title: Heavy chain variable region, light chain variable region, and heavy chain CDR3 influences on the mono- and polyreactivity and on the affinity of human monoclonal rheumatoid factors.
abstract: Monoreactive high affinity pathologic autoantibodies were supposed previously to derive through somatic mutation from polyreactive low affinity autoantibodies that are encoded by a small set of unmutated V region genes in fetal and neonatal B cells. However, recent data exploring the physiologically expressed Ab repertoire and the importance of the stochastically generated heavy chain CDR3 (H-CDR3) in autoreactivity suggest that this scheme is incomplete. Here we analyzed via gene-swapping experiments and site-directed mutagenesis the relative contributions of the mutations in the light chain variable region (VL) and the heavy chain variable region (VH) domains and of the H-CDR3 in the autoreactivity of two IgM rheumatoid factors (RF), one a polyreactive low affinity Ab, the other a monoreactive high affinity Ab. These two RFs derived from the same V kappa III (humkv325) and VH1 (51p1) genes, but differed from each other by a few mutations and by the structure of the H-CDR3. The analysis of the reactivity patterns of different combinations of wild-type and in vitro engineered hybrid gene products clearly demonstrates the main influence of the H-CDR3 in the autoAb activity profiles. The results directly demonstrate the previously proposed hypothesis, namely, that the H-CDR3 plays a critical role in distinguishing poly- from monospecific RF. However, the data also indicate that self polyreactivity is a very fragile property and is dependent upon the primary structure of the VH segment.
authors.affiliation: Laboratory of Immunopathology, University Hospitals of Strasbourg, France.
authors.forename: R
authors.initials: R
authors.lastname: Crouzier
authors.forename: T
authors.initials: T
authors.lastname: Martin
authors.forename: J L
authors.initials: JL
authors.lastname: Pasquali
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D001667:Binding, Competitive / Q000276:immunology; D003429:Cross Reactions / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D015321:Gene Rearrangement / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000032:analysis / Q000235:genetics / Q000276:immunology*; D007147:Immunoglobulin Light Chains / Q000032:analysis / Q000235:genetics / Q000276:immunology*; D007135:Immunoglobulin Variable Region / Q000032:analysis / Q000235:genetics / Q000276:immunology*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed / Q000235:genetics; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000276:immunology; D012217:Rheumatoid Factor / Q000276:immunology*; D014162:Transfection / Q000235:genetics
chemical_list: D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D007135:Immunoglobulin Variable Region; D011994:Recombinant Proteins; D012217:Rheumatoid Factor