Structure-function analysis of Lyn kinase association with lipid rafts andinitiation of early signaling events after Fc epsilon receptor I aggregation

The first step in immunoreceptor signaling is represented by ligand-depende nt receptor aggregation, followed by receptor phosphorylation mediated by t yrosine kinases of the Src family. Recently, sphingolipid- and cholesterol- rich plasma membrane microdomains, called lipid rafts, have been identified and proposed to function as platforms where signal transduction molecules may interact with the aggregated immunoreceptors. Here we show that aggrega tion of the receptors with high affinity for immunoglobulin E (Fee RI) in m ast cells is accompanied by a co-redistribution of the Src family kinase Ly n. The co-redistribution requires Lyn dual fatty acylation, Src homology 2 (SH2) and/or SH3 domains, and Lyn kinase activity, in cis or in trans. Palm itoylation site-mutated Lyn, which is anchored to the plasma membrane but e xhibits reduced sublocalization into lipid rafts, initiates the tyrosine ph osphorylation of Fc epsilon RI subunits, Syk protein tyrosine kinase, and t he linker for activation of T cells, along with an increase in the concentr ation of intracellular Ca2+. However, Lyn mutated in both the palmitoylatio n and myristoylation sites does not anchor to the plasma membrane and is in capable of initiating Fc epsilon RI phosphorylation and early signaling eve nts. These data, together with our finding that a constitutively tyrosine-p hosphorylated Fc epsilon RI does not exhibit an increased association with lipid rafts, suggest that Fc epsilon RI phosphorylation and early activatio n events can be initiated outside of lipid rafts.