MOLECULAR GENETIC ANALYSIS OF LYMPHOCYTE FUNCTION

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The focus of this laboratory has been the MHC class I molecule, classically considered a molecular heterodimer consisting of a 46 kd polymorphic heavy chain, exemplified by the murine H-2K, D, and L molecules (analogous to the human HLA-A, -B, and -C molecules), that is non-covalently assembled with the 12 kd monomorphic light chain, Beta2-microglobulin. Our objectives in these studies have been to examine a number of specific aspects of the biosynthesis, structure, and function of the class I molecule, including: 1) the measurement of the biochemical parameters that govern the binding of the MHC class I molecule to antigenic peptide; 2) the influence of Beta2-microglobulin and other serum factors on functional and biochemical binding of MHC class I molecules to antigenic peptides; 3) the expression, folding, assembly, structure, and stability of class I molecules produced in: a) an in vitro RNA-dependent translation system and b) an insect virus/moth ovary cell expression system; 4) the expression folding, assembly, structure, and stability of domain and sub-domain fragments of class I molecules generated by recombinant DNA methods, and expressed in tissue culture cells; 5) the function of soluble MHC class I molecules in a transgenic mouse model system; and 6) the production of single chain MHC class I molecules to examine the role of the heavy chain/Beta2-microglobulin interaction in the function and stability of these molecules and as the basis for high level production in prokaryotic systems.