MHC class II molecules are loaded with their peptide cargo in endosomal compartments. This process is catalyzed by the class ll-like molecule, HLA-DM (DM; H2-M in mice). HLA-DO (DO; H2-O in mice), an MHC-encoded, class ll-like molecule, associates with DM both during and after transport to endosomal compartments. Intriguingly, unlike all other components of the class II processing pathway, DO expression is restricted to B cells and thymic epithelial cells. It is now clear that DO modifies the peptide loading activity of DM. Less clear is the net effect of DO; DO can promote or inhibit class II peptide loading depending on the experimental system. The restricted expression of DO/H2-O to B cells has led us to hypothesize that H2-O exerts its affects predominantly on antigens (Ags) internalized via the B cell receptor (BCR). Our preliminary data shows that BCR ligation causes rapid dissociation of H2-O from H2-M and, also a dramatic compartmentalization of free H2-M with the internalized BCR. Focusing the critical components of Ag presentation within one compartment clearly suggests a mechanism by which H2-O substantially enhances class II presentation only after the BCR engagement of the BCR with Ag. Such an """"""""Ag focusing"""""""" mechanism would appear to be critical for T-dependent B cell immune responses, particularly for B cells expressing low affinity BCR. These concepts will be directly examined in this application.
In aim 1 we will 1) confirm and extend our studies examining H2-O dissociation from H2-M in BCR-ligated cells; 2) identify the subcellular compartments to which H2-O and H2-M traffic; and 3) examine the impact that H2-O dissociation has on class II presentation.
In aim 2, we will determine the role H2-O plays in T cell-dependent immune response in vivo. The effect of H2-O expression on T cell-dependent, antigen specific antibody responses and germinal center formation will be examined. We will also study the impact of H2-O on somatic hypermutation, affinity maturation and B cell positive selection. The proposed studies will elucidate the role of H2-O in the class II antigen-processing pathway in vivo, during immune responses. These studies are relevant for tumor immunity, autoimmunity and vaccine development.
