The molecular mechanisms that lead to the expression of newly synthesized mediators of inflammation are the central focus of our studies. We utilize a mast cell model, RBL-2H3 cells, due to the robust production of both inflammatory and immunoregulatory cytokines and the induction of their synthesis via the well characterized high affinity receptor for IgE (FcEpsilonRI). In particular the steps involved in the signaling pathway, beginning with IgE-antigen activation of the high affinity receptor for IgE and resulting in the induction of cytokine genes, are being explored. In the past year we have focused principally on the regulation of kinases, receptor adapter molecules and transcription factors known to be critical for the induction of gene expression. Three principal areas of investigation are underway: 1) Studies on the regulation of molecules purported to link cell surface activation of receptors with induction of gene expression. 2) Exploring the regulation of receptor-associated protein kinase C (PKC) delta in response to aggregation of the high affinity receptor for IgE. 3) Development of stably transfected mast cell lines over-expressing the individual isozymes of PKC to enable studies on cellular targets of enzyme activity. Our new results show: 1) The calcium independent PKC-delta is tyrosine phosphorylated in response to receptor aggregation. This tyrosine phosphorylation of PKC delta inhibits its ability to recognize the receptor gamma chain as a substrate, but does not inactivate the enzyme since it readily phosphorylates other substrates. This result suggests a novel intracellular mechanism for regulation of substrate recognition. 2) In collaborative studies we have identified the site of phosphorylation as tyrosine 52 in the regulatory domain of PKC delta. 3) Studies on the regulation of Vav, a protein thought to play a role in MAP kinase activation and therefore gene transcription, show that this protein which we have found to associate with the FcEpsilonRI exists as a multicomponent complex in association with the adaptor molecule Grb2, the serine/threonine kinase Raf-1 and the p42 MAP Kinase. Evidence is provided of a link between receptor and MAP kinase activation.
Showing the most recent 10 out of 49 publications
