Fc ϵRI-mediated expression of mRNA for c-fos in rat basophilic leukemia cells does not require ongoing aggregation of the receptor

Abstract

Cross-linkage of the high-affinity receptor for IgE (Fc ϵRI) by a polyvalent ligand, leads to activation of mast cells and basophils. We have studied Fc ϵRI-mediated expression of RNA coding for the protoonogene, c-fos, in rat basophilic leukemia (RBL) cells and specifically have examined the requirements for ongoing receptor aggregation in the generation of this signal. RBL cells were sensitized with IgE specific for 2,4-dinitrophenyl (DNP) and incubated at 37°C in the presence of DNP 24BSA or BSA alone. Following activation for 0 to 30 min, the reaction was terminated. RNA was isolated and separated on denaturing gels, blotted to nylon membranes and hybridized with a 32P-labelled cDNA probe for c-fos. Messenger RNA for c-fos is detectable as early as 5–10 min following the addition of antigen and increases in a time-dependent fashion over 30 min. Unexpectedly, the addition of the hapten, 10 −4M DNP-lysine, 5 min after the addition of antigen (which causes immediate cessation of exocytosis) does not dramatically alter the amount of message detected at 30 min. This effect is present as early as 2 min after cross-linking of the receptor and occurs at various doses of the aggregating stimulus. Thus, in contrast to the case with exocytosis and other well-described intracellular events, Fc ϵRI-mediated increases in the level of mRNA for c-fos does not require ongoing aggregation of Fc ϵRI.