The objectives of this research are to understand the relationship between structure and function of class II major histocompatibility complex (MHC) molecules and how they regulate the immune response in mouse and man, and to determine genetic susceptibility to a wide variety of autoimmune and related diseases. There are now cDNA and/or genomic clones for murine A-alpha, A-beta, E-alpha, and E-beta; for HLA-DR-alpha, DR-beta, and DC-beta; and for genomic sequences in a protochordate tunicate Botryllus, which are homologous to murine and human class I and class II MHC cDNA sequences. These clones will be used: (1) for synthesis, isolation, characterization, and sequencing of HLA-DR-beta, DC-beta cDNA clones for the known DR haplotypes and specifically for HLA-DR and DCl-5 and for variants subtypes of HLA-DR2,3, and 4, DR-beta, and DC-beta coding sequences. Allelic sequence polymorphism will guide synthesis of oligonucleutide allele-specific probes for typing the known DR and DC alleles and subtypes of these alleles. Allelic sequence polymorphism and allele-specific probes will be used to analyze more precisely the relationship between class II MHC type and susceptibility to disease in man; (2) to characterize and sequence genomic and cDNA clones in class I and class II MHC-like genes and gene products in the protochordate tunicate Botryllus (sp. Monterey). These sequences will be used to generate synthetic peptides and antibodies for use in analyzing expression and function of these gene products with the aim of understanding their function in an organism lacking a vertebrate immune response but expressing MHC-related gene products; (3) to synthesize peptides for allele-specific sequences of murine and human class II MHC molecules for use as immunogens to elicit chain-specific and epitope-specific antisera and monoclonal antibodies for use in functional and biochemical analysis of expression and function. Synthetic peptides homologous to similar genes in Botryllus will also be synthesized and utilized in a similar manner; and (4) to devise strategies for obtaining high-level expression of cDNA sequences encoding murine or human alpha and beta class II MHC molecules. The plan is to devise cloning strategies to achieve high-level expression of protein products of parts or all of particular cDNA sequences.
The aim i s to develop the methodology for producing sufficient amounts for biochemical characterization and collaboration with laboratories expert in X-ray crystallography. (CS)
