Restriction in V kappa gene use and antigen selection in anti-myeloperoxidase response in mice.
Anti-neutrophil cytoplasmic Abs, directed primarily toward myeloperoxidase (MPO) and proteinase 3, are detected in the majority of patients with distinct forms of small vessel vasculitides and pauci-immune necrotizing glomerulonephritis. However, the origin of these autoantibodies remains unknown. We studied the V region gene use in murine anti-MPO Abs derived from Spontaneous Crescentic Glomerulonephritis/Kinjoh mice. A total of 13 anti-MPO-producing hybridomas were generated from four unimmunized mice. Ten of the 13 hybridomas (corresponding to 3 of 4 clones) expressed Vkappa1C but differed in their use of VH genes. The remaining three hybridomas expressed a Vkappa5 gene. Anti-MPO hybridomas from individual mice were derived from single clones as deduced by sequence similarity and splice-site identity. We found a statistically significant bias of amino acid replacement mutations to the complementarity-determining regions (CDR) in the Vkappa1C-expressing hybridomas. Intriguingly, all 10 Vkappa1C hybridomas share a lysine to glutamate mutation in the CDR1. To determine the effects of somatic V gene mutations on binding to MPO, we generated an anti-MPO Ab with an unmutated Vkappa1C L chain and compared its ability to bind MPO with its mutated counterpart. The mutated hybridoma-derived Ab has a 4.75-fold higher avidity for MPO than the unmutated Ab. These results suggest that: 1) the L chain plays a dominant role in determining Ab specificity to MPO, 2) the anti-MPO Ab response is oligoclonal, consistent with Ag selection, and 3) MPO is a driving Ag in the murine anti-MPO Ab response.
Jethwa
H S
HS
Department of Pathology and Laboratory Medicine, University of North Carolina, Chapel Hill 27599, USA.
Clarke
S H
SH
Itoh-Lindstrom
Y
Y
Falk
R J
RJ
Jennette
J C
JC
Nachman
P H
PH
D000595:Amino Acid Sequence; D000818:Animals; D001323:Autoantibodies / Q000096:biosynthesis* / Q000235:genetics; D001324:Autoantigens / Q000276:immunology* / Q000378:metabolism; D001666:Binding Sites, Antibody / Q000235:genetics; D015321:Gene Rearrangement / Q000235:genetics*; D005803:Genes, Immunoglobulin; D018095:Germ-Line Mutation / Q000276:immunology; D006801:Humans; D006825:Hybridomas / Q000276:immunology / Q000378:metabolism; D007135:Immunoglobulin Variable Region / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D007145:Immunoglobulin kappa-Chains / Q000096:biosynthesis / Q000235:genetics*; D051379:Mice; D008815:Mice, Inbred Strains; D008969:Molecular Sequence Data; D009195:Peroxidase / Q000276:immunology* / Q000378:metabolism
D001323:Autoantibodies; D001324:Autoantigens; D007135:Immunoglobulin Variable Region; D007145:Immunoglobulin kappa-Chains; D009195:Peroxidase