Structure and expression of Fcγ receptors on mouse suppressor T cell hybridomas

Antigen-specific and idiotype-specifric mouse suppressor T cell hybridomas were analyzed for the presence and specificity of Fcγ receptors (FcγR) by EA rosetting and by flow microfluorometry with the use of monoclonal antibodies. We found that four hybridomas expressed FcγR specific for IgG1 and IgG2b, one of which became FcγR^- during prolonged culture. Four other hybridomas and the fusion parent, BW5147, consistently lacked FcγR. The ¹²⁵I-labeled FcγR were isolated from surface radioiodinated hybridoma cells solubilized with 1% Nonidet P-40, were purified by using single or repetitive chromatography on mouse IgG-Sepharose columns, and were analyzed by SDS-PAGE. An ¹²⁵I-labeled 56,000 to 61,000 M(r) macromolecule was isolated from each of the FcγR⁺ hybridomas, but from none of the FcγR^- hybridomas nor from BW5147 cells. This macromolecule rebound to insolubilized mouse IgG1, IgG2b, and human Fc fragments, but not to insolubilized mouse IgG2a, IgG3, or IgA or human F(ab')₂ fragments, consistent with the specificity observed for FcγR on intact hybridoma cells. The mouse suppressor T cells FcγR differs in size and specificity from mouse B cell FcγR. A 70,000 M(r) protein expressed on all hybridomas and on BW5147 cells was radiolabeled and, despite preclearing with ovalbumin-Sepharose, bound to the mouse IgG-Sepharose columns, presumably due to mouse antibodies to gp-70. This macromolecule was completely and specifically removed by using goat antiserum to gp-70.