The antigen receptor of B lymphocytes (BCR) plays a central role in the development of B cells and in their inactivation or activation following antigen contact. This receptor acts both as a high efficiency system for uptake of antigen and presentation to helper T cells and as a signal transducing receptor that activates intracellular protein tyrosine kinases. Both functions appear to be important for B cell activation and antibody production whereas the signaling role seems to be primarily responsible for the inactivation that contributes to immunological tolerance. It is this signaling function of the BCR that is the focus of the proposed experiments. The BCR is a complex between membrane immunoglobulin (mIg) and two other polypeptides called Ig-alpha and Ig- beta. In the first specific aim, the structural elements of the BCR involved in signaling will be characterized. We shall try to interfere with the expression of endogenous Ig-beta, so that the importance of the cytoplasmic domain of this subunit in BCR signaling can be assessed. We shall continue our characterization of the roles of the cytoplasmic domains of Ig-alpha and Ig-beta in signaling and in particular will determine the structural requirements for binding of these domains to the tyrosine kinases Lyn, Fyn and Syk. In addition, experiments will be done to determine whether any other B cell proteins bind to the tyrosine phosphorylated forms of the signaling motifs found in Ig-alpha or Ig-beta. In the second specific aim, the mechanism of protein tyrosine kinase activation by the BCR will be characterized. A cDNA encoding Syk has been isolated. It will be introduced into a non-lymphoid system to see if this will restore signaling capability to the BCR expressed in these cells following transfection. An epitope-tagged version of wild type and mutated Syk will be introduced into B cell lines to study the structural requirements for its interaction with the BCR. Efforts will also be made to examine the roles of Src-family tyrosine kinases in BCR signaling. The experiments of the third specific aim, will examine the role of Ras in BCR signaling. In the fourth specific aim, experiments are proposed to characterize the BCR-triggered events that induce growth arrest and apoptosis in B lymphoma cell lines. These experiments are designed to test current models for the mechanism by which antigen contact triggers BCR signaling and to examine how some of the consequences of this signaling are mediated.
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