The molecular basis of processing and presenting peptide antigens by MHC class I and class II proteins to T cells is now well understood. A third pathway of presentation of lipid antigens by CD1 proteins to T Cells has recently been described. The non-MHC encoded CD1 bears structural homology to both MHC Class I and Class II. T cell recognition of CD1 restricted lipid antigens functionally parallels that of T cell recognition of Class Il restricted peptides in many ways: 1) T cells show fine specificity for closely related antigens 2) Antigen is recognized only in the context of the appropriate """"""""restricting"""""""" CD1 isotype 3) A chloroquine sensitiVe processing step is required for recognition. CD1 restriction of alpha-beta CD8+, alpha-beta """"""""double negative"""""""" and gamma-delta T cell populations has now been demonstrated. The CD1 restricted antigens are nonpeptide lipid and glycolipid antigens including mycobacterial cell wall derived mycolic acid and lipoarabinomannan (LAM). The study of molecular models of a new biochemical class of T cell antigens broadly supports clinical research of autoimmune and infectious diseases. The candidate will define the molecular events underlying lipid antigen presentation and recognition. Heterogeneous preparations of mycolic acid will be purified into homogenous species and tested in T cell proliferation assays to define the contributions of specific chemical structures (R groups, chain length) to antigenicity. The complete chemical structure of a mycolic acid species will be determined with gas chromatography and mass spectroscopy (GC-MS). The candidate will radiolabel the lipid antigens and demonstrate that they bind to CD1 by immunoprecipitating CD1/lipid antigen complexes. The structure of the processed, CD1 bound antigen will be determined with conventional GC-MS and with Fourier Transform Ion Cyclotron Resonance Mass Spectroscopy (FTICR-MS). The candidate will identify candidate subcellular compartments of the macrophage for the loading of lipid antigen onto CD1 by defining compartments of colocalization using immunofluorescence and electron microscopy. Using cellular fractionation, cellular subcompartments will be directly examined for the presence of CD1/LAM complexes. The candidate will clone and transfect the TCR from LAM and mycolic acid specific T cell clones into the TCR- cell line, Jurkat. The structural elements of the TCR with functional importance for antigen recognition will be defined by TCR alpha or beta chain substitutions and CDR3 point mutations. The relevance of lipid T cell antigens outside of mycobacterial immunity will be explored by making T cell lines specific for Lipopolysaccharide and lipoteichoic acid, glycolipids of the gram negative and gram positive bacterial cell walls that have chemical and biological similarities with LAM. These studies are included as part of a training program in basic immunology for the candidate for this Mentored Clinical Scientist Award. The program will build on the candidate's previous experience, teaching laboratory techniques in molecular biology, cell biology and cellular immunology. Concurrently, it will provide strong theoretical background in basic immunology. With this training the candidate will develop the skills necessary to establish himself as an independent investigator in immunology. At the end of this training program, the applicant intends to obtain a faculty position and pursue an academic career in basic research.
