Several common human disease states, including ankylosing spondylitis, post-infectious reactive arthritis, and acute anterior uveitis, are genetically associated with the serologic marker HLA- B27. Persuasive evidence has accumulated that the class 1 major histocompatibility (MHC) antigen identified as HLA-B27 on cell surfaces is directly involved in disease pathogenesis. The mechanism of this involvement is unknown, but is presumably related to the unique antigenic structure of the HLA-B27 molecule. The overall goal of the proposed experiments is to enlarge substantially the existing knowledge regarding the antigenic structure of the HLA-B27 molecule. This will be accomplished through four specific aims: 1) A panel of mutant B27 genes will be produced by oligonucleotide-mediated site-directed mutagenesis; these mutant genes will encode B27 molecules with amino acid substitutions at sites predicted to be important in the alloantigenic determinants of B27. These molecules will then be expressed in cells in vitro. 2) Transgenic mice expressing either the wild-type HLA-B27 gene, the mutant HLAB27 genes, or the related HLA-B7 gene will be created by microinjection of fertilized ova. 3) Monoclonal antibodies with restricted specificity for HLA-B27 will be generated from trangenic mice expressing either the B7 gene or mutant B27 genes, immunized with cells from the wild-type B27 transgenic mice. 4) These antibodies, along with other monoclonal and polyclonal anti-B27 antibodies, will be tested for binding to the B27 mutant molecules, and the resulting patterns of reactivity will be used to generate a computer-assisted map of antigenic determirants on the native B27 molecule, based on the recently published crystallographic map of HLA-A2. It is anticipated that this project will provide sufficient resourses and information for the rational design of successful experimental models of the pathogenesis of B27-related disease.
Showing the most recent 10 out of 32 publications
