Systemic lupus erythematosus (SLE) is the prototypic immune complex disease. The disease manifestations are due to the development of autoantibodies to nuclear antigens (ANA) including chromatin, DNA, histones and extractable nuclear antigens. These autoantibodies interact with endogenous nuclear material leading to immune complex formation and tissue damage. There is increasing evidence that the development of ANA is due to immunization with nuclear antigens released by cells following death by apoptosis or necrosis. The clearance, processing and presentation of these autoantigens must therefore play an important role in the generation of ANA. The proposed work is based on the findings that the acute phase serum protein, C-reactive protein (CRP), binds specifically to two of the major autoantigens recognized by autoantibodies from lupus patients, chromatin and Sm/RNP. CRP may be involved in their clearance since the functional activities of CRP include ligand recognition, complement activation and enhancement of phagocytosis. Serum levels of CRP increase up to 1000 fold during the acute phase response, but CRP levels remain low in SLE patients. A receptor for CRP (CRP-R) on phagocytic cells has been demonstrated, but it has not been completely characterized or cloned. The first major goal of this proposal is to characterize and clone the CRP-R. The CRP-R will be isolated by affinity chromatography of K-562 cell extracts. Microsequencing of receptor bands will be used to design primers and probes for screening a K-562 cDNA library. Monoclonal and polyclonal antibodies to the CRP-R will be prepared. Previous studies demonstrated CRP binding to the high affinity receptor for IgG on mononuclear cells as well as to the CRP-R. The interaction of CRP with FcgRI will be examined by quantitative binding studies using recombinant FcgRI expressed in COS cells. The sequence YLGGP in CRP, which is homologous to the LLGGP sequence in IgG previously demonstrated to be essential for FcR binding, will be mutated to YEGGP. Additional CRP mutants will be constructed, expressed in baculovirus and used in receptor binding studies to CRP-R and FcgRI as well as signal transduction studies in mononuclear cell lines. Functional studies will delineate the ability of CRP to stimulate signal transduction and opsonic activities through each of the two receptors.
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