FC-EPSILON-RI-MEDIATED RECRUITMENT OF P53 56(LYN) TO DETERGENT-RESISTANT MEMBRANE DOMAINS ACCOMPANIES CELLULAR SIGNALING/

Detergent-resistant plasma-membrane structures, such as caveolae, have been implicated in signaling, transport, and vesicle trafficking func tions, Using sucrose gradient ultracentrifugation, we have isolated lo w-density, Triton X-100-insoluble membrane domains from RBL-2H3 mucosa l mast cells that contain several markers common to caveolae, includin g a src-family tyrosine kinase, p53/56(lyn). Aggregation of Fc epsilon RI, the high-affinity IgE receptor, causes a significant increase in the amount of p53/56(lyn) associated with these low-density membrane d omains. Under our standard conditions for lysis, IgE-Fc epsilon RI fra ctionates with the majority of the solubilized proteins, whereas aggre gated receptor complexes are found at a higher density in the gradient . Stimulated translocation of p53/56(lyn) is accompanied by increased tyrosine phosphorylation of several proteins in the low-density membra ne domains as well as enhanced in vitro tyrosine kinase activity towar d these proteins and an exogenous substrate, With a lower detergent-to -cell ratio during lysis, significant Fc epsilon RI remains associated with these membrane domains, consistent with the ability to coimmunop recipitate tyrosine kinase activity with Fc epsilon RI under similar l ysis conditions [Pribluda, V, S., Pribluda, C. gr Metzger, H, (1994) P roc. Natl. Acad. Sci, USA 91, 11246-11250]. These results indicate tha t specialized membrane domains may be directly involved in the couplin g of receptor aggregation to the activation of signaling events.