Characterization of an IgE Receptor Isolated from Cultured B-Type Lymphoblastoid Cells

Abstract

For the purpose of characterizing an IgE receptor found on a subpopulation of B lymphocytes, cultured Wil-2WT cells, which bind IgE, were labeled either biosynthetically with ¹⁴C-leucine or enzymatically with ¹²⁵I by the lactoperoxidase method. The cells were lysed with NP-40, and the cleared supernatant was passed over IgE Sepharose-4B immunoadsorbent columns. The material eluted from the column was analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Two radioactively labeled proteins having extrapolated m.w. of 86,000 and 47,000 were consistently found in eluates from the IgE columns. A 23,000 dalton band, which may have been a degradation product of the other bands, was often also observed. Reduction of the proteins did not change their m.w., suggesting that they represented single polypeptide chains. Because of their lability, the proteins could only be recovered when three enzyme inhibitors were added to the cell lysates. The eluate from the IgE immunoadsorbent column, incorporated into complete Freund's adjuvant, was injected into a goat. The resulting antiserum, after absorption with cultured lymphoid cells that do not bind IgE, specifically inhibited IgE rosette formation on Wil-2WT cells as well as on other lines of cultured lymphoblastoid cells, chronic lymphatic leukemia, and tonsil and spleen lymphocytes that bind IgE. After absorption with IgE, the antiserum did not cause histamine release from normal or allergic donors' basophils. When the F(ab′)₂ fragments of the antiserum coupled to Sepharose-4B were used to isolate proteins from radiolabeled Wil-2WT lysates, both the reduced and unreduced isolated material again showed three components of 86,000, 47,000 and 23,000 m.w. These proteins were not found in lysates of Raji cells, which do not bind IgE. The results suggest that these three proteins are derived from the IgE receptor either as subunits or degradation products and that the IgE Fc receptor on lymphoid cells is antigenically different from that on basophils and mast cells.