Figure 5.
Tyrosine phosphorylation of FcγRIIA after crosslinking FcγR. (A) J2/3 (top) or J2/3-CD7 (bottom) cells were incubated with mAb IV.3 (anti-FcγRII) or with mAbs IV.3 and 3G8 (anti-FcγRIII) for 30 min on ice and then warmed 10 min to 37°C. (B) J2/3 cells were incubated with various combinations of mAbs specific for FcγRII, FcγRIII, CD48, or CD59. In both panels, crosslinking F(ab′)2 fragments of goat anti–mouse antibodies were added for various amounts of time. At each time point, an aliquot was removed, lysed, and FcγRIIA immunoprecipitated. Proteins were separated by SDS-PAGE, and blots were probed with anti-phosphotyrosine. Cocrosslinking of GPI- but not transmembrane-anchored FcγRIIIB diminishes tyrosine phosphorylation of FcγRIIA. Blots shown are representative of at least five experiments.