MOLECULAR MECHANISMS MEDIATING CD22 ACCESSORY FUNCTION

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B cell activation is dependent on ligand binding and cross-liking of the antigen receptor resulting in activation of multiple protein tyrosine kinases. Tyrosine phosphorylation of antigen receptor-associated proteins is critical for entry into the cell cycle and proliferative expansion of the activated B cell. CD22 is a B lineage-restricted transmembrane glycoprotein that is rapidly phosphorylated on tyrosine in response to antigen receptor cross-linking. Recent studies have demonstrated that CD22 functions as an adhesion molecule and an accessory protein that regulates signal transduction via the B cell antigen receptor. Simultaneous cross- linking of CD22 and membrane immunoglobulin significantly decreases the threshold of activation required for entry of the B cell cycle. Moreover, the lack of CD22 expression has been shown to alter signal transduction processes initiated by ligand binding to the antigen receptor. Thus, it is apparent that membrane immunoglobulin and CD22 are both important for B cell activation by virtue of the fact that they act in concert to regulate the generation of signals that lead to increased gene transcription within the cell. We propose to examine the molecular mechanisms that regulate the accessory function of CD22 in the B cell. Studies will be performed to identify the regions of CD22 that are important for its interaction with the antigen receptor and the protein tyrosine phosphatase, CD45, both of which have been shown to regulate tyrosine phosphorylation of CD22. The specific antigen receptor-associated protein tyrosine kinase(s) that is responsible for phosphorylation of CD22 will be identified. Regulation of CD22 tyrosine phosphorylation by the relevant protein tyrosine kinase and the protein tyrosine phosphatase, CD45, will be examined in order to identify the specific target residues on CD22 that are substrates for these enzymes. Additionally, the physiological importance of CD22 phosphorylation will be examined with regard to the binding of SH2-containing proteins. Experiments will be performed to measure phosphotyrosine-dependent interactions between CD22 and SH2-containing signal transduction proteins as well as to define the specific tyrosine residues to which these proteins bind. Finally, studies will be performed to determine the physiological importance of accessory protein binding to CD22. CD22-negative cell lines will be reconstituted with mutated forms of CD22 that no longer associate with specific SH2-containing proteins. The ability of mutated CD22 molecules to reconstitute signal transduction via the antigen receptor will be assessed. These structure/function studies will contribute significantly to our understanding of the molecular basis by which CD22 regulates B cell activation. In view of the fact the CD22 appears to play an important role in regulating and/or potentiating entry of the B cell into the cell cycle, these studies may shed light on processes related to autoimmune disease and/or cancer that result in aberrant activation and proliferation of B cells.