The signals regulating human B cell function have not been thoroughly explored. We have focused on the mechanisms by which signals delivered through the B cell surface molecule, CD40, regulate human B cell function. These signals have been shown to be essential for germinal center formation, immunoglobulin heavy chain switching, and development of B cell memory, but the exact details of the signaling cascades involved have not been delineated. To address these questions, in vitro model systems using both B cell lines and primary B cells have been developed. The objectives of the research was to determine the signaling and adapter molecules involved in CD40 induced activation of the transcription factors, NKkappaB and AP-1. Using a transient transfection system and the B cell line, Ramos, the pathways coupling CD40 engagement and activation of NKkappa have been delineated. These involve both TRAF dependent and TRAF independent pathways and exhibit remarkable redundancy. Gene regulation by microarray analysis and proteomics have been confirmed. Experiments employing multi-parameter flow cytometry have confirmed the notable complexity of the signaling pathway resulting in activation of NFKappaB resulting from occupancy of CD40. Despite the complexity, the demonstration of points at which this signaling cascade can be inhibited suggest novel means to regulate B cell function therapeutically. In summary, the results of the experiments have delineated the mechanisms by which engagement of CD40 can activate NFkappaB. Further delineation of this pathway should provide unique targets that could be exploited to regulate B cell activity.
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