The objective of this proposal is to determine the in vivo functional role of effector proteins that interact with the B cell co-receptor CD22 by generating transgenic mice that express wild type or altered CD22 on the CD22-/- background. CD22 is a B cell specific transmembrane glycoprotein that regulates the threshold of signaling via the B cell antigen receptor (BCR). Therefore, CD22 plays an important role in regulating the balance between tolerance and immunity in the B cell. The cytoplasmic domain of CD22 contains six tyrosine residues, one or more of which are phosphorylated in response to BCR cross-linking. The resultant phosphotyrosine motifs function as docking sites for the recruitment of SH2 domain-containing effector proteins. Studies performed in this laboratory have determined that CD22 recruits three """"""""classes"""""""" of effector proteins including the inhibitory effector protein SHP-1 and the stimulatory effector proteins PLOgamma, PI 3-K, Grb2 and Syk. Additionally, studies performed in this laboratory have determined that CD22 physically interacts with AP50, the medium chain subunit of the AP-2 complex, via a tyrosine-containing motif. Thus it is likely that CD22 expression, and presumably its function, are regulated through its association with clathrin-coated pits. Although it is apparent the CD22 negatively regulates signal transduction via the BCR; questions remain regarding the functional role of stimulatory and inhibitory effector proteins that associate with it. Therefore, studies are proposed to determine the functional importance of effector proteins that are recruited to CD22.
The specific aims of the proposal include the following: 1) to reconstitute CD22-/- mice with wild type and altered forms of murine CD22 that no longer bind to selected effector proteins; 2) to utilize the CD22 transgenic mice to determine the physiologic importance of effector protein binding to the cytoplasmic domain of CD22; 3) to define the molecular and biochemical processes that regulate CD22 expression; and 4) to determine the physiologic importance of the interaction between CD22 and the AP-2 complex in vivo. The proposed studies will precisely determine the mechanism by which CD22 regulates BCR signal transduction in vivo. The results from these studies will provide information to further our understanding of the molecular processes that regulate the balance between tolerance and immunity. This information can then be used to facilitate the development of biotherapeutic agents that will modulate the immune system to control disease.
