When trying to increase FcγRIIB phosphorylation in B cells and mast cells, we found that concentrations of ligands optimal for FcγRIIB phosphorylation failed to induce SHP-1 recruitment. SHP-1 was, however, recruited by FcγRIIB when hyperphosphorylated following cell treatment with pervanadate. Our data suggest that FcγRIIB phosphorylation may not be sufficient in vivo to enable the recruitment of SHP-1 but that (pathological?) conditions that would hyperphosphorylate FcγRIIB might enable SHP-1 recruitment.