FcɛRIγ associates with CD4/PIR-A3 via Arg⁶³² in 293T cells. 293T cells were transfected with 500 ng of pcDNA3, CD4/PIR-A3, or CD4/PIR-A3^R632L cDNA in the absence or presence of 500 ng of FcɛRIγ DNA. (A) A tyrosine phosphoprotein is associated with CD4/PIR-A3 after pervanadate stimulation. The transfected cells were stimulated with pervanadate and lysed in 1% Brij-96 buffer, and whole cell lysates were immunoprecipitated with anti-CD4. Immune complexes were eluted in nonreducing Laemmli sample buffer and separated by 8% SDS-PAGE followed by immunoblotting with antiphosphotyrosine. (B) FcɛRIγ associates with CD4/PIR-A3 via Arg⁶³² in 293T cells. The transfected cells were lysed and processed as above, using reducing Laemmli sample buffer followed by separation with 4% to 20% SDS-PAGE and immunoblotting with antimurine FcɛRIγ.