The helper T cell recognition of protein antigens requires that the antigen be processed and presented by an Ia expressing antigen presenting cell (APC). This involves the internalization of the antigen into an acidic intracellular vesicle where the antigen is denatured, often involving proteolysis, and the resulting processed antigen transported to and held on the APC surface where it is recognized by the specific T cell along with Ia. While the requirement for APC function in helper T cell activation is well documented, the molecular mechanisms underlying the phenomenon remain to be elucidated. This proposal represents an approach to further our understanding of these processes, taking advantage of recently developed native antigen- and peptide antigen- antibody conjugates which allow antigens to be targeted to APC surfaces. Such conjugates synthesized using the well characterized soluble globular protein antigen, cytochrome c, and a variety of antibodies specific for APC surface structures will provide a means of introducing appropriately radiolabeled native antigens into the APC in a directed fashion allowing an elucidation of the intracellular processing pathway. Several key questions will be address which include: Into which intracellular compartment is antigen transported? Where does the initial proteolytic cleavage event occur? What are the proteolytic products of that cleavage? What is the content of such vesicles with regard to proteolytic enzymes, Ia, and other potential accessory molecules? Can antigen enter the processing pathway by binding to any APC surface structure? Can intracellular cytoplasmically synthesized proteins enter the processing pathway? Peptides-antibody conjugates will be used to target the processed form of the antigen to the APC surface to investigate the mechanism by which peptides become associated with Ia and the T cell receptor. Both native antigen-and peptide- antibody conjugates will be tested for their efficacy as immunogens in vivo and the mechanisms underlying increased immunogenicity investigated. New information gained from the proposed studies will be applied toward the design of immunogens and therapeutic immunoconjugates for use in vivo.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI027957-02
Application #
3142269
Study Section
Experimental Immunology Study Section (EI)
Project Start
1990-01-01
Project End
1994-12-31
Budget Start
1991-01-01
Budget End
1991-12-31
Support Year
2
Fiscal Year
1991
Total Cost
Indirect Cost
Name
Northwestern University at Chicago
Department
Type
Schools of Arts and Sciences
DUNS #
City
Evanston
State
IL
Country
United States
Zip Code
60201
Cheng, P C; Brown, B K; Song, W et al. (2001) Translocation of the B cell antigen receptor into lipid rafts reveals a novel step in signaling. J Immunol 166:3693-701
Sproul, T W; Malapati, S; Kim, J et al. (2000) Cutting edge: B cell antigen receptor signaling occurs outside lipid rafts in immature B cells. J Immunol 165:6020-3
Wagle, N M; Kim, J H; Pierce, S K (1999) CD19 regulates B cell antigen receptor-mediated MHC class II antigen processing. Vaccine 18:376-86
Cheng, P C; Dykstra, M L; Mitchell, R N et al. (1999) A role for lipid rafts in B cell antigen receptor signaling and antigen targeting. J Exp Med 190:1549-60
Cheng, P C; Steele, C R; Gu, L et al. (1999) MHC class II antigen processing in B cells: accelerated intracellular targeting of antigens. J Immunol 162:7171-80
Wagle, N M; Faassen, A E; Kim, J H et al. (1999) Regulation of B cell receptor-mediated MHC class II antigen processing by FcgammaRIIB1. J Immunol 162:2732-40
Schafer, P H; Malapati, S; Hanfelt, K K et al. (1998) The assembly and stability of MHC class II-(alpha beta)2 superdimers. J Immunol 161:2307-16
Wagle, N M; Kim, J H; Pierce, S K (1998) Signaling through the B cell antigen receptor regulates discrete steps in the antigen processing pathway. Cell Immunol 184:1-11
Song, W; Wagle, N M; Banh, T et al. (1997) Wortmannin, a phosphatidylinositol 3-kinase inhibitor, blocks the assembly of peptide-MHC class II complexes. Int Immunol 9:1709-22
Schafer, P H; Green, J M; Malapati, S et al. (1996) HLA-DM is present in one-fifth the amount of HLA-DR in the class II peptide-loading compartment where it associates with leupeptin-induced peptide (LIP)-HLA-DR complexes. J Immunol 157:5487-95

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