The Journal of Immunology, Vol 143, Issue 8 2617-2625, Copyright © 1989 by American Association of Immunologists
Studies of protein kinase C in the rat basophilic leukemia (RBL-2H3) cell reveal that antigen-induced signals are not mimicked by the actions of phorbol myristate acetate and Ca2+ ionophore
JR Cunha-Melo, HM Gonzaga, H Ali, FL Huang, KP Huang and MA Beaven
Laboratory of Chemical Pharmacology, National Heart, Lung, and Blood Institute, Bethesda, MD 20892.
Exogenous activators of protein kinase C such as PMA in combination with a
Ca2+ ionophore (A23187), cause secretion in rat basophilic (RBL- 2H3)
cells,but they do so through stimulatory signals that are not the same as
those generated by Ag or oligomers of IgE. On the one hand, the synergy
between PMA and A23187 and the suppression of Ag-mediated signals
(hydrolysis of inositol phospholipids and rise in concentration of
cytosolic Ca2+) by PMA were totally dependent on protein kinase C. The loss
of synergistic and inhibitory actions of PMA, for example, correlated with
the loss of protein kinase C (as determined by immunoblotting techniques)
when cells were continuously exposed to PMA. Furthermore, the
permeabilization of RBL-2H3 cells resulted in the loss of both protein
kinase C and the inhibitory action of PMA, but both were retained if cells
were exposed to PMA before permeabilization Ag- induced secretion, on the
other hand, was not as dependent on the presence of protein kinase C. The
potent inhibitor of this enzyme, staurosporine, which blocked completely
the secretory response to the combination of PMA and A23187, did not
inhibit Ag-induced secretion except at concentrations (greater than 10 nM)
that inhibited Ag- stimulated hydrolysis of inositol phospholipids as well.
Also RBL-2H3 cells still showed some secretory-response (approximately 25%
of normal) to Ag when cells were depleted (greater than 98%) of protein
kinase C by prolonged treatment with PMA. Previous studies have indicated
that the secretory response to PMA and A23187 is much lower than that
elicited by Ag when the concentrations of stimulants were matched to give
the same increase in concentrations of cytosolic Ca2+.
