STUDIES OF HLA CLASS II GENES

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Studies are proposed to obtain further information about sequences of expressed HLA class II DR and DQ genes that are responsible for the expression of determinant recognized by T lymphocytes in allo- or restricted recognition. Homozygous typing cells (HTCs), from consanguineous matings wherever possible, will be studied. DR4+ and DR2+ HTCs will be the primary focus of the proposed experiments for several reasons, including the fact that several "Dw subtypes" of each of these serologically defined specificities have been defined. Individuals who are DR4+ can be divided into a number of groups based on the Dw specificities that they express as defined with HTCs, i.e. specificities recognized by T lymphocytes; these "subtype" specificities include Dw4, Dw10, Dw13, Dw14, and Dw15. Likewise, DR2+ individuals can be divided into those expressing Dw2, Dw12, and LD-MN2. We consider it a special strength of this application that the same HTCs proposed herein for study by DNA techniques have been, and are continuing to be, studied in our laboratories including studies of the DR and DQ proteins, functional characterization of responding T lymphocytes, and definition of determinants recognized in allo- and restricted recognition. This should allow correlations of structure as determined by protocols in the present application with function and thus allow a better understanding of the molecular basis of cellular interactions. A number of techniques, including in vivo recombination, intra-exonic shuffling and site directed mutagenesis, are proposed to pinpoint precisely which sequences are involved in expression of determinants recognized by T lymphocytes. Normal and mutagenized genes will be studied by transfection. In addition, peptides which include sequences of interest will be evaluated in an antigen presenting cell system. Attempts will be made to generate monoclonal antibodies/antisera to determinants of interest. We believe these studies will contribute to diagnosis by providing the background information needed to study this same gene family in "disease haplotypes".