Impaired secretion and increased insolubilization of IgE‐receptor complexes in mycophenolic acid‐treated (guanine nucleotide‐depleted) RBL‐2H3 mast cells

In RBL‐2H3 rat leukemic mast cells, cross‐linking anti‐DNP IgE‐receptor complexes with multivalent antigen (DNP‐BSA) activates a signal transduction pathway leading to Ca 2+ influx and secretion. Cross‐linking IgE‐receptor complexes also stimulates a pathway that inactivates (desensitizes) receptors;this pathway becomes important at high concentrations of cross‐linking antigen. Recent evidence that antigen‐induced secretion is impaired by mycophenolic acid (MPA), an inhibitor of guanine nucleotide synthesis de novo, has implicated a GTP‐binding protein (G protein) in the signaling pathway. Other recent studies have indicated that the conversion of cross‐linked receptors to a detergent‐insoluble (cytoskeleton‐associated) form at high antigen concentrations is correlated with the loss of signaling activity. Here we show that secretion elicited by an optimal concentration of antigen (0.05 μg/ml DNP‐BSA) is only inhibited by about 25% in guanine nucleotide‐depleted cells, whereas secretion elicited by 5 μg/ml DNP‐BSA, a concentration in the range that causes the high‐dose inhibition of secretion, is inhibited by more than 60%. We also show that IgE‐receptor complexes are insolubilized in response to 5 but not 0.05 μg/ml DNP‐BSA in both control and guanine nucleotide‐depleted cells. Importantly, the extent of insol‐ubilization elicited by 5 μg/ml DNP‐BSA is increased by more than 60% in the guanine nucleotide‐depleted samples. These results raise the possibility that guanine nucleotide depletion reduces the secretory response to high antigen concentrations in two ways: by inhibiting the G protein‐coupled signaling pathway and by increasing the availability of receptors to the pathway leading to receptor insolubilization and inactivation.