Distinct functions of the FcεR1 γ and β subunits in the control of FcεR1-mediated tyrosine kinase activation and signaling responses in RBL- 2H3 mast cells

Abstract

In RBL-2H3 rat tumor mast cells, cross-linking the high affinity IgE receptor, FcεR1, activates the protein-tyrosine kinases Lyn and Syk and initiates a series of responses including protein-tyrosine phosphorylation, inositol 1,4,5-trisphosphate synthesis, Ca2+ mobilization, secretion, membrane ruffling, and actin plaque assembly. The development of chimeric receptors containing cytoplasmic domains of individual subunits of the heterotrimeric (αβγ2) FcεR1 has simplified analyses of early signaling events in RBL-2H3 cells. Here, RBL-2H3 cells were transfected with cDNAs encoding the extra-cellular and transmembrane domains of the interleukin-2 receptor a subunit (the Tac antigen) joined to the C-terminal cytoplasmic domains of the FcεR1 γ and β subunits (TTγ and TTβ). Both sequences contain tyrosine activation motifs implicated in antigen receptor signal transduction. TTγ and TTβ are expressed independently of the native FcεR1, as demonstrated by the ability of Tac cross-linking agents to trigger the clustering and internalization through coated pits of both chimeric receptors without co-clustering the FcεR1. A full range of signaling activities is induced by TTγ cross-linking; the TTγ-induced responses are slower and, except for Lyn activation, smaller than the FcεR1-induced responses. In striking contrast, TTβ cross-linking elicits no tyrosine phosphorylation or signaling responses, it impairs basal activities measured in secretion and anti-PY (anti-phosphotyrosine antibody) immune complex kinase assays, and it antagonizes FcεR1-induced Lyn and Syk activation, protein-tyrosine phosphorylation, and signaling responses. We hypothesize that the isolated β subunit binds a specific kinase or coupling protein(s) required for signaling activity, sequestering it from the signal-transducing γ subunit. Binding the same kinase or coupling protein to the β subunit of the intact FcεR1 may serve instead to present it to the adjacent γ subunit, resulting in enhanced kinase activation and signaling responses.