Session Type: Abstract Submissions (ACR)
Background/Purpose: We have recently identified a variant in intron 1 of complement receptor 2 (CR2/CD21) that is associated with decreased risk of lupus (rs1876453; Pmeta=4.2×10-4, OR=0.85). Its effect was strongest in subjects with anti-dsDNA antibodies (Pmeta=7.6×10-7, OR=0.71), suggesting a preferential association with this B cell-driven endophenotype. Peripheral blood B cells from healthy subjects with the rs1876453 minor allele had increased levels of complement receptor 1 (CR1/CD35) resulting in an altered CR1:CR2 ratio on the cell surface. CR1 binds the C3b and iC3b fragments of C3 and is a required cofactor for the degradation of C3b to the CR2-specific ligand C3dg. While crosslinking of CR2 and the B cell antigen receptor is known to lower the activation threshold, the role of CR1 during complement-dependent B cell activation remains unclear. We hypothesize that increased CR1 levels associated with the protective allele facilitate complement processing and modify B cell activation.
Methods: A novel CR1 ligand (biot-C3b) was developed by biotinylating C3b in the thioester domain, the site of antigen attachment. The biotin orientation enables multimers of biot-C3b to be formed that mimic the natural C3b-coated immune complexes that are targets for CR1-mediated processing. Multimeric and monomeric CR1 ligands were generated by incubating biot-C3b with and without streptavidin, respectively. For analysis of CR1-mediated processing, ligand binding was evaluated by flow cytometry using primary B cells and cofactor activity was determined by incubating CR1 (soluble or cell bound) with biot-C3b and factor I followed by SDS-PAGE.
Results: Multimers of biot-C3b were processed by CR1 more efficiently than monomers as demonstrated by increased B cell binding and more complete degradation to C3dg. In solution, higher CR1 concentrations resulted in more rapid conversion of biot-C3b to its degradation products. On primary B cells, elevated levels of CR1 associated with the protective allele enhanced biot-C3b processing with increased binding capacity (p<0.05) and hastened degradation kinetics.
Conclusion: Here, we show that increases in CR1 levels, both in solution and on primary cells, enhance the processing of biot-C3b multimers. These data suggest that the elevated B cell CR1 levels associated with the protective minor allele at rs1876453 will result in increased generation of C3dg-coated complexes that bind CR2. Further characterization of the downstream effects of these augmented signals through CR1 and CR2 are likely to provide mechanistic insights into the protective effect of rs1876453 as well as enhance the understanding of how the complement system modulates B cell activation.
B. M. Giles,
S. A. Boackle,
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/alterations-in-b-cell-complement-processing-related-to-a-lupus-associated-variant-in-complement-receptor-2/