Structural basis for the interaction of lyn kinase with the unphosphorylated high affinity receptor for IgE Fc(RI)

Studies from our laboratory and others suggest that mast ceil responses initiated by aggregation of FccRI result from the transphosphorylation of the receptor by constitutively bound I.yn kinase. We showed (J.Biol.Chem.272:24072(1997)) that this constitutive interaction involves the unique domain of Lyn kinase by transfecting constructs of Lyn into CHO-FceRI transfectants and monitoring their effect on aggregation-induced phosphorylatiori of receptor tyrosines (PY). The current studies probed for the corresponding site of interaction on FcfRI. Prior studies by ourselves and others using either in vitro binding studies or the yeast two-hybrid system had implicated the ('-terminal rytoplasmic domain of the receptor's 3 chain (c). In order to examine (his interaction in more detail in vivo, a construct containing the extra cellular and transmembrane portion of the IL-2 receptor a chain (TAC) fused to, if (TTi?) was stably transfected into the CHO FctRI transfectants. Dramatic inhibition ( 70-90%) of Q and 7 PY was observed upon antigen-induced aggregation of the FctRI compared to cells transacted with empty vector. Since the transfectants express several FccRI for each molecule of TT.5, the TT5e construct may have a higher affinity for or be more accessible to Lyn kinase than native J,.. TTJ from either resting or activated cells were not tyrosine phosphorylated, consistent with a constitutive interaction between c and the unique domain of Lyn. Unlike a previous report on RBL TT.3 transfectants, aggregation nf ITJ was not required and failed tn enliance the inhibition in nur ~t tidic.i.